To see the other types of publications on this topic, follow the link: CDC2 Protein.
Dissertations / Theses on the topic 'CDC2 Protein'
Create a spot-on reference in APA, MLA, Chicago, Harvard, and other styles
Consult the top 50 dissertations / theses for your research on the topic 'CDC2 Protein.'
Next to every source in the list of references, there is an 'Add to bibliography' button. Press on it, and we will generate automatically the bibliographic reference to the chosen work in the citation style you need: APA, MLA, Harvard, Chicago, Vancouver, etc.
You can also download the full text of the academic publication as pdf and read online its abstract whenever available in the metadata.
Browse dissertations / theses on a wide variety of disciplines and organise your bibliography correctly.
1
Randall, Susan. "Interactions among the mitogen-activated protein kinase cascades and the identification of a novel cdc2-related protein kinase." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1997. http://www.collectionscanada.ca/obj/s4/f2/dsk2/ftp04/mq29271.pdf.
Full text
2
Potapova, Tamara. "Exploring mechanisms that control the activity of cyclin-dependent kinase 1 during mitotic transitions in somatic cells." Oklahoma City : [s.n.], 2009.
Find full text
3
Bhaduri, Samyabrata. "Regulation of CDK1 Activity during the G1/S Transition in S. cerevisiae through Specific Cyclin-Substrate Docking: A Dissertation." eScholarship@UMMS, 2014. http://escholarship.umassmed.edu/gsbs_diss/871.
Full textAbstract:
Several cell cycle events require specific forms of the cyclin-CDK complexes. It has been known for some time that cyclins not only contribute by activating the CDK but also by choosing substrates and/or specifying the location of the CDK holoenzyme. There are several examples of B-type cyclins identifying certain peptide motifs in their specific substrates through a conserved region in their structure. Such interactions were not known for the G1 class of cyclins, which are instrumental in helping the cell decide whether or not to commit to a new cell cycle, a function that is non-redundant with B-type cylins in budding yeast. In this dissertation, I have presented evidence that some G1 cyclins in budding yeast, Cln1/2, specifically identify substrates by interacting with a leucine-proline rich sequence different from the ones used by B-type cyclins. These “LP” type docking motifs determine cyclin specificity, promote phosphorylation of suboptimal CDK sites and multi-site phosphorylation of substrates both in vivo and in vitro. Subsequently, we have discovered the substrate-binding region in Cln2 and further showed that this region is highly conserved amongst a variety of fungal G1 cyclins from budding yeasts to molds and mushrooms, thus suggesting a conserved function across fungal evolution. Interestingly, this region is close to but not same as the one implicated in B-type cyclins to binding substrates. We discovered that the main effect of obliterating this interaction is to delay cell cycle entry in budding yeast, such that cells begin DNA replication and budding only at a larger than normal cell size, possibly resulting from incomplete multi-site phosphorylation of several key substrates. The docking-deficient Cln2 was also defective in promoting polarized bud morphogenesis. Quite interestingly, we found that a CDK inhibitor, Far1, could regulate the Cln2-CDK1 activity partly by inhibiting the Cln2-substrate interaction, thus demonstrating that docking interactions can be targets of regulation. Finally, by studying many fungal cyclins exogenously expressed in budding yeast, we discovered that some have the ability to make the CDK hyper-potent, which suggests that these cyclins confer special properties to the CDK. My work provides mechanistic clues for cyclinspecific events during the cell cycle, demonstrates the usefulness of synthetic strategies in problem solving and also possibly resolves long-standing uncertainties regarding functions of some cell cycle proteins.
4
Kommajosyula, Naveen. "Regulation of DNA Replication Origins in Fission Yeast: A Dissertation." eScholarship@UMMS, 2009. https://escholarship.umassmed.edu/gsbs_diss/436.
Full textAbstract:
Cells need to complete DNA replication in a timely and error-free manner. To ensure that replication is completed efficiently and in a finite amount of time, cells regulate origin firing. To prevent any errors from being transmitted to the next generation, cells have the checkpoint mechanism.
The S-phase DNA damage slows replication to allow the cell to repair the damage. The mechanism of replication slowing by the checkpoint was not clear in fission yeast, Schizosaccharomyces pombe, at the start of my thesis. The downstream targets of the DNA damage checkpoint in fission yeast were also unclear. I worked on identifying the downstream targets for the checkpoint by studying if Cdc25, a phosphatase, is a target of the checkpoint.
Work from our lab has shown that origin firing is stochastic in fission yeast. Origins are also known to be inefficient. Inefficient origins firing stochastically would lead to large stretches of chromosome where no origins may fire randomly leading to long replication times, an issue called the random gap problem. However, cells do not take a long time to complete replication and the process of replication itself is efficient. I focused on understanding the mechanism by which cells complete replication and avoid the random gap problem by attempting to measure the firing efficiency of late origins.
Genome-wide origin studies in fission yeast have identified several hundred origins. However, the resolution of these studies can be improved upon.
I began a genome-wide origin mapping study using deep sequencing to identify origins at a greater resolution compared to the previous studies. We have extended our origin search to two other Schizosaccharomyces species- S. octosporus and S. japonicus.There have been no origin mapping studies on these fission yeasts and identifying origins in these species will advance the field of replication.
My thesis research shows that Cdc25 is not a target of the S-phase DNA damage checkpoint. I showed that DNA damage checkpoint does not target Cdc2-Y15 to slow replication. Based on my preliminary observation, origin firing might be inhibited by the DNA damage checkpoint as a way to slow replication. My efforts to measure the firing efficiency of a late replicating sequence were hindered by potentially unidentified inefficient origins firing at a low rate and replicating the region being studied. Studying the origin efficiency was maybe further complicated by neighboring origins being able to passively replicate the region. To identify origins in recently sequenced Schizosaccharomyces species, we initiated the genome-wide origin mapping. The mapping was also done on S. pombe to identify inefficient origins not mapped by other mapping studies. My work shows that deep sequencing can be used to map origins in other species and provides a powerful tool for origin studies.
5
Choi, Sung Hugh. "The Role of Dynamic Cdk1 Phosphorylation in Chromosome Segregation in Schizosaccharomyces pombe: A Dissertation." eScholarship@UMMS, 2010. https://escholarship.umassmed.edu/gsbs_diss/453.
Full textAbstract:
The proper transmission of genetic materials into progeny cells is crucial for maintenance of genetic integrity in eukaryotes and fundamental for reproduction of organisms. To achieve this goal, chromosomes must be attached to microtubules emanating from opposite poles in a bi-oriented manner at metaphase, and then should be separated equally through proper spindle elongation in anaphase. Failure to do so leads to aneuploidy, which is often associated with cancer. Despite the presence of a safety device called the spindle assembly checkpoint (SAC) to monitor chromosome bi-orientation, mammalian cells frequently possess merotelic kinetochore orientation, in which a single kinetochore binds microtubules emanating from both poles. Merotelically attached kinetochores escape from the surveillance mechanism of the SAC and when cells proceed to anaphase cause lagging chromosomes, which are a leading cause of aneuploidy in mammalian tissue cultured cells. The fission yeast monopolin complex functions in prevention of mal-orientation of kinetochores including merotelic attachments during mitosis. Despite the known importance of Cdk1 activity during mitosis, it has been unclear how oscillations in Cdk1 activity drive the dramatic changes in chromosome behavior and spindle dynamics that occur at the metaphase/anaphase transition. In two separate studies, we show how dynamic Cdk1 phosphorylation regulates chromosome segregation. First, we demonstrate that sequential phosphorylation and dephosphorylation of monopolin by Cdk1 and Cdc14 phosphatase respectively helps ensure the orderly execution of two discrete steps in mitosis, namely sister kinetochore bi-orientation at metaphase and spindle elongation in anaphase. Second, we show that elevated Cdk1 activity is crucial for correction of merotelic kinetochores produced in monopolin and heterochromatin mutants.
6
Yassenko, Marina. "Modifications post-traductionnelles de la sous-unité régulatrice RIIα de la protéine kinase dépendante de l'AMP-cyclique au cours du cycle cellulaire." Paris 11, 2000. http://www.theses.fr/2000PA11T070.
Full textAbstract:
Dans les cellules de mammifères, les effets multiples de l'adénosine monophosphate cyclique (AMPc) sont assurés par l'intermédiaire de la protéine kinase dépendante de l'AMPc (PKA). Deux isoenzymes de la PKA, la PKA de type I et la PKA de type II, diffèrent selon la nature de leur sous-unités régulatrices (RI ou RII). Les modifications post-traductim:melles de ces isoformes majeures des sous-unités régulatrices RIα et RIIα de la PKA, majoritairement présentes dans les cellules HeLa, ont été étudiées au cours du cycle cellulaire. Des techniques de synchronisation cellulaire et le marquage par photoaffinité des sous-unités régulatrices de la PKA avec le 8-N3 [32P] AMPc ont été utilisés. Les deux résultats majeurs de cette étude sont : la mise en évidence d'une double phosphorylation de RIIα sur deux sites différents (par la sous-unité catalytique de la PKA et par une des kinases mitotiques (probablement p34cdc2), dans les cellules en mitose et la modification de l'activité de liaison à l'AMPc pour les deux isoformes RI et RII au cours de la transition G1/S. Ces deux effets se traduisent : 1) par un changement d'affinité de RIIα à la transition G2/M pour les AKAPs, protéines qui assurent sa localisation subcellulaire ; 2) par la modification du statut oxyditif de la cellule. Les processus oxydatifs par les radicaux libres des sous-unités régulatrices des PKAs, déjà mentionnés dans des maladies telles que le psoriasis, modifieraient l'activité PKA au cours du cycle cellulaire. L'étude présente met en évidence une régulation fine de l'activité des PKAs en fonction des modifications post-traductionnelles de leurs sous-unités régulatrices.
