Journal articles: 'G proteins ; Drosophila'

1

Wadsworth, Samuel C. "Drosophila src family proteins." Comparative Biochemistry and Physiology Part B: Comparative Biochemistry 97, no. 3 (January 1990): 403–6. http://dx.doi.org/10.1016/0305-0491(90)90135-g.

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Schmidt, C. J., S. Garen-Fazio, Y. K. Chow, and E. J. Neer. "Neuronal expression of a newly identified Drosophila melanogaster G protein alpha 0 subunit." Cell Regulation 1, no. 1 (November 1989): 125–34. http://dx.doi.org/10.1091/mbc.1.1.125.

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Guanine nucleotide-binding proteins (G proteins) mediate signals between activated cell-surface receptors and cellular effectors. A bovine G-protein alpha-subunit cDNA has been used to isolate similar sequences from Drosophila genomic and cDNA libraries. One class, which we call DG alpha 0, hybridized to position 47A on the second chromosome of Drosophila. The nucleotide sequence of the protein coding region of one cDNA has been determined, revealing an alpha subunit that is 81% identical with rat alpha 0. The cDNA hybridizes strongly to a 3.8 kb mRNA and weakly with a 5.3 kb message. Antibodies raised against a trp-E-DG alpha 0 fusion protein recognized a 39,000 Da protein in Drosophila extracts. In situ hybridization to adult Drosophila sections combined with immunohistochemical studies revealed expression throughout the optic lobes and central brain and in the thoracic and abdominal ganglia. DG alpha 0 message and protein were also detected in the antennae, oocytes, and ovarian nurse cells. The neuronal expression of this gene is similar to mammalian alpha 0, which is most abundantly expressed in the brain.

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van Lohuizen, Maarten, Marieke Tijms, Jan Willem Voncken, Armin Schumacher, Terry Magnuson, and Ellen Wientjens. "Interaction of Mouse Polycomb-Group (Pc-G) Proteins Enx1 and Enx2 with Eed: Indication for Separate Pc-G Complexes." Molecular and Cellular Biology 18, no. 6 (June 1, 1998): 3572–79. http://dx.doi.org/10.1128/mcb.18.6.3572.

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ABSTRACT The Polycomb group (Pc-G) constitutes an important, functionally conserved group of proteins, required to stably maintain inactive homeobox genes repressed during development. Drosophila extra sex combs (esc) and its mammalian homolog embryonic ectoderm development (eed) are special Pc-G members, in that they are required early during development when Pc-G repression is initiated, a process that is still poorly understood. To get insight in the molecular function of Eed, we searched for Eed-interacting proteins, using the yeast two-hybrid method. Here we describe the specific in vivo binding of Eed to Enx1 and Enx2, two mammalian homologs of the essential DrosophilaPc-G gene Enhancer-of-zeste[E(z)]. No direct biochemical interactions were found between Eed/Enx and a previously characterized mouse Pc-G protein complex, containing several mouse Pc-G proteins includingmouse polyhomeotic (Mph1). This suggests that different Pc-G complexes with distinct functions may exist. However, partial colocalization of Enx1 and Mph1 to subnuclear domains may point to more transient interactions between these complexes, in support of a bridging role for Enx1.

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Katanayeva, Natalya, Damir Kopein, Reto Portmann, Daniel Hess, and Vladimir L. Katanaev. "Competing Activities of Heterotrimeric G Proteins in Drosophila Wing Maturation." PLoS ONE 5, no. 8 (August 23, 2010): e12331. http://dx.doi.org/10.1371/journal.pone.0012331.

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Wolfgang, W. J., F. Quan, N. Thambi, and M. Forte. "Restricted spatial and temporal expression of G-protein alpha subunits during Drosophila embryogenesis." Development 113, no. 2 (October 1, 1991): 527–38. http://dx.doi.org/10.1242/dev.113.2.527.