7
Martino, Lisa. "Rôle et régulation de la kinase PLK-1 lors de l'entrée en mitose dans l'embryon de Caenorhabditis elegans." Thesis, Sorbonne Paris Cité, 2018. http://www.theses.fr/2018USPCC225.
Full textAbstract:
Lors de la division cellulaire, une cellule mère doit dupliquer (interphase) puis ségréger son matériel génétique de façon égale entre les deux cellules filles (mitose). Entre ces deux étapes, la cellule subit une réorganisation drastique gouvernée par l’acteur majeur Cdk1-Cycline B, conduisant à l’entrée en mitose. L’activation de cette kinase est régulée par une boucle d’auto-amplification où les premières molécules de Cdk1-Cycline B stimulent l’activation des suivantes. Il a été montré que la kinase Plk1 initie cette boucle d’auto-amplification en stimulant les activateurs et en réprimant les inhibiteurs de Cdk1-Cycline B en amont. Pour que cette kinase soit totalement active, elle doit être elle-même activée par Aurora A, en présence de son co-activateur Bora. Il est crucial de comprendre comment tous ces acteurs se coordonnent dans l’espace et dans le temps pour déclencher l’entrée en mitose car un dérèglement pourrait amener à une ségrégation de l’ADN anarchique, conduisant à la formation de tumeurs et l’apparition de cancers. Au cours de ma thèse, j’ai tout d’abord contribué à la mise en évidence d’un mécanisme conservé d’activation de Plk1 dans les cellules humaines et chez C. elegans (PLK-1), impliquant le co-activateur Bora ou SPAT-1 chez C. elegans. Nous avons montré que la phosphorylation de SPAT-1 par Cdk1-Cycline B induit son interaction avec PLK-1, ce qui promeut la phosphorylation de PLK-1 par Aurora A et donc son activation in vitro. Ce mécanisme phospho-dépendant de SPAT-1 est important in vivo pour contrôler dans le temps l’entrée en mitose. De plus, l’activation de Plk1 in vitro avec les protéines humaines suggèrent fortement une conservation du mécanisme. Nous avons ensuite montré que la phosphorylation de Bora et de SPAT-1 par Cdk1 sur les résidus S41, S112, S137 et S119, S190, T229 respectivement, est nécessaire à leur interaction avec Plk1/PLK-1, déclenchant ensuite l’activation de Plk1/PLK-1 et l’entrée en mitose. Ces résultats démontrent que Bora/SPAT-1 phosphorylée fait partie de la boucle d’auto-amplification de Cdk1-Cycline B via l’activation de Plk1, permettant à terme d’activer de façon irréversible les acteurs de l’entrée en mitose. Par la suite, je me suis focalisée sur le rôle de PLK-1 dans la rupture de l’enveloppe nucléaire en utilisant l’embryon de C. elegans comme système modèle. Après avoir démontré que PLK-1 est cruciale pour la rupture de l’enveloppe nucléaire dans les embryons, j’ai observé une localisation de PLK-1 à l’enveloppe nucléaire avant sa rupture et j’ai identifié un complexe de nucléoporines impliqué dans ce processus. En effet, NPP-1, NPP-4 et NPP-11 dont la fonction est de réguler le transport nucléo-cytoplasmique, ont également un second rôle dans le recrutement de PLK-1 aux pores nucléaires. PLK-1 interagit avec ses substrats phosphorylés par deux types de mécanismes d’amorçage Plk1-dépendant et indépendant, impliquant une autre kinase en amont comme Cdk1-Cycline B par exemple. J’ai montré que le recrutement de PLK-1 aux pores dépend des deux mécanismes, nécessitant donc une coordination entre Cdk1-Cycline B et PLK-1. Une fois que PLK-1 est au centre du pore nucléaire, elle peut alors probablement phosphoryler de nombreuses nucléoporines et participer au désassemblage des pores, conduisant à la rupture de l’enveloppe nucléaireDuring cell division, a mother cell duplicates (interphase) and then segregate its genetic material equally between the two daughter cells (mitosis). Between these two stages, the cell undergoes a drastic reorganization governed by the major actor Cdk1-Cyclin B, leading to mitotic entry. The activation of this kinase is regulated by an auto-amplification loop where the first molecules of Cdk1-Cyclin B stimulate activation of the following. Plk1 kinase has been shown to initiate this self-amplification loop by stimulating activators and repressing upstream Cdk1-Cyclin B inhibitors. For this kinase to be fully active, it must itself be activated by Aurora A, in the presence of its coactivator Bora. It is crucial to understand how all these actors coordinate in space and time to trigger mitotic entry because a disruption could lead to a segregation of anarchic DNA, leading to the formation of tumors and the appearance of cancers. During my thesis, I first contributed to demonstrate a conserved mechanism of Plk1 activation in human cells and in C. elegans (PLK-1), involving the coactivator Bora or SPAT-1 in C. elegans. We have shown that the phosphorylation of SPAT-1 by Cdk1-Cyclin B induces its interaction with PLK-1, which promotes the phosphorylation of PLK-1 by Aurora A and thus its activation in vitro. This phosphory-dependent mechanism of SPAT-1 is important in vivo for controlling the entry into mitosis over time. In addition, activation of Plk1 in vitro with human proteins strongly suggests conservation of the mechanism. We then showed that the phosphorylation of Bora and SPAT-1 by Cdk1 on residues S41, S112, S137 and S119, S190, T229 respectively, is necessary for their interaction with Plk1 / PLK-1, then triggering the activation of Plk1 / PLK-1 and mitotic entry. These results demonstrate that phosphorylated Bora / SPAT-1 is part of the self-amplification loop of Cdk1-Cyclin B via the activation of Plk1, ultimately enabling irreversible activation of the actors of mitotic entry. Subsequently, I focused on the role of PLK-1 in nuclear envelope breakdown using the C. elegans early embryo as a model system. After demonstrating that PLK-1 is crucial for the nuclear envelope breakdown in embryos, I observed a localization of PLK-1 to the nuclear envelope before its rupture and I identified a nucleoporin complex involved in this process. Indeed, NPP-1, NPP-4 and NPP-11 whose function is to regulate nucleo-cytoplasmic transport, also have a second role in the recruitment of PLK-1 to nuclear pores. PLK-1 interacts with its phosphorylated substrates by two types of Plk1-dependent and independent priming mechanisms, involving another upstream kinase such as Cdk1-Cyclin B for example. I have shown that the recruitment of PLK-1 to the pores depends on both mechanisms, thus requiring coordination between Cdk1-Cyclin B and PLK-1. Once PLK-1 is at the center of the nuclear pore, it can probably phosphorylate many nucleoporins and participate in the disassembly of pores, leading to tnuclear envelope breakdown
8
Borysov, Sergiy I. "B-Raf is an essential component of the mitotic machinery critical for activation of MAPK signaling during mitosis in Xenopus egg extracts." [Tampa, Fla] : University of South Florida, 2006. http://purl.fcla.edu/usf/dc/et/SFE0001759.
Full text
9
Anscombe, Elizabeth. "Targeting protein-protein interactions for cancer therapy." Thesis, University of Oxford, 2012. http://ora.ox.ac.uk/objects/uuid:6155f526-5e56-454c-819d-9510fb6f9e02.
Full textAbstract:
Protein-protein interactions (PPIs) are key drug targets and recent breakthroughs in this area are providing insight into the types of molecules needed to selectively and potently inhibit a target traditionally seen as untractable. The rules that have been used to design classic substratecompetitive drugs (for example Lipinski's rule of five) may not apply in this new field in the same way. Here I present work performed in three systems that are well-validated drug targets for oncogenesis: the CDK2/cyclin A complex, the PLK1 Polobox domain and MDM2. In each case the site of the protein-protein interaction is defined and understood and the rationale for pharmaceutical intervention is clear. I use these as a model system to evaluate the characteristics of drugs that target protein-protein interaction sites and present work on the development of inhibitors as potential leads for subsequent drug development. In Chapter 1 I introduce the problems, challenges and rewards of PPI drug development; in Chapter 2 I present co-crystal structures of MDM2 with isoindolinone inhibitors; in Chapter 3 I detail attempts to co-crystallise the Plk1 Polobox with inhibitors and screen potential inhibitors; in Chapter 4 I present the results of screening to identify inhibitors of Cyclin A recruitment; and in Chapter 5 I discuss other strategies for inhibition of the CDK2/cyclin A complex, including results with a covalent inhibitor. Through these projects I have been able to demonstrate the wide applicability of the PPI inhibition approach, identify key features of drugs able to inhibit PPIs and contribute to drug design in each system.
10
LEBEL-BINAY, SOPHIE. "Caracterisation fonctionnelle d'une nouvelle proteine transmembranaire : cd82." Paris 7, 1995. http://www.theses.fr/1995PA077277.
Full textAbstract:
La glycoproteine cd82 a ete identifiee et clonee dans notre laboratoire pour son expression preferentielle sur les lignees cellulaires sensibles a la lyse par les effecteurs nk/lak. Cette molecule de 267 acides amines (35 a 100 kda) est une proteine de type iii avec 4 segments transmembranaires, des extremites nh2 et cooh intracytoplasmiques et un grand domaine extracellulaire. Elle appartient a la famille des proteines tetra span transmembranaires (tst) qui comprend cd9, cd37, cd53, cd63 et cd81. Cd82. Elle est exprimee a la surface de nombreux types cellulaires. Son expression est augmentee apres activation ou differenciation des cellules mononuclees (lymphocytes, monocytes). Cd82 possede des proprietes de transduction sur la lignee monocytaire u937 (mobilisation de calcium intracellulaire). Cd82 est une molecule de costimulation de l'activation des lymphocytes t. Les lymphocytes t stimules in vitro par des acs anti-cd3 et anti-cd82 immobilises, se differencient et produisent de l'il-2 et de l'ifng
11
Pessoa-Brandão, Luis. "Genetic and molecular studies of Saccharomyces cerevisiae Cdc7-Dbf4 kinase function in DNA damage-induced mutagenesis /." Connect to full text via ProQuest. IP filtered, 2005.