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Of the known signal transduction mechanisms, the most evolutionarily ancient is mediated by a family of heterotrimeric guanine nucleotide binding proteins or G proteins. In simple organisms, this form of sensory transduction is used exclusively to convey signals of developmental consequence. In metazoan organisms, however, the developmental role of G-protein-coupled sensory transduction has been more difficult to elucidate because of the wide variety of signals (peptides, small molecules, odorants, hormones, etc.) that use this form of sensory transduction. We have begun to examine the role of G-protein-coupled signaling during development by investigating the expression during Drosophila embryogenesis of a limited set of G proteins. Since these proteins are a common component of all G-protein-coupled signaling systems, their developmental pattern of expression should indicate when and where programmed changes in gene activity are initiated by, or involve the participation of, G-protein-coupled signaling events. We have focused on the spatial and temporal expression pattern of three different Drosophila G-protein alpha subunits by northern blot analysis, in situ hybridization and immunocytochemistry using antibodies directed to peptides specifically found in each alpha subunit. From the spatial and temporal restriction of the expression of each protein, our results suggest that different forms of G-protein-coupled sensory transduction may mediate developmental interactions during both early and late stages of embryogenesis and may participate in a variety of specific developmental processes such as the establishment of embryonic position, the ontogeny of the nervous system and organogenesis.

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Chamberlin, H. M., and J. H. Thomas. "The bromodomain protein LIN-49 and trithorax-related protein LIN-59 affect development and gene expression in Caenorhabditis elegans." Development 127, no. 4 (February 15, 2000): 713–23. http://dx.doi.org/10.1242/dev.127.4.713.

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We have molecularly characterized the lin-49 and lin-59 genes in C. elegans, and found their products are related to Drosophila trithorax group (trx-G) proteins and other proteins implicated in chromatin remodelling. LIN-49 is structurally most similar to the human bromodomain protein BR140, and LIN-59 is most similar to the Drosophila trx-G protein ASH1. In C. elegans, lin-49 and lin-59 are required for the normal development of the mating structures of the adult male tail, for the normal morphology and function of hindgut (rectum) cells in both males and hermaphrodites and for the maintenance of structural integrity in the hindgut and egg-laying system in adults. Expression of the Hox genes egl-5 and mab-5 is reduced in lin-49 and lin-59 mutants, suggesting lin-49 and lin-59 regulate HOM-C gene expression in C. elegans as the trx-G genes do in Drosophila. lin-49 and lin-59 transgenes are expressed widely throughout C. elegans animals. Thus, in contrast to the C. elegans Polycomb group (Pc-G)-related genes mes-2 and mes-6 that function primarily in the germline, we propose lin-49 and lin-59 function in somatic development similar to the Drosophila trx-G genes.

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Kopein, Damir, and Vladimir L. Katanaev. "Drosophila GoLoco-Protein Pins Is a Target of Gαo-mediated G Protein–coupled Receptor Signaling." Molecular Biology of the Cell 20, no. 17 (September 2009): 3865–77. http://dx.doi.org/10.1091/mbc.e09-01-0021.

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G protein–coupled receptors (GPCRs) transduce their signals through trimeric G proteins, inducing guanine nucleotide exchange on their Gα-subunits; the resulting Gα-GTP transmits the signal further inside the cell. GoLoco domains present in many proteins play important roles in multiple trimeric G protein–dependent activities, physically binding Gα-subunits of the Gαi/o class. In most cases GoLoco binds exclusively to the GDP-loaded form of the Gα-subunits. Here we demonstrate that the poly-GoLoco–containing protein Pins of Drosophila can bind to both GDP- and GTP-forms of Drosophila Gαo. We identify Pins GoLoco domain 1 as necessary and sufficient for this unusual interaction with Gαo-GTP. We further pinpoint a lysine residue located centrally in this domain as necessary for the interaction. Our studies thus identify Drosophila Pins as a target of Gαo-mediated GPCR receptor signaling, e.g., in the context of the nervous system development, where Gαo acts downstream from Frizzled and redundantly with Gαi to control the asymmetry of cell divisions.

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Yamamoto, Y., F. Girard, B. Bello, M. Affolter, and W. J. Gehring. "The cramped gene of Drosophila is a member of the Polycomb-group, and interacts with mus209, the gene encoding Proliferating Cell Nuclear Antigen." Development 124, no. 17 (September 1, 1997): 3385–94. http://dx.doi.org/10.1242/dev.124.17.3385.