Find full text
12
Sarrab, Ramadan. "Role of CD2 associated protein in podocyte differentiation." Thesis, University of Bristol, 2011. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.550317.
Full textAbstract:
The glomerular ultrafiltration barrier contains highly terminally differentiated podocytes with major processes and foot processes interlinked by ultrathin slit diaphragms. A number of molecules that are associated with nephrosis and podocyte damage have been described and these discoveries have given insight into the mechanisms that lead to podocyte injury. One of these molecules is CD2-associated protein (CD2AP), which is a crucial protein for slit- diaphragm assembly and function. In spite of the fact that CD2AP knockout causes nephrotic syndrome in mice and the heterozygous +/- mouse is prone to proteinuria, little is known about the relevance of this molecule in human renal pathology. In this thesis I studied the effect of a disease causing CD2AP mutation on the human podocyte phenotype. I identified the dramatic effects of the CD2AP mutation on the morphology of the cells and on the expression of mesenchymal, epithelial and other markers. I found that in contrast to wild type podocytes, CD2AP mutant podocytes acquired the characteristics of dedifferentiated cells. I compared the phenotypic characteristics of the CD2AP mutant podocytes with podocytes carrying mutations in the transcription factor WTI which is essential for normal nephrogenesis. Surprisingly, this study detected a similar de-differentiated phenotype in the CD2AP mutant podocytes to that seen in podocytes carrying a mutation in the WTl gene. In view of these phenotypic similarities I studied whether there is a functional link between CD2AP and WTI which confer these similarities in cellular phenotype. CD2AP is also known to bind to WTIP (WTl interacting protein), a molecule that binds WTI and can shuttle to the nucleus during podocyte injury. In this study, I found that WTIP might provide the functional link between CD2AP and WTl. In addition, in this study I have established the first conditionally immortalized human glomerular mesangial cell line with unique migratory properties, which will be an important adjunct in studies of representative glomerular cells, as well as in eo-culture studies. Overall this thesis demonstrates that there are clear implications for a novel role for CD2AP in the development and progression of glomerular disease. This thesis also discusses a novel conditionally immortalized human mesangial cell line that represents a new tool for the study of human mesangial cell biology in vitro.
13
Smith, Gregory R. "Identification and characterization of GTPase activating proteins for CDC42 /." view abstract or download file of text, 2001. http://wwwlib.umi.com/cr/uoregon/fullcit?p3024536.
Full textAbstract:
Thesis (Ph. D.)--University of Oregon, 2001.Typescript. Includes vita and abstract. Includes bibliographical references (leaves 90-98). Also available for download via the World Wide Web; free to University of Oregon users.
14
Taylor, Vanessa Claire. "Biology of the CD52 antigen, a major glycoprotein of human lymphocytes." Thesis, University of Cambridge, 1995. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.242883.
Full text
15
Hulstrand, Alissa Marie. "Characterization of a Cdc42 effector protein (Cep4l) and a novel role for Cdc42 in xenopus neurogenesis and Fgf signaling." Diss., University of Iowa, 2013. https://ir.uiowa.edu/etd/4859.
Full textAbstract:
The vegetal cortex of the Xenopus oocyte is enriched for several mRNAs critical for early embryonic developmental processes, including germ layer specification and dorsoventral axis formation. A recent microarray screen for other vegetally localized RNAs identified several hundred novel cortex-enriched transcripts, which may have undiscovered roles in early development. In order to better elucidate the functions of localized mRNAs in early development, I characterized the spatiotemporal expression patterns and developmental functions of two novel transcripts, TRIO and F-actin binding protein (triobp) and Cdc42 effector protein 4-like (cep4l).
Overexpression and loss-of-function experiments failed to identify a critical role for TrioBP in early Xenopus development. For Cep4l, I found that overexpression of Cep4l induced primary neuron formation throughout the epidermis, preferentially inducing primary sensory neurons. This increase came at the expense of neighboring non-neuronal ciliated and ion-secreting cells, suggesting a role for Cep4l in neural boundary formation. Additionally, I have shown that Cep4l binds specifically to Cdc42 through its known Cdc42/Rac-interactive binding (CRIB) domain, and that this activation was necessary for Cep4l function.
Morpholino (MO) oligonucleotide based inhibition of Cep4l protein synthesis resulted in decreased primary sensory neurogenesis. Additionally, I have shown that Cdc42 itself is required for sensory neurogenesis. Furthermore, I find that Fgf8a, an isoform of Fgf8 previously known to regulate neuronal development, but not the Fgf8b isoform, regulates the association of Cep4l and Cdc42. Importantly, I further show that Cep4l and Cdc42 are required for the ability of Fgf8a to induce sensory neurons.
Overall, this work suggests a novel role for Cep4l and Cdc42 in the regulation of primary sensory neuronal fate downstream of a unique Fgf8 signaling pathway. I propose that binding of Fgf8a to its receptors activates Cdc42 and recruits Cep4l, which could serve as a scaffold for integrating additional signaling pathways involved in controlling sensory neuron fate.
16
Watson, Joanna. "Structural and biochemical insight into the interactions of Cdc42 with TOCA1 and N-WASP." Thesis, University of Cambridge, 2017. https://www.repository.cam.ac.uk/handle/1810/268520.
Full textAbstract:
Cdc42 is a member of the Rho family of small GTPases, which, together with its homologues RhoA and Rac1, controls a multitude of cellular functions via the actin cytoskeleton. Cdc42 exerts its effects on the cytoskeleton via effector proteins of the Wiskott-Aldrich Syndrome (WASP) family and the Transducer of Cdc42-dependent Actin assembly (TOCA) family. The WASP family and their activation by Cdc42 have been thoroughly studied in vitro and are well understood. Conversely, understanding of the TOCA family remains limited by a lack of biochemical, biophysical and structural insight. An investigation of the TOCA1-Cdc42 interaction is described here, revealing a relatively low affinity interaction with a dissociation constant in the micromolar range. This is 10-100x weaker than other Rho-effector interactions and suggests that TOCA1 must first be co-localised with Cdc42 to achieve stable binding in vivo. The solution NMR structure of the Cdc42 binding HR1 domain of TOCA1 provides the first structural data on this protein and reveals some interesting structural features that may relate to binding affinity and specificity. A structural model of the Cdc42-HR1 complex provides further insight into differential specificities and affinities of GTPase-effector interactions. NMR and actin polymerisation assays provide insight into the pathway of Cdc42/TOCA1/WASP-dependent actin assembly, suggesting unidirectional displacement of TOCA1 by N-WASP. A comparison of the Cdc42- TOCA1 model with an NMR structure of Cdc42 in complex with the GTPase binding domain of WASP reveals a possible mechanism by which an ‘effector handover’ from TOCA1 to N-WASP could take place. Small GTPases such as Cdc42 are lipid modified and membrane anchored via their C- termini in vivo, so in vitro studies using truncated, unmodified GTPases are limited in their biological interpretation. This project also aimed to develop methods to study full length and membrane-anchored GTPases in vitro. Lipid modified protein was produced, which showed a weak affinity for liposomes, and so structural studies of membrane anchored protein are within reach. Further method development is now required to achieve stable membrane anchoring of lipid modified GTPases for detailed NMR studies.
17
Petersen, Birgit Otzen. "Regulation of mammalian CDC6 by CDK phosphorylation and proteasome dependent degradation." Thesis, Open University, 1999. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.298212.
Full text
18
Small, Lawrence Edward. "PAR Proteins Regulate CDC-42-Dependent Myosin Dynamics During C. elegans Zygote Polarization." The Ohio State University, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=osu1461086954.
Full text
19
Lundgren, Andreas. "The ABC of the cell cycle: roles of the mammalian Cdc25 isoforms /." Stockholm, 2006. http://diss.kib.ki.se/2006/91-7140-639-5/.
Full text
20
Ni, Tao. "Structural and functional study of MACPF/CDC superfamily proteins." Thesis, University of Oxford, 2016. https://ora.ox.ac.uk/objects/uuid:50793dea-6aa6-4922-b7d8-43c50079639b.
Full textAbstract:
The thesis mainly focuses on structural study of proteins in membrane attack complex- perforin/cholesterol-dependent cytolysins (MACPF/CDC) superfamily, in particular, Astrotactins from human and perforin-like proteins (PLPs) from Toxoplasma and Plasmodium. Both of these subfamily proteins are implicated in human diseases and structurally uncharacterised before. Astrotactins (ASTNs) have been shown to play a crucial role in enabling neural migration along glial fibres. While ASTN1 directly forms contacts between the neuron and the fibre, ASTN2 is responsible for extracting ASTN1 from contacts at the lagging edge of the cell and recycling them to the leading edge. ASTN2 is associated with endosomes, with the majority of its structure (at the C-terminus) projecting into their lumen, anchored by a pair of transmembrane â-helices. Here we present the structure of this "endodomain" region of ASTN2, and find it to consist of a unique combination of polypeptide folds comprising a membrane attack com- plex/perforin (MACPF) domain, an EGF-like domain, a previously-unobserved form of fibronectin type III (Fn(III)) domain and an annexin-like domain. Taken together, the structural characterisation provides a framework for better understanding the mechanism of the ASTNs and related proteins in neural and other forms of vertebrate development. Perforin-like proteins from Toxoplasma and Plasmodium (TgPLP1 and PPLPs) are critical for normal life cycle progression of these parasites, and knockout out of any of them results in significant defects in their life cycle, entrapping the parasites within the host cells and thus limiting their ability to egress. Here we present the crystal structures of TgPLP1 MACPF domain and C-terminal domain at 2.0 Å and 1.1 Å, respectively. We also presents the MACPF domain assembled in helical and hexameric ring form, which indicates the possibility of pore-formation by 6 subunits. This is the first structure of perforin-like protein from Apicomplexan parasites and provides a structural basis to elucidate the function of PLPs in toxoplasmosis and malaria pathogenesis. The final section is a continuation of a previous study in our lab on pleckstrin homology (PH) domain of kindlins. We determined the crystal structure of kindlin-3 PH domain and characterised its lipid and membrane binding properties. Using nanodiscs incorporated with different lipids as model membrane system to study the interaction between inositol phosphate lipids and kindlin-3 PH domain, together with molecular dynamic simulation studies (in collaboration with Mark Sansom's group, University of Oxford), we propose that a subset of PH domains is able to bind to multiple inositol phosphates simultaneously and so via an avidity effect have its interaction with target membranes strengthened.