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We have isolated and molecularly characterized the cramped (crm) gene of Drosophila melanogaster, and show that it can be classified as a Polycomb-group (Pc-G) gene. crm mutants exhibit typical Pc-G mutant phenotypes, reminiscent of ectopic homeotic gene expression, with additional sex comb teeth found on mesothoracic and metathoracic legs, and proximodistal transformations of the tarsal segments. crm encodes an 693 amino acids protein, with no significant homology to known proteins. We used polyclonal antibodies raised against bacterially expressed truncated CRM protein to show that the crm gene product is localized to the nucleus during embryogenesis. This nuclear localization appears to be restricted to S-phase nuclei, as CRM immunostaining disappears at mitosis. We found that this cell-cycle-dependent staining pattern was identical to that of Proliferating Cell Nuclear Antigen (PCNA). Furthermore, we provide evidence for co-localization of CRM and PCNA proteins in salivary gland polytene nuclei, and for a genetic interaction between crm and mus209, the Drosophila gene encoding PCNA. Together, our data suggest that these two proteins are involved in a common regulatory pathway and highlight possible interactions between Pc-G-mediated silencing and DNA replication in Drosophila.

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9

Bunker, C. A., and R. E. Kingston. "Transcriptional repression by Drosophila and mammalian Polycomb group proteins in transfected mammalian cells." Molecular and Cellular Biology 14, no. 3 (March 1994): 1721–32. http://dx.doi.org/10.1128/mcb.14.3.1721.

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The Polycomb group (Pc-G) genes are essential for maintaining the proper spatially restricted expression pattern of the homeotic loci during Drosophila development. The Pc-G proteins appear to function at target loci to maintain a state of transcriptional repression. The murine oncogene bmi-1 has significant homology to the Pc-G gene Posterior sex combs (Psc) and a highly related gene, Suppressor two of zeste [Su(z)2]. We show here that the proteins encoded by bmi-1 and the Pc-G genes Polycomb (Pc) and Psc as well as Su(z)2 mediate repression in mammalian cells when targeted to a promoter by LexA in a cotransfection system. These fusion proteins repress activator function by as much as 30-fold, and the effect on different activation domains is distinct for each Pc-G protein. Repression is observed when the LexA fusion proteins are bound directly adjacent to activator binding sites and also when bound 1,700 bases from the promoter. These data demonstrate that the products of the Pc-G genes can significantly repress activator function on transiently introduced DNA. We suggest that this function contributes to the stable repression of targeted loci during development.

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10

Bunker, C. A., and R. E. Kingston. "Transcriptional repression by Drosophila and mammalian Polycomb group proteins in transfected mammalian cells." Molecular and Cellular Biology 14, no. 3 (March 1994): 1721–32. http://dx.doi.org/10.1128/mcb.14.3.1721-1732.1994.

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The Polycomb group (Pc-G) genes are essential for maintaining the proper spatially restricted expression pattern of the homeotic loci during Drosophila development. The Pc-G proteins appear to function at target loci to maintain a state of transcriptional repression. The murine oncogene bmi-1 has significant homology to the Pc-G gene Posterior sex combs (Psc) and a highly related gene, Suppressor two of zeste [Su(z)2]. We show here that the proteins encoded by bmi-1 and the Pc-G genes Polycomb (Pc) and Psc as well as Su(z)2 mediate repression in mammalian cells when targeted to a promoter by LexA in a cotransfection system. These fusion proteins repress activator function by as much as 30-fold, and the effect on different activation domains is distinct for each Pc-G protein. Repression is observed when the LexA fusion proteins are bound directly adjacent to activator binding sites and also when bound 1,700 bases from the promoter. These data demonstrate that the products of the Pc-G genes can significantly repress activator function on transiently introduced DNA. We suggest that this function contributes to the stable repression of targeted loci during development.

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11

Christis, Chantal, and Sean Munro. "The small G protein Arl1 directs the trans-Golgi–specific targeting of the Arf1 exchange factors BIG1 and BIG2." Journal of Cell Biology 196, no. 3 (January 30, 2012): 327–35. http://dx.doi.org/10.1083/jcb.201107115.