21
Murray, Alison Jane. "Mutational studies on the dimerization and structure of the metastable protein CD2 domain 1." Thesis, University of Bristol, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.262836.
Full text
22
Ferreira, Sandra Mara 1982. "ARHGAP21 inibe a secreção de insulina estimulada por glicose através da modulação aa FAK, CDC42 E PKC'dzeta'." [s.n.], 2011. http://repositorio.unicamp.br/jspui/handle/REPOSIP/313943.
Full textAbstract:
Orientadores: Antonio Carlos Boschiero, Everardo Magalhães CarneiroDissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia
Made available in DSpace on 2018-08-18T17:06:06Z (GMT). No. of bitstreams: 1 Ferreira_SandraMara_M.pdf: 1558318 bytes, checksum: d69677a862aa6ba8e96725b7d5c76a87 (MD5) Previous issue date: 2011
Resumo: Introdução e objetivos: A ARHGAP21 é uma Rho-GAP que promove a ativação de um fator intrínseco da Rho-GTPase Cdc42 responsável pela hidrólise de GTP à GDP e inativação da atividade da proteína. ARHGAP21 se associa à PKC? em cardiomiócitos e à porção C-terminal da FAK em glioblastomas, onde inibe a ativação de Rho-GTPases, e, com isso, o rearranjo do citoesqueleto de actina. A Cdc42, FAK e PKC? estão envolvidas na secreção de insulina estimula por glicose, diferenciação e proliferação de ilhotas pancreáticas. O objetivo deste trabalho foi investigar em células beta pancreáticas: 1) a expressão e localização da ARHGAP21; 2) o efeito da glicose na expressão das proteínas PKC?, FAK e Cdc42 e sua associação à ARHGAP21 e; 3) a função da ARHGAP21 no controle da secreção de insulina estimulada por glicose. Materiais e Métodos: A expressão e localização da ARHGAP21 em células MIN6 foram avaliadas por imunoflorescência. Células MIN6 foram tratadas na presença de 5,6 ou 22 mM de glicose ou, na ausência ou presença de insulina (0,2 U/ml) por 3 dias. Após a extração protéica a expressão das proteínas ARHGAP21, PKC, FAK e Cdc42 foi avaliada por Western blot. Células MIN6 foram incubadas em solução contendo 22 mM de glicose e coletadas em diferentes tempos (0, 5, 15, e 30 min) para avaliação da interação ARHGAP21/FAK e ARHGAP21/PKC? por imunoprecipitação e, para análise do grau de fosforilação tirosina 397 e 925 da FAK e em treonina 410 da PKC?. A expressão dessas proteínas também foi avaliada em ilhotas pancreáticas de camundongos Swiss neonatos previamente tratados por dois dias com 1 nM de anti-sense anti-ARHGAP21 ou mismatch. Essas ilhotas foram incubadas com 2,8 ou 16,7 mM de glicose e a secreção de insulina avaliada. Resultados: A ARHGAP21 localiza-se no citoplasma, mais acentuadamente na região da membrana plasmática. Glicose reduziu a expressão da ARHGAP21 e PKC? e não alterou a expressão da FAK e Cdc42, enquanto a insulina não alterou a expressão de nenhuma das proteínas estudadas. A glicose também promoveu a dissociação ARHGAP21/FAK, levando ao aumento na fosforilação da FAK seguido de aumento na associação ARHGAP21/PKC? e redução na fosforilação da PKC? em Thr 410. Animais Knockdown para ARHGAP21 apresentaram menor expressão de PKC? e maior expressão de Cdc42, além de um aumento na secreção de insulina por ilhotas pancreáticas tanto em condição sub- (2,8 mM) quanto supra-estimulatória (16,7 mM) de glicose. Conclusão: ARHGAP21 modula Cdc42 e FAK, proteínas responsáveis pelo rearranjo do citoesqueleto de actina e extrusão do grânulo de insulina, reduzindo sua expressão e atividade, o que inibe a secreção de insulina. Além disso, observamos que a glicose modula as interações ARHGAP21/FAK e ARHGAP21/PKC?
Abstract: Background/Aims: ARHGAP21 is a Rho-GAP that promotes activation of the Rho-GTPase Cdc42 intrinsic factor, which is responsible for the hydrolysis of GTP to GDP and those proteins inactivation. ARHGAP21 associates with PKC? in cardiomyocytes and to the C-terminal portion of FAK in glioblastoma, inhibiting Rho-GTPases and actin cytoskeleton rearrangement. Cdc42, FAK and PKC? promote glucose-stimulated insulin secretion, differentiation and proliferation in pancreatic islets. The aim this study was to assess in pancreatic beta cells: 1) ARHGAP21 expression and localization; 2) glucose effect in PKC?, FAK e Cdc42 expression and association to ARHGAP21; 3) The role of ARHGAP21 on glucose-stimulated insulin secretion. Materials and Methods: ARHGAP21 expression and localization in MIN6 cells were evaluated by immunofluorescence. MIN6 cells were treated with glucose (5.6 mM and 22 mM) or insulin (0.2 U/ml) for 3 days. After protein extraction, the ARHGAP21, PKC?, FAK e Cdc42 expressions were evaluated by Western blot. MIN6 cells were exposed for 0, 5, 15, e 30 min to 22 mM of glucose. ARHGAP21/FAK and ARHGAP21/PKC? interactions were evaluated for immunopreciption and phosphorylation of FAK in tyrosine 397 and 925 and phosphorylation of PKC? in threonine 410. The expression of these proteins also was evaluated in pancreatic islets of neonates Swiss mice previously treated for 2 days with 1 nM of ARHGAP21 antisense or mismatch oligonucleotides. These islets were incubated with 2.8 mM or 16.7 mM of glucose and insulin secretion was evaluated. Results: ARHGAP21 is localized in the cytoplasm, mainly the plasmatic membrane. Glucose reduced ARHGAP21 and PKC? expression but not altered FAK and Cdc42 expression, while insulin had no effect whatsoever on the expression of the studied proteins. Glucose also dissociated ARHGAP21/FAK, leading to increased FAK phosphorylation, which was followed by ARHGAP21/PKC? association and reduced Thr410 PKC? phosphorylation. ARHGAP21 Knockdown mice pancreatic islets had lower PKC? and higher Cdc42 expression, and also increased insulin secretion in both sub- (2.8 mM) and supra-stimulatory (16.7 mM) of glucose conditions. Conclusions: ARHGAP21 modulates Cdc42 and FAK, proteins responsible for rearrangement of actin cytoskeleton and extrusion of insulin vesicles, and reducing their expression leads to inhibited insulin secretion. Furthermore, we observed that glucose modulates ARHGAP21/FAK and ARHGAP21/ PKC? interactions
Mestrado
Fisiologia
Mestre em Biologia Funcional e Molecular
23
Neutzner, Melanie. "Regulatoren des Zellteilungszyklus der Hefe Saccharomyces cerevisiae : die Polo-Kinase Cdc5 und der Ubiquitinierungsfaktor Hct1 /." [S.l. : s.n.], 2003. http://www.bsz-bw.de/cgi-bin/xvms.cgi?SWB10605153.
Full text
24
Primeau, Martin. "Novel mechanisms of regulation of the Cdc42 GTPase- activating protein CdGAP/ARHGAP31, a protein involved in cell migration and adhesion." Thesis, McGill University, 2011. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=96901.
Full textAbstract:
The Rho GTPases form a family of enzymes that control numerous cellular processes including cell migration and proliferation through effects on the cytoskeleton, membrane trafficking and cell adhesion. The activity of these molecular switches is modulated by GTPase-activating proteins (GAPs), a group of negative-regulators which includes Cdc42-GTPase-Activating Protein (CdGAP). This protein specifically negatively-regulates the Rho GTPases Cdc42 and Rac1. In this study, we show that CdGAP is regulated by lipid-, protein- and intramolecular-interactions. First, we demonstrate that a polybasic region (PBR) of CdGAP preceding the GAP domain and found in numerous Rho family GAPs is required for CdGAP specific association with phosphatidilinositol-3,4,5-trisphosphate (PI(3,4,5)P3). We show that the binding of PI(3,4,5)P3 is required for CdGAP-mediated GAP activity in vitro, and that an intact PBR is required for its CdGAP-mediated GAP activity in vivo. Second, we characterize the binding site for the negative-regulator of CdGAP Intersectin-1 located in the Basic-Rich (BR) domain of CdGAP. We present evidence that this interaction mediated by the SH3D domain of Intersectin requires one to three lysine residues located in the Basic-Rich (BR) domain of CdGAP. Thirdly, we show that CdGAP is negatively-regulated by its C-terminal domain. This observation is part of a study that links two human CdGAP gene mutations to a syndrome which presents a combination of aplasia cutis congenita (ACC) and terminal transverse limb defects (TTLD). In this syndrome, the deletion-mutant gene products which lack the residual amino-acid of CdGAP at its C-terminus have an increased activity compared to wild-type proteins. We show that this C-terminus can bind to the GAP domain of CdGAP, providing a model to explain how the absence of the C-terminus induces this syndrome. In summary, this work provides novel insight into understanding the mechanisms of regulation of CdGAP, a protein involved in cell migration and adhesion with unexpected roles related to human diseases.Les Rho GTPases forment une famille d'enzymes qui contrôlent de nombreux processus cellulaires, tels que la migration cellulaire et la prolifération, grâce à leurs effets sur le cytosquelette, le trafic membranaire et l'adhésion cellulaire. L'activité de ces interrupteurs moléculaires est modulée par les protéines activatrices de GTPases (GAPs), un groupe de régulateurs négatifs qui inclu CdGAP (Cdc42-GTPase activating protein). Cette protéine régule négativement les Rho GTPases Cdc42 et Rac1 de façon spécifique. Dans la présente étude, nous montrons que CdGAP est régulée par des interactions lipidiques, protéiques et intramoléculaire. Premièrement, nous démontrons qu'une région polybasique (PBR), précédant le domaine GAP et retrouvée dans plusieurs GAP de la famille Rho, est requise pour l'association spécifique de CdGAP avec le phosphatidilinositol-3,4,5-trisphosphate (PI(3,4,5)P3). Nos résultats suggèrent que l'activation des GAP requiert la liaison du PI(3,4,5)P3 à CdGAP dans un contexte in vitro et un PBR intact pour que CdGAP provoque ses effets GAP-dépendants dans un contexte in vivo. Deuxièmement, nous caractérisons le site de liaison du régulateur négatif de CdGAP Intersectin-1. Ce site est localisé dans le domaine riche en résidus basiques (BR) de CdGAP. Nous suggérons que cette interaction, médiée par le domaine SH3D d'Intersectin, requiert de un à trois résidus lysine dans le domaine BR de CdGAP. Troisièmement, nous montrons que CdGAP est régulé de manière négative par son propre domaine C-terminal. Cette observation fait partie d'une étude qui associe deux mutations humaines du gène CdGAP à un syndrôme présentant une combinaison d'aplasie cutis congenita (ACC) et de malformation des doigts et des orteils (TTLD). Les gènes mutants produisent des protéines tronquées qui ont une activité GAP supérieure à la protéine de type sauvage. Nous montrons que ce C-terminal peut lier le domaine GAP de CdGAP, supportant un modèle expliquant comment l'absence du C-terminal induit ce syndrome. En bref, ce travail présente un nouvel aperçu des mécanismes de régulation de CdGAP, une protéine impliquée dans la migration cellulaire et dans l'adhésion des cellules en plus d'être directement impliquée dans une maladie humaine.
25
Rowan, Wendy Caroline. "Characterisation of function and regulation of the CD52 antigen on T and B lymphocytes." Thesis, Imperial College London, 1999. http://hdl.handle.net/10044/1/7566.
Full text
26
Dart, Anna Elizabeth. "The role of the adaptor protein Nck in phagocytosis and other Cdc42-dependent signalling pathways." Thesis, Imperial College London, 2010. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.520954.
Full text
27
Head, Jared G. "Structure and function of the #beta#-sheet proteins, titin and CD2." Thesis, University of Bristol, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.265387.
Full text
28
Trautmann, Susanne. "Functions of the Cdc14-Family Phosphatase Clp1p in the Cell Cycle Regulation of Schizosaccharomyces pombe: A Dissertation." eScholarship@UMMS, 2005. https://escholarship.umassmed.edu/gsbs_diss/10.
Full textAbstract:
In order to generate healthy daughter cells, nuclear division and cytokinesis need to be coordinated. Premature division of the cytoplasm in the absence of chromosome segregation or nuclear proliferation without cytokinesis might lead to aneuploidy and cancer.
The cyclin dependent kinases, CDKs, are a main regulator of the cell cycle. Timely increase and decrease in their activity is required for cell cycle progression. To enter mitosis, mitotic CDK activity needs to rise. CDK activity stays elevated until chromosome segregation is completed and exit from mitosis requires decrease in CDK activity.
Observations in several experimental systems suggest that coordination of cytokinesis with the nuclear cycle is regulated through CDK activity. Prolonged high CDK activity, as it occurs when chromosome segregation is delayed, was found to oppose cytokinesis. Prevention of cytokinesis through high CDK activity may therefore provide a mechanism to prevent precocious cell division in the absence of chromosome segregation. To prevent polyploidy when cell division is delayed, progression through the next nuclear cycle should be inhibited until cytokinesis is completed, presumably by the inhibition of CDK activity.
In the fission yeast Schizosaccharomyces pombe, a signaling cascade called Septation Initiation Network (SIN) is required for the coordination of cytokinesis with the nuclear cycle. The SIN is essential for cytokinesis, triggering the execution of cell division through constriction of the actomyosin ring. The activation of the SIN signaling cascade, and thus cytokinesis, is opposed by high CDK activity, preventing precocious cytokinesis.
S. pombe delay entry into the next nuclear division in response to delayed cytokinesis due to defects in the contractile ring until cytokinesis is completed thereby preventing the accumulation of multinucleate, non viable cells. This safeguard against multinucleate cells is termed the cytokinesis checkpoint. The cytokinesis checkpoint keeps CDK activity low, preventing nuclear cycle progression. The SIN is required for the cytokinesis checkpoint and therefore is a key coordinator between nuclear cycle and cytokinesis. How the SIN functions in the cytokinesis checkpoint was not known.
Cdc14-family phosphatases are highly conserved from yeast to humans, but were only characterized in Saccharomyces cerevisiae at the time this thesis was initiated. Cdc14 had been identified as the effector of a signaling cascade homologous to the SIN, called the mitotic exit network (MEN), which is required for exit from mitosis. This thesis describes the identification of the S. pombe Cdc14-like phosphatase Clp1p as a component of the cytokinesis checkpoint. Clp1p opposes CDK activity, and Clp1p and the SIN activate each other in a positive feedback loop. This maintains an active cytokinesis checkpoint and delays mitotic entry. We further found that Clp1p regulates chromosome segregation.
Concluding, this thesis describes discoveries adding to the characterization of the cytokinesis checkpoint and the function of Clp1p. While others found that Cdc14-family phosphatases, including Clp1p, have similar catalytic functions, we show that their biological function may be quite different between organisms, possibly due to different biological challenges.
29
Han, Bong Kwan. "The G1 cyclin Cln3p regulates vacuole homeostasis through phosphorylation of a scaffold protein, Bem1p, in Saccharomyces cerevisiae." Texas A&M University, 2005. http://hdl.handle.net/1969.1/4795.
Full textAbstract:
How proliferating cells maintain the copy number and overall size of their organelles is
not clear. In the budding yeast Saccharomyces cerevisiae the G1 cyclins Cln1,2,3p
control initiation of cell division by regulating the activity of the cyclin-dependent
kinase (Cdk) Cdc28p. We show that Cln3p controls vacuolar (lysosomal) biogenesis and
segregation. First, loss of Cln3p, but not Cln1p or Cln2p, resulted in vacuolar
fragmentation. Although the vacuoles of cln3ÃÂ cells were fragmented, together they
occupied a large space, which accounted for a significant fraction of the overall cell size
increase in cln3ÃÂ cells. Second, cytosol prepared from cells lacking Cln3p had reduced
vacuolar homotypic fusion activity in cell-free assays. Third, vacuolar segregation was
perturbed in cln3ÃÂ cells. Our findings reveal a novel role for a eukaryotic G1 cyclin in
cytoplasmic organelle biogenesis and segregation.
Furthermore we show that the scaffold protein Bem1p, a critical regulator of
Cdc42p activity, is a downstream effector of Cln3p/Cdc28p complex. The Cdc42p
GTPase is known to be required for vacuole fusion. Our results suggest that Ser72 on
Bem1p is phosphorylated by Cdc28p in a Cln3p-dependent manner to promote vacuole fusion. Replacing Ser72 with Asp, to mimic phosphorylation at an optimal Cdkconsensus
site located in the first SH3 domain of Bem1p, suppressed vacuolar
fragmentation in cells lacking Cln3p. Using in vivo and in vitro assays, we found that
Cln3p was unable to promote vacuole fusion in the absence of Bem1p or in the presence
of a non-phosphorylatable Bem1p-Ser72Ala mutant. Furthermore, activation of Cdc42p
also suppressed vacuolar fragmentation in the absence of Cln3p. Our results provide a
mechanism that links cyclin-dependent kinase activity with vacuole fusion through
Bem1p and the Cdc42p GTPase cycle.
30
Stromme, Adrianna. "The characterization of the cytoskeleton and associated proteins in the formation of wound-induced contractile arrays /." Thesis, McGill University, 2008. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=116078.
Full textAbstract:
The cytoskeleton is an intrinsic aspect of all cells, and is essential for many cellular events including cell motility, endocytosis, cell division and wound healing. Remodeling of the cytoskeleton in response to these cellular activities leads to significant alterations in the morphology of the cell. One such alteration is the formation of an actomyosin contractile array required for cytokinesis, wound healing and embryonic development.Cellular structure and shape depends upon tensional prestress brought about by the organization of cytoskeletal components. Using the Xenopus laevis oocyte wound healing model, it is first described how diminished cellular tension affects the balance of the Rho family of GTPases, and subsequently prevents the formation of actomyosin contractile arrays. This suggests that cellular tension in the cell is not created at the level of the cytoskeletal elements but rather via the upstream signaling molecules: RhoA and Cdc42.
The role of N-WASP (Neural-Wiscott Aldrich Syndrome Protein), a mediator of Arp2/3 based actin polymerization, is next examined for its putative role in cellular wound healing. Xenopus laevis oocytes injected with mutant N-WASP constructs reveals in vivo evidence that functional N-WASP is required for appropriate contractile array formation and wound closure.
Lastly, it is revealed that the cellular structures involved with single cell wound healing in other model systems are also important for the initial repair of severed muscle cells. Actin, non-muscle myosin-II, microtubules, sarcomeric myosin and Cdc42 are all recruited and reorganized at the edge of damaged C2C12 myotubes. This data promotes the possibility that an actomyosin array may be established in injured muscle cells as well.
31
Souza, Renan Crocci de. "Investigação de parceiros moleculares de Cdc42 em linhagens de células humanas submetidas a estresse genotóxico." Universidade de São Paulo, 2016. http://www.teses.usp.br/teses/disponiveis/46/46131/tde-15092016-090036/.