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The small G protein Arf1 regulates Golgi traffic and is activated by two related types of guanine nucleotide exchange factor (GEF). GBF1 acts at the cis-Golgi, whereas BIG1 and its close paralog BIG2 act at the trans-Golgi. Peripheral membrane proteins such as these GEFs are often recruited to membranes by small G proteins, but the basis for specific recruitment of Arf GEFs, and hence Arfs, to Golgi membranes is not understood. In this paper, we report a liposome-based affinity purification method to identify effectors for small G proteins of the Arf family. We validate this with the Drosophila melanogaster Arf1 orthologue (Arf79F) and the related class II Arf (Arf102F), which showed a similar pattern of effector binding. Applying the method to the Arf-like G protein Arl1, we found that it binds directly to Sec71, the Drosophila ortholog of BIG1 and BIG2, via an N-terminal region. We show that in mammalian cells, Arl1 is necessary for Golgi recruitment of BIG1 and BIG2 but not GBF1. Thus, Arl1 acts to direct a trans-Golgi–specific Arf1 GEF, and hence active Arf1, to the trans side of the Golgi.

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12

Sinka, Rita, Alison K. Gillingham, Vangelis Kondylis, and Sean Munro. "Golgi coiled-coil proteins contain multiple binding sites for Rab family G proteins." Journal of Cell Biology 183, no. 4 (November 10, 2008): 607–15. http://dx.doi.org/10.1083/jcb.200808018.

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Vesicles and other carriers destined for the Golgi apparatus must be guided to the correct cisternae. Golgins, long coiled-coil proteins that localize to particular Golgi subdomains via their C termini, are candidate regulators of vesicle sorting. In this study, we report that the GRIP domain golgins, whose C termini bind the Arf-like 1 G protein on the trans-Golgi, can also bind four members of the Rab family of G proteins. The Rab2-, Rab6-, Rab19-, and Rab30-binding sites are within the coiled-coil regions that are not required for Golgi targeting. Binding sites for two of these Rabs are also present on two coiled-coil proteins of the cis-Golgi, the Drosophila melanogaster orthologues of GM130 and GMAP-210. We suggest an integrated model for a tentacular Golgi in which coiled-coil proteins surround the Golgi to capture and retain Rab-containing membranes, excluding other structures such as ribosomes. Binding sites for diverse Rabs could ensure that incoming carriers are captured on first contact and moved to their correct destination within the stack.

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Fuse, Naoyuki, Kanako Hisata, Alisa L. Katzen, and Fumio Matsuzaki. "Heterotrimeric G Proteins Regulate Daughter Cell Size Asymmetry in Drosophila Neuroblast Divisions." Current Biology 13, no. 11 (May 2003): 947–54. http://dx.doi.org/10.1016/s0960-9822(03)00334-8.

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14

Granderath, S., A. Stollewerk, S. Greig, C. S. Goodman, C. J. O'Kane, and C. Klambt. "loco encodes an RGS protein required for Drosophila glial differentiation." Development 126, no. 8 (April 15, 1999): 1781–91. http://dx.doi.org/10.1242/dev.126.8.1781.

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In Drosophila, glial cell development depends on the gene glial cells missing (gcm). gcm activates the expression of other transcription factors such as pointed and repo, which control subsequent glial differentiation. In order to better understand glial cell differentiation, we have screened for genes whose expression in glial cells depends on the activity of pointed. Using an enhancer trap approach, we have identified loco as such a gene. loco is expressed in most lateral CNS glial cells throughout development. Embryos lacking loco function have an normal overall morphology, but fail to hatch. Ultrastructural analysis of homozygous mutant loco embryos reveals a severe glial cell differentiation defect. Mutant glial cells fail to properly ensheath longitudinal axon tracts and do not form the normal glial-glial cell contacts, resulting in a disruption of the blood-brain barrier. Hypomorphic loco alleles were isolated following an EMS mutagenesis. Rare escapers eclose which show impaired locomotor capabilities. loco encodes the first two known Drosophila members of the family of Regulators of G-protein signalling (RGS) proteins, known to interact with the alpha subunits of G-proteins. loco specifically interacts with the Drosophila alphai-subunit. Strikingly, the interaction is not confined to the RGS domain. This interaction and the coexpression of LOCO and Galphai suggests a function of G-protein signalling for glial cell development.

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Jones, Clark A., Joyce Ng, Aidan J. Peterson, Kelly Morgan, Jeffrey Simon, and Richard S. Jones. "The Drosophila esc and E(z) Proteins Are Direct Partners in Polycomb Group-Mediated Repression." Molecular and Cellular Biology 18, no. 5 (May 1, 1998): 2825–34. http://dx.doi.org/10.1128/mcb.18.5.2825.

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ABSTRACT The extra sex combs (esc) and Enhancer of zeste [E(z)] proteins are members of the Drosophila Polycomb group (Pc-G) of transcriptional repressors. Here we present evidence for direct physical interaction between the esc and E(z) proteins using yeast two-hybrid and in vitro binding assays. In addition, coimmunoprecipitation from embryo extracts demonstrates association of esc and E(z) in vivo. We have delimited the esc-binding domain of E(z) to an N-terminal 33-amino-acid region. Furthermore, we demonstrate that site-directed mutations in the esc protein previously shown to impair esc function in vivo disrupt esc-E(z) interactions in vitro. We also show an in vitro interaction between the heed and EZH1 proteins, which are human homologs of esc and E(z), respectively. These results suggest that the esc-E(z) molecular partnership has been conserved in evolution. Previous studies suggested that esc is primarily involved in the early stages of Pc-G-mediated silencing during embryogenesis. However, E(z) is continuously required in order to maintain chromosome binding by other Pc-G proteins. In light of these earlier observations and the molecular data presented here, we discuss how esc-E(z) protein complexes may contribute to transcriptional silencing by the Pc-G.

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Kaushik, Rachna, Fengwei Yu, William Chia, Xiaohang Yang, and Sami Bahri. "Subcellular Localization of LGN During Mitosis: Evidence for Its Cortical Localization in Mitotic Cell Culture Systems and Its Requirement for Normal Cell Cycle Progression." Molecular Biology of the Cell 14, no. 8 (August 2003): 3144–55. http://dx.doi.org/10.1091/mbc.e03-04-0212.

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Mammalian LGN/AGS3 proteins and their Drosophila Pins orthologue are cytoplasmic regulators of G-protein signaling. In Drosophila, Pins localizes to the lateral cortex of polarized epithelial cells and to the apical cortex of neuroblasts where it plays important roles in their asymmetric division. Using overexpression studies in different cell line systems, we demonstrate here that, like Drosophila Pins, LGN can exhibit enriched localization at the cell cortex, depending on the cell cycle and the culture system used. We find that in WISH, PC12, and NRK but not COS cells, LGN is largely directed to the cell cortex during mitosis. Overexpression of truncated protein domains further identified the Gα-binding C-terminal portion of LGN as a sufficient domain for cortical localization in cell culture. In mitotic COS cells that normally do not exhibit cortical LGN localization, LGN is redirected to the cell cortex upon overexpression of Gα subunits of heterotrimeric G-proteins. The results also show that the cortical localization of LGN is dependent on microfilaments and that interfering with LGN function in cultured cell lines causes early disruption to cell cycle progression.

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Kim, Junhyong, and John R. Carlson. "Gene discovery by e-genetics: Drosophila odor and taste receptors." Journal of Cell Science 115, no. 6 (March 15, 2002): 1107–12. http://dx.doi.org/10.1242/jcs.115.6.1107.

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A new algorithm that examines DNA databases for proteins that have a particular structure, as opposed to a particular sequence, represents a novel`e-genetics' approach to gene discovery. The algorithm has successfully identified new G-protein-coupled receptors, which have a characteristic seven-transmembrane-domain structure, from the Drosophila genome database. In particular, it has revealed novel families of odor receptors and taste receptors, which had long eluded identification by other means. The two new gene families, the Or and Gr genes, are expressed in neurons of olfactory and taste sensilla and are highly divergent from all other known G-protein-coupled receptor genes. Modification of the algorithm should allow identification of other classes of multitransmembrane-domain protein.

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Yi, Peng, Aaron N. Johnson, Zhe Han, Jiang Wu, and Eric N. Olson. "Heterotrimeric G Proteins Regulate a Noncanonical Function of Septate Junction Proteins to Maintain Cardiac Integrity in Drosophila." Developmental Cell 15, no. 5 (November 2008): 704–13. http://dx.doi.org/10.1016/j.devcel.2008.10.001.