Full textAbstract:
A proteína Cdc42 (Cell Division Cycle 42) é um membro da família das Rho GTPases, sinalizadores intracelulares conhecidos pelo seu papel na regulação do citoesqueleto. Essa proteína e capaz de ciclar entre um estado ativo (ligado à GTP) e um estado inativo (ligado à GDP) e essa ativação é modulada por diversas proteínas, conhecidas como GEFs (guanine nucleotide-exchange factors), GAPs (GTPase-activating proteins) e GDIs (guanine nucleotide-dissociation inhibitors). Trabalhos recentes têm demonstrado um papel de Cdc42 na apoptose e na senescência, respostas relacionadas e comumente desencadeadas por estresse genotóxico. Neste contexto este trabalho procurou identificar interações de Cdc42 com outras proteínas, que podem ou não estar envolvidas nos mecanismos de resposta ao dano do DNA. Para isso foram utilizadas as linhagens celulares HeLa e MRC-5 submetidas a tratamento com radiação ultravioleta tipo C, a fim de provocar danos no DNA. Foram realizados dois diferentes tratamentos em cada uma das linhagens com diferentes tempos de incubação pós radiação UV, visando a busca de proteínas envolvidas em uma resposta rápida ou tardia ao dano causado. Os lisados celulares desses tratamentos foram submetidos ao pull-down com proteínas recombinantes GST, GST-Cdc42WT (Selvagem) e GST-Cdc42V12 (Mutação constitutivamente ativa). As proteínas purificadas foram digeridas e submetidas à análise por espectrometria de massa e os dados obtidos foram utilizados para a construção de redes de interação proteica. Dentre as proteínas identificadas as que despertaram maior atenção foram: Proibitina-2 (PHB2) encontrada nas amostras incubadas por 48 horas pós irradiação e Cullina-4A (CUL4A) e P53, encontradas em amostra incubada por 5 minutos pós radiação. Essas proteínas possuem papéis em apoptose e reparo de DNA e foram observadas em posições muito próximas de Cdc42 nas redes de interação, fazendo delas interessantes alvos para futuras validações de interação proteica por análises experimentais distintasThe Cdc42 protein (Cell Division Cycle 42) is a member of the Rho family of GTPases, intracellular signalling molecules well known for their role in the cytoskeleton regulation. This protein cycles between an active state (GTP-bound) and an inactive state (GDP-bound) and this regulation is modulated by proteins known as GEFs, GAPs and GDIs. Recent studies demonstrated roles for Cdc42 in apoptosis and senescence, cellular responses commonly triggered by genotoxic stress. This work sought to identify Cdc42 interactions with other proteins that possibly involved in response to DNA damage mechanisms. To reach this aims we used HeLa and MRC-5 cell lines submitted to treatments with ultraviolet C radiation to induce DNA damage. Two experimental conditions were used in each cell line with different times and doses post UV irradiation in order to search for proteins involved in either rapid or delayed response to the installed DNA damage. Cell lysates obtained from these treatments were subjected to pull-down experiments using recombinant proteins GST, GST-Cdc42-WT (Wild type) and GST-Cdc42-V12 (constitutively active mutant). Purified proteins were digested by trypsin, analyzed by mass spectrometry and th obtained data were used for the construction of protein-protein interaction (PPI) networks. Among the identified proteins those that seem more relevant to the aims of this project were: Prohibitin-2 (PHB2), found in samples incubated 48 hours post irradiation; Cullin-4A (CUL4A) and P53, found in samples incubated 5 minutes after radiation. These proteins have roles in apoptosis and DNA repair and were observed in close proximity to Cdc42 in PPI networks, making them interesting targets for future validation by different experimental approaches
32
El, Dika Mohammed. "Régulation de la phase M du cycle cellulaire par CDK1, PP2A et CDC6." Thesis, Rennes 1, 2013. http://www.theses.fr/2013REN1S068.
Full textAbstract:
L'objectif de cette thèse est de mieux comprendre la régulation de la phase M du cycle cellulaire. Nos expériences ont été effectuées dans des extraits acellulaires d’embryons de Xenopus laevis. Tout d'abord, nous montrons que le moment de l'entrée en phase M est précisément déterminé par un équilibre entre l'activité de la protéine kinase CDK1 et l’activité d’une protéine phosphatase sensible à l'acide okadaïque, PP2A. Nous montrons également le rôle de la protéine CDC6 dans la régulation de l'entrée dans la première phase M embryonnaire. En effet, CDC6 inhibe CDK1 et à travers cette action régule la dynamique de cette kinase lors de l'entrée et de la progression en phase M. Ces résultats mettent en évidence un nouveau contrôle qui précise le moment du clivage embryonnaire. Ce contrôle joue un rôle clé dans la coordination entre les mécanismes de régulation du cycle cellulaire et le programme de développement de l'embryonThe aim of this thesis is to understand better the regulation of the M-phase of the cell cycle. Experiments were done in cell-free extracts of Xenopus laevis one-cell embryos. Firstly, we show that the timing of the M-phase entry is precisely determined by a balance between the activity of CDK1 kinase and okadaic acid sensitive phosphatase, mainly PP2A. Secondly, we show the role of CDC6 protein in regulation of the entry into the first embryonic M-phase. CDC6 inhibits CDK1 and through this action regulates the dynamic of this kinase upon M-phase entry and during M-phase progression. This mechanism discovered during my PhD allows controlling precisely the timing of embryonic cleavage. This control plays a key role in coordinating the cell cycle regulating machinery and the development program of the embryo
33
Johnson, Julian A. "The Production of Designed Potential Protein Contrast Agents and their Encapsulation in Albumin Microspheres." Digital Archive @ GSU, 2008. http://digitalarchive.gsu.edu/biology_theses/20.
Full textAbstract:
Using protein design, a series of metal binding proteins have been designed, allowing the local factors that contribute to metal affinity and thermostability to be studied. Those proteins with the highest metal binding affinities had the lowest apo-form Tm and the largest ÄTm upon metal binding. In this thesis, major steps have been taken toward applying the engineered protein to MR imaging. The progress of magnetic resonance imaging is hindered by low specificity and rapid elimination of FDA-approved MRI contrast agents. The engineered protein contrast agent has been conjugated to a cancer-specific targeting peptide and encapsulated in albumin microspheres to provide tandem passive and active tumor targeting. Also, a simple, high-yield purification method has been developed.
34
Leung, Daisy W. "Biochemical and biophysical characterization of the allosteric equilibrium of the Wiskott-Aldrich Syndrome protein." Access to abstract only; dissertation is embargoed until after 12/20/2006, 2005. http://www4.utsouthwestern.edu/library/ETD/etdDetails.cfm?etdID=131.
Full text
35
Godoy, Patricia Diogo de Melo. "Estudos sobre o componente ORC1/CDC6 da maquinaria de pré-replicação dos tripanossomas." Universidade de São Paulo, 2010. http://www.teses.usp.br/teses/disponiveis/42/42135/tde-18102010-112445/.
Full textAbstract:
Em eucariotos, a origem de replicação é reconhecida por um complexo ORC, a proteína Cdc6 e outras proteínas. Nos tripanossomas, encontramos somente uma proteína similar a Orc1 e Cdc6, que chamaremos de Orc1/Cdc6. Nesta tese serão descritos os estudos realizados sobre Orc1/Cdc6 do Trypanosoma cruzi e do Trypanosoma brucei. As proteínas recombinantes dos tripanossomas apresentam atividade de ATPase e são capazes de substituir Cdc6 em ensaio de complementação em leveduras. A indução do silenciamento do gene de Orc1/Cdc6 por RNAi resulta em células anucleadas. Orc1/Cdc6 é expressa durante todo o ciclo celular das formas replicativas, permanecendo associada à cromatina. No caso do T.cruzi, Orc1/Cdc6 é expressa não só nas formas replicativas, mas também nas formas não replicativas. Nestas últimas, a proteína expressa não interage com o DNA, este resultado sugere que a ausência desta interação deve contribuir para ausência da duplicação do DNA nas formas infectivas do T. cruzi.In eukaryotes, the replication origin is recognized by a complex ORC, Cdc6 and other proteins. The trypanosomes contain only one protein, we named it Orc1/Cdc6. Here we show that the recombinant Orc1/Cdc6 from T.cruzi (TcOrc1/Cdc6) and from T.brucei (TbOrc1/Cdc6) present ATPase activity, replaced yeast Cdc6 in a phenotypic complementation assay. The induction of Orc1/Cdc6 silencing by RNA interference in T.brucei resulted in enucleated cells. Orc1/Cdc6 is expressed during the entire cell cycle and in all stages of the life cycle of trypanosomes, remaining associated with chromatin in all stages of the cell cycle. This association is different among the stages from T. cruzi life cycle. In the non replicative ones, Orc1/Cdc6 does not interact with DNA. The lack of pre-replication machinery-DNA interaction in T. cruzi non-replicative stages might contribute to the absence of DNA replication in these stages.
36
Tiedt, Barbara. "Cleavage of DNA replication proteins Mcm3 and Cdc6 and differential gene expression in B cell apoptosis." [S.l. : s.n.], 2003. http://www.diss.fu-berlin.de/2003/166/index.html.
Full text
37
Vicente-García, José Julio. "Identification of new activated Cdc42 kinase (ACK1) binding proteins and characterisation of the ACK1-STAT3 interaction." Thesis, University of Cambridge, 2009. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.611738.
Full text
38
Raman, Malavika. "Identification of intracellular signaling pathways regulated by the TAO family of mammalian STE20p kinases." Access to abstract only; dissertation is embargoed until after 5/15/2007, 2006. http://www4.utsouthwestern.edu/library/ETD/etdDetails.cfm?etdID=163.
Full text
39
Lee, Hsiau-Wei. "Determining The Site Specific Metal Binding and Structural Properties of EF-Hand Protein Using Grafting Approach." Digital Archive @ GSU, 2008. http://digitalarchive.gsu.edu/chemistry_diss/26.