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19

Smart, Renee, Aidan Kiely, Morgan Beale, Ernesto Vargas, Colm Carraher, Andrew V. Kralicek, David L. Christie, Chen Chen, Richard D. Newcomb, and Coral G. Warr. "Drosophila odorant receptors are novel seven transmembrane domain proteins that can signal independently of heterotrimeric G proteins." Insect Biochemistry and Molecular Biology 38, no. 8 (August 2008): 770–80. http://dx.doi.org/10.1016/j.ibmb.2008.05.002.

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20

Wang, Hongyan, Kian Hong Ng, Hongliang Qian, David P. Siderovski, William Chia, and Fengwei Yu. "Ric-8 controls Drosophila neural progenitor asymmetric division by regulating heterotrimeric G proteins." Nature Cell Biology 7, no. 11 (October 16, 2005): 1091–98. http://dx.doi.org/10.1038/ncb1317.

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Hampoelz, Bernhard, Oliver Hoeller, Sarah K. Bowman, Dara Dunican, and Juergen A. Knoblich. "Drosophila Ric-8 is essential for plasma-membrane localization of heterotrimeric G proteins." Nature Cell Biology 7, no. 11 (October 16, 2005): 1099–105. http://dx.doi.org/10.1038/ncb1318.

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Yao, C. A., and J. R. Carlson. "Role of G-Proteins in Odor-Sensing and CO2-Sensing Neurons in Drosophila." Journal of Neuroscience 30, no. 13 (March 31, 2010): 4562–72. http://dx.doi.org/10.1523/jneurosci.6357-09.2010.

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Matunis, E. L., M. J. Matunis, and G. Dreyfuss. "Characterization of the major hnRNP proteins from Drosophila melanogaster." Journal of Cell Biology 116, no. 2 (January 15, 1992): 257–69. http://dx.doi.org/10.1083/jcb.116.2.257.

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To better understand the role(s) of hnRNP proteins in the process of mRNA formation, we have identified and characterized the major nuclear proteins that interact with hnRNAs in Drosophila melanogaster. cDNA clones of several D. melanogaster hnRNP proteins have been isolated and sequenced, and the genes encoding these proteins have been mapped cytologically on polytene chromosomes. These include the hnRNP proteins hrp36, hrp40, and hrp48, which together account for the major proteins of hnRNP complexes in D. melanogaster (Matunis et al., 1992, accompanying paper). All of the proteins described here contain two amino-terminal RNP consensus sequence RNA-binding domains and a carboxyl-terminal glycine-rich domain. We refer to this configuration, which is also found in the hnRNP A/B proteins of vertebrates, as 2 x RBD-Gly. The sequences of the D. melanogaster hnRNP proteins help define both highly conserved and variable amino acids within each RBD and support the suggestion that each RBD in multiple RBD-containing proteins has been conserved independently and has a different function. Although 2 x RBD-Gly proteins from evolutionarily distant organisms are conserved in their general structure, we find a surprising diversity among the members of this family of proteins. A mAb to the hrp40 proteins crossreacts with the human A/B and G hnRNP proteins and detects immunologically related proteins in divergent organisms from yeast to man. These data establish 2 x RBD-Gly as a prevalent hnRNP protein structure across eukaryotes. This information about the composition of hnRNP complexes and about the structure of hnRNA-binding proteins will facilitate studies of the functions of these proteins.

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Lev, Sima, John Hernandez, Ricardo Martinez, Alon Chen, Greg Plowman, and Joseph Schlessinger. "Identification of a Novel Family of Targets of PYK2 Related to Drosophila Retinal Degeneration B (rdgB) Protein." Molecular and Cellular Biology 19, no. 3 (March 1, 1999): 2278–88. http://dx.doi.org/10.1128/mcb.19.3.2278.