Full textAbstract:
Calmodulin is an essential EF-hand protein with a helix-loop-helix calcium binding motif. Understanding Ca(II) dependent activation of calmodulin and other EF-hand proteins is limited by Ca(II)-induced conformational change, multiple and cooperative binding of Ca(II) ions, and interactions between the paired EF-hand motifs. The goal of this research project is to probe key determinants for calcium binding properties and pairing interactions at the site specific level using a grafting approach and high resolution NMR. An individual Ca(II) binding site of the EF-hand motifs of calmodulin was grafted into a non-calcium dependent protein, CD2, to bypass limitations associated with natural EF-hand proteins and peptide fragments. Using high resolution NMR, we have shown that the grafted EF-loop III of calmodulin in the host protein retains its native conformation with a strong loop and β-conformation preference. Grafted ligand residues in the engineered protein are directly involved in binding of Ca(II) and La(III). The NMR studies support our hypothesis that both ligand arrangement and dynamic properties play essential role in tuning Ca(II) binding affinities. Using pulse-field diffusion NMR and protein engineering, we further demonstrated that grafted EF- loop remains as a monomer. Although the EF-loop with flanking helices dimerizes in the presence of Ca(II). Additionally, removal of conserved hydrophobic residues at the flanking helices of the EF-hand motif leads to be monomer in the absence and presence of metal ions. Our results suggest that conserved hydrophobic residues are essential for the pair-paired interaction in the coupled EF-hand protein. We have shown that our developed grafting approach can be applied to probe intrinsic Ca(II) binding affinities of different Ca(II) binding sites.
40
Mutavchiev, Delyan Rumenov. "Regulation of fission yeast cell polarity by stress-response pathways." Thesis, University of Edinburgh, 2017. http://hdl.handle.net/1842/29006.
Full textAbstract:
Cell polarisation is a key biological process crucial for the functioning of essentially all cells. Regulation of cell polarity is achieved through various processes determined by both internal and external factors. An example of the latter is that cell polarity can be disrupted or lost as a consequence of a variety of external stresses. When facing such stresses, cells adapt to unfavourable conditions by activating a range of molecular signalling pathways, collectively termed ‘stress response’. Despite the connections between external stress and cell polarity, whether stress-response signalling regulates cell polarisation and what the molecular basis for such regulation remains an open question. The fission yeast Schizosaccharomyces pombe presents an excellent biological platform to study the complexity of cell polarity regulation on a systematic level. This study is aimed at understanding the functional relationship between stress-response signalling and maintenance of cell polarity in this model organism. The findings presented in this thesis set the basis for establishing a functional link between the activation of the S.pombe stress-response pathway and the activity of the master regulator of cell polarity- the Rho GTPase Cdc42. Here, I describe experiments that identify an active involvement of the stress-response mitogen-activated kinase (MAPK) Sty1 in the dispersal of active Cdc42 from the sites of growth. This new role for Sty1 occurs independently from its involvement in transcription regulation and other previously identified signalling pathways involving Sty1. Furthermore, I also find that Sty1’s involvement in Cdc42 regulation has direct implications for fission yeast physiology as it is essential for the maintenance of cellular quiescence upon nitrogen starvation. This thesis also focuses on identifying the targets of Sty1 orchestrating the active Cdc42 disruption. Here, I describe a candidate-based approach, where I investigate the role of proteins from the Cdc42 regulatory network during Sty1 activation. Additionally, I present a global phospho-proteomics approach to identify novel targets of Sty1 and offer preliminary findings which might explain Sty1’s involvement in Cdc42 regulation.
41
Takahashi, Satoe. "Plasma Membrane Localization of Signaling Proteins in Yeast: a Dissertation." eScholarship@UMMS, 2008. https://escholarship.umassmed.edu/gsbs_diss/364.
Full textAbstract:
In response to external stimuli, many intracellular signaling proteins undergo dynamic changes in localization to the plasma membrane. Using the Saccharomyces cerevisiaemating pathway as a model, I investigated the molecular interactions that govern plasma membrane localization of signaling proteins, and how the plasma membrane compartmentalization of a signaling complex influences the overall signaling behavior of the pathway.
Signaling proteins often consist of multiple interaction domains that collectively dictate their localization and function. Ste20 is a p21-activated kinase (PAK) that functions downstream of the Rho-type GTPase Cdc42 to activate several mitogen-activated protein (MAP) kinase pathways in budding yeast, including the mating pathway. I identified a short domain in Ste20 that directly binds to membrane lipids via electrostatic interaction. A mutation in this domain abolishes both the localization and function of Ste20. Thus, the previously known Cdc42 binding is necessary but not sufficient; instead, direct membrane binding by Ste20 is also critical. By replacing this domain with heterologous membranebinding domains, I demonstrated that phospholipid specificity is not essential in vivo. Functionally important short membrane-binding domains were also found in the Cdc42 effectors Gic1 and Gic2, indicating that generic membrane binding can work in concert with the CRIB domain to regulate activation of Cdc42 targets. These results underscore the importance of cooperation between protein-protein and protein-membrane interaction in achieving proper localization of signaling proteins at the cell cortex.
At the system level, MAP kinase cascades can be graded or switch-like. The budding yeast mating pathway exhibits a graded response to increasing levels of pheromone. Previously the scaffold protein Ste5 was hypothesized to contribute to this graded response. To test this idea, I activated the pathway in a variety of ways and measured the response at the single cell level. I found that the graded response is not perturbed by the deletion of negative regulators of the pathway whereas the response became switch-like when the pathway was activated by a crosstalk stimulus that bypasses the upstream components. Interestingly, activation of the pathway in the cytoplasm using the graded expression of MAPKKK resulted in an ultrasensitive response. In contrast, activation of the pathway at the plasma membrane using the graded expression of membranetargeted active pathway components remained graded. In these settings, the scaffold protein Ste5 increased ultrasensitivity when limited to the cytosol; however, if Ste5 was allowed to function at the plasma membrane, signaling was graded. The results suggest that, in the mating pathway, the inherently ultrasensitive MAPK cascade is converted to a graded system by the scaffoldmediated assembly of signaling complexes at the plasma membrane. Therefore, the plasma membrane localization of Ste5 helps shape the input-output properties of the mating MAPK pathway in a manner that is suitable for the biology of mating.
Taken together, this thesis underscores the importance of plasma membrane localization during mating pathway signaling in yeast. The examples described here provide further appreciation of how multiple interaction domains can function together to achieve specific targeting of the signaling proteins, as well as advances in understanding the role of scaffold proteins in modulating signaling behavior to promote graded signaling at the plasma membrane.
42
Ciolli, Mattioli Camilla. "Post-transcriptional mechanisms contributing to RNA and protein localization: study of local translation and alternative 3′UTRs in induced neurons." Doctoral thesis, Humboldt-Universität zu Berlin, 2019. http://dx.doi.org/10.18452/20702.
Full textAbstract:
Die asymmetrische Verteilung von mRNA und Proteinen innerhalb einer Zelle definiert die Polarität. Dies ermöglicht eine strikte Regulierung der Genexpression in Raum und Zeit. Ich habe in dieser Arbeit untersucht, wie das Soma und die Neuriten in induzierten Neuronen sich hinsichtlich ihres Transkriptoms und Translatoms unterscheiden. Eine räumliche ribosomale Profilanalyse ergab, dass die Hälfte des lokalen Proteoms durch die mRNA-Lokalisierung und der lokalen Translation definiert wird. Dies sind Prozesse, die durch die synergistische Aktivität von trans- und cis-agierenden Elementen durchgeführt werden. In dieser Arbeit konzentrierte ich mich auf MOV10 als trans-agierendes Element und die alternativen 3′UTRs als cis-agierende Elemente, um ihre Rolle in der Asymmetrie zu untersuchen. MOV10 ist eine RNA-Helikase, welche an vielen Aspekten des RNA-Metabolismus beteiligt ist. Mit den Methoden RIP und PAR-CLIP konnte ich zeigen, dass sowohl MOV10-Ziele als auch MOV10 selbst in den Neuriten lokalisiert sind. Aus ̈erdem ist MOV10 möglicherweise an der translationalen Repression mitinvolviert.
In der Tat konnte ich unter den MOV10-Protein-Interaktoren mehrere Proteine identifizieren, welche an der translationalen Repression beteiligt sind, wie z.Bsp. AGO2, FMR1, und TRIM71. Für die Identifizierung der cis-agierenden Elemente führte ich das "Mapping" von alternativen 3′UTRs durch. Diese Analyse zeigte mehrere Gene, die differentiell lokalisierte 3′UTR-Isoformen exprimieren. Insbesondere habe ich mich auf Cdc42 konzentriert. Ich konnte beweisen, dass die beiden Isoformen von Cdc42 auf mRNA-Ebene unterschiedlich lokalisiert sind und dass die 3′UTR der entscheidende Faktor für die mRNA- und Proteinlokalisierung ist. Darüber hinaus habe ich mehrere RBPs identifiziert, die an der Cdc42-Lokalisierung beteiligt sind. Diese Analyse zeigt, dass für die differenzierte Lokalisierung von funktional unterschiedlichen alternativen Protein-Isoformen die Verwendung von alternativen 3′UTR Isoformen als neu-entdeckter Mechanismus eine entscheidende Rolle spielt.Asymmetric distribution of mRNA and proteins inside a cell defines polarity, which allow tight regulation of gene expression in space and time. In this thesis I investigated how asymmetric distribution characterizes the somatic and neuritic compartments of in induced neurons, in terms of transcriptome and translatome. Spatial ribosome profiling analysis revealed that half of the local proteome is defined by mRNA localization and local translation. These, are processes accomplished by the synergistic activity of trans- and cis-acting elements. I focused on MOV10 as trans-acting element, and on alternative 3′UTRs as cis-elements, to investigate their role in asymmetry. MOV10 is an RNA helicase which participates to many aspects of RNA metabolism. With RIP and PAR-CLIP I showed that MOV10 targets are localized to the neurites, consistently with MOV10-neuritic localization, and that MOV10 might be involved in translational repression. Indeed, among MOV10 protein interactors, I identified several proteins involved in translational repression, i.e. AGO2, FMR1, and TRIM71. On the side of cis-elements, I performed mapping of alternative 3′UTRs. This analysis identified several genes expressing differentially localized 3′UTR isoforms. In particular, I focused on Cdc42. I showed that the two isoforms of Cdc42 are differentially localized at mRNA level, and that the 3′UTR is the driver of mRNA and protein localization. Moreover, I identified several RBPs that might be involved in Cdc42 localization. This analysis points to usage of alternative 3′UTR isoforms as a novel mechanism to provide for differential localization of functionally diverse alternative protein isoforms.
43
Searle, Jennifer. "The Role of PKA in the DNA Damage Checkpoint." University of Cincinnati / OhioLINK, 2005. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1123003066.
Full text
44
Bretou, Marine. "Regulation of the dynamics of the fusion pore : importance of the SNARE protein synaptobrevin 2 and of the Rho GTPase Cdc42." Paris 7, 2010. http://www.theses.fr/2010PA077157.
Full textAbstract:
L'exocytose nécessite la formation d'un pore de fusion. Le pore initial est étroit ; seules de petites molécules sont libérées. Quand le pore s'élargit les macromolécules sont libérées. J'ai étudié le rôle de deux protéines sur la dilatation du pore: la protéine SNARE synaptobrévine 2 (Syb2), et la Rhô GTPase Cdc42. L'assemblage des SNAREs fournirait l'énergie nécessaire à la fusion. L'insertion d'un espacer dans le domaine juxtamembranaire de Syb2 ne modifie pas la fréquence des événements d'exocytose détectés par ampérométrie à 1|jM [Ca2+], mais empêche l'apparition d'une composante de sécrétion à de plus fortes [Ca2+]. Les événements peuvent être classés en deux groupes, liés à la vitesse et au degré de dilatation des pores; l'allongement de Syb2 réduit la population de pics rapides, mais n'affecte pas celle des pics lents. Les événements lents seraient dus à un assemblage partiel des SNAREs, alors que ceux dits rapides résulteraient d'un assemblage plus serré, assurant ainsi une dilatation rapide du pore. Cdc42 contrôle la dynamique de l'actine. Diminuer son expression dans les cellules BON réduit le nombre de granules fusionnant complètement avec la membrane, mais n'affecte pas leur recrutement et leur liaison à la membrane. Réduire l'expression de Cdc42 diminue le nombre de hauts pics dus à une dilatation rapide et complète des pores, et augmente le nombre de pieds seuls, dus à des pores ne s'élargissant pas. L'augmentation de tension de la membrane corrige les effets dus à l'absence de Cdc42 ; sa diminution par dépolymérisation de l'actine imite les effets obtenus en son absence. Cdc42 contrôlerait la dilatation du pore en modulant la tension de membraneExocytosis ends with the formation of a fusion pore. The initial pore is narrow, only small molecules flow through it. The pore then enlarges, releasing larger secretory products. I studied the role of two proteins on the dilation of the pore: the SNARE protein synaptobrevin 2 (Syb2), and the Rho GTPase Cdc42. Zippering of SNAREs in opposed membranes might give energy to catalyze fusion. Inserting a linker between the SNARE core and the transmembrane domain of Syb2 did not modify the frequency of exocytotic events detected by amperometry at 1|jM free [Ca2+] but prevented the occurrence of an extra component of release at higher [Ca2+]. Analysis of these events led to their classification into two groups, due to the rate and extent of dilation of the pore; lengthening Syb2 reduced the population of fast spikes, leaving the slow one unchanged. Slow fusion events might be due to a partial zippering of the SNAREpin while fast fusion events require a tight one, i. E. A short intermembrane distance to assure rapid dilation of the pore. Cdc42 controls actin dynamics. TIRFM experiments showed that its silencing in BON cells reduced the number of granules undergoing full fusion, with little effect on their recruitment and docking at the membrane. Using amperometry, we showed that this silencing reduced the number of high spikes due to fast and complete dilation of the pore, and increased stand-alone foot signals reflecting pores failing to enlarge. Increasing membrane tension rescued the effects of silencing while decreasing it through actin depolymerization mimicked Cdc42 silencing. Cdc42 might control fusion pore dilation by modulating membrane tension
45
Legagneux, Vincent. "Deux etudes biochimiques de la cellule stressee chez les mammiferes : acceleration de l'echange du phosphate sur la proteine de choc thermique hsp90; stimulation des activites proteine-kinases de type cdc2 et rna polymerase ii ctd-kinase." Paris 7, 1990. http://www.theses.fr/1990PA077145.
Full textAbstract:
Le choc thermique, ainsi que certaines formes de stress chimique, ont des effets toxiques sur les activites enzymatiques et le metabolisme cellulaire. Pendant la phase de recuperation qui suit le choc thermique, l'expression des proteines de choc thermique (hsps) est specifiquement augmentee. Au terme de cette phase, les fonctions cellulaires sont restaurees. On pense actuellement que les hsps sont directement impliquees dans ce phenomene de recuperation cellulaire en interagissant avec les proteines denaturees qui s'accumulent dans les cellules stressees. Dans un premier temps, nous avons mis en evidence une acceleration de l'echange du phosphate sur la hsp90 dans les cellules humaines hela soumises a un choc thermique. Ce phenomene s'accompagne d'une plus grande phosphorylabilite de la hsp90 in vitro, par une enzyme qui a des caracteristiques communes avec la caseine-kinase ii. Nous pensons que cet echange du phosphate pourrait refleter une acceleration des reactions d'association/dissociation entre la hsp90 et les proteines denaturees. Nous avons d'autre part etudie le devenir des activites proteine-kinases dans les cellules stressees. Deux activites sont stimulees par un choc thermique ou un stress a l'arsenite de sodium: une kinase capable de phosphoryler le domaine c-terminal de la rna polymerase ii (ctd-kinase) et une histone h1-kinase du type cdc2. Nous avons montre que ces deux activites etaient portees par des molecules differentes. Une etude sur la transcription elevee du gene hsp86 dans les cellules de carcinome embryonnaire murin est egalement presentee
46
Puglise, Jason Matthew. "Roles of the Rac/Cdc42 effector proteins Pak and PIX in cytokinesis, ciliogenesis, and cyst formation in renal epithelial cells." University of Cincinnati / OhioLINK, 2010. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1288980975.
Full text
47
Clarkson, Nicholas. "Coordination of extracellular and intracellular interactions in immune regulation by the CD2/SLAM family of Leukocyte Surface Proteins." Thesis, University of Oxford, 2009. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.526553.
Full text
48
Cheng, Haitao. "Protein structure prediction and conformational transitions I. Improvement of protein secondary structure prediction : II. Pathways of conformational transition originating in phosphorylation : a study of CDK2 using targeted molecular dynamics and coarse grained models /." [Ames, Iowa : Iowa State University], 2009. http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&res_dat=xri:pqdiss&rft_dat=xri:pqdiss:3360333.
Full text
49
Zhou, Yubin. "Exploring the Role of Calcium Ions in Biological Systems by Computational Prediction and Protein Engineering." Digital Archive @ GSU, 2007. http://digitalarchive.gsu.edu/chemistry_diss/17.
Full textAbstract:
Ca2+, a signal for death and life, is closely involved in the regulation of numerous important cellular events. Ca2+ carries out its function through its binding to Ca2+-receptors or Ca2+-binding proteins. The EF-hand protein, with a helix-loop-helix Ca2+-binding motif, constitutes one of the largest protein families. To facilitate our understanding of the role of Ca2+ in biological systems (denoted as calciomics) using genomic information, an improved pattern search method (http://www.chemistry.gsu.edu/faculty/Yang/Calciomics.htm) for the identification of EF-hand and EF-like Ca2+-binding proteins was developed. This fast and robust method allows us to analyze putative EF-hand proteins at the genome-wide level and further visualize the evolutionary scenario of the EF-hand protein family. This prediction method further enables us to locate a putative viral EF-hand Ca2+-binding motif within the rubella virus nonstructural protease that cleaves the nonstructural protein precursor into two active replicase components. A novel grafting approach has been used to probe the metal-binding properties of this motif by engineering the predicted 12-residue Ca2+-coordinating loop into a non-Ca2+-binding scaffold protein, CD2 domain 1. Structural and conformational studies were further performed on a purified, bacterially-expressed NS protease minimal metal-binding domain spanning the Zn2+- and EF-hand Ca2+-binding motif. It was revealed that Ca2+ binding induced local conformational changes and increased thermal stability. Furthermore, functional studies were carried out using RUB infectious cDNA clone and replicon constructs. Our studies have shown that the Ca2+ binding loop played a structural role in the NS protease and was specifically required for optimal stability under physiological conditions. In addition, we have predicted and characterized a calmodulin-binding domain in the gap junction proteins connexin43 and connexin44. Peptides encompassing the CaM binding motifs were synthesized and their ability to bind CaM was determined using various biophysical approaches. Transient expression in HeLa cells of two mutant Cx43-EYFP constructs without the putative CaM-binding site eliminated the Ca2+-dependent inhibition of gap junction permeability. These results provide the first direct evidence that CaM binds to a specific region of the ubiquitous gap junction protein Cx43 and Cx44 in a Ca2+-dependent manner, providing a molecular basis for the well-characterized Ca2+-dependent inhibition of Cx43-containing gap junctions.
50
Doucet, Sandrine. "Etude fonctionnelle de RhoG, une petite GTPase apparentée a Rac et Cdc42. Localisation sub-cellulaire et caractérisation de partenaires et effecteurs." Montpellier 1, 1996. http://www.theses.fr/1996MON1T030.
Full text