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ABSTRACT The protein tyrosine kinase PYK2 has been implicated in signaling pathways activated by G-protein-coupled receptors, intracellular calcium, and stress signals. Here we describe the molecular cloning and characterization of a novel family of PYK2-binding proteins designated Nirs (PYK2 N-terminal domain-interacting receptors). The three Nir proteins (Nir1, Nir2, and Nir3) bind to the amino-terminal domain of PYK2 via a conserved sequence motif located in the carboxy terminus. The primary structures of Nirs reveal six putative transmembrane domains, a region homologous to phosphatidylinositol (PI) transfer protein, and an acidic domain. The Nir proteins are the human homologues of the Drosophila retinal degeneration B protein (rdgB), a protein implicated in the visual transduction pathway in flies. We demonstrate that Nirs are calcium-binding proteins that exhibit PI transfer activity in vivo. Activation of PYK2 by agents that elevate intracellular calcium or by phorbol ester induce tyrosine phosphorylation of Nirs. Moreover, PYK2 and Nirs exhibit similar expression patterns in several regions of the brain and retina. In addition, PYK2-Nir complexes are detected in lysates prepared from cultured cells or from brain tissues. Finally, the Nir1-encoding gene is located at human chromosome 17p13.1, in proximity to a locus responsible for several human retinal diseases. We propose that the Nir and rdgB proteins represent a new family of evolutionarily conserved PYK2-binding proteins that play a role in the control of calcium and phosphoinositide metabolism downstream of G-protein-coupled receptors.

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Nagao, R. T., E. Czarnecka, W. B. Gurley, F. Schöffl, and J. L. Key. "Genes for low-molecular-weight heat shock proteins of soybeans: sequence analysis of a multigene family." Molecular and Cellular Biology 5, no. 12 (December 1985): 3417–28. http://dx.doi.org/10.1128/mcb.5.12.3417.

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Soybeans, Glycine max, synthesize a family of low-molecular-weight heat shock (HS) proteins in response to HS. The DNA sequences of two genes encoding 17.5- and 17.6-kilodalton HS proteins were determined. Nuclease S1 mapping of the corresponding mRNA indicated multiple start termini at the 5' end and multiple stop termini at the 3' end. These two genes were compared with two other soybean HS genes of similar size. A comparison among the 5' flanking regions encompassing the presumptive HS promoter of the soybean HS-protein genes demonstrated this region to be extremely homologous. Analysis of the DNA sequences in the 5' flanking regions of the soybean genes with the corresponding regions of Drosophila melanogaster HS-protein genes revealed striking similarity between plants and animals in the presumptive promoter structure of thermoinducible genes. Sequences related to the Drosophila HS consensus regulatory element were found 57 to 62 base pairs 5' to the start of transcription in addition to secondary HS consensus elements located further upstream. Comparative analysis of the deduced amino acid sequences of four soybean HS proteins illustrated that these proteins were greater than 90% homologous. Comparison of the amino acid sequence for soybean HS proteins with other organisms showed much lower homology (less than 20%). Hydropathy profiles for Drosophila, Xenopus, Caenorhabditis elegans, and G. max HS proteins showed a similarity of major hydrophilic and hydrophobic regions, which suggests conservation of functional domains for these proteins among widely dispersed organisms.

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Nagao, R. T., E. Czarnecka, W. B. Gurley, F. Schöffl, and J. L. Key. "Genes for low-molecular-weight heat shock proteins of soybeans: sequence analysis of a multigene family." Molecular and Cellular Biology 5, no. 12 (December 1985): 3417–28. http://dx.doi.org/10.1128/mcb.5.12.3417-3428.1985.

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Soybeans, Glycine max, synthesize a family of low-molecular-weight heat shock (HS) proteins in response to HS. The DNA sequences of two genes encoding 17.5- and 17.6-kilodalton HS proteins were determined. Nuclease S1 mapping of the corresponding mRNA indicated multiple start termini at the 5' end and multiple stop termini at the 3' end. These two genes were compared with two other soybean HS genes of similar size. A comparison among the 5' flanking regions encompassing the presumptive HS promoter of the soybean HS-protein genes demonstrated this region to be extremely homologous. Analysis of the DNA sequences in the 5' flanking regions of the soybean genes with the corresponding regions of Drosophila melanogaster HS-protein genes revealed striking similarity between plants and animals in the presumptive promoter structure of thermoinducible genes. Sequences related to the Drosophila HS consensus regulatory element were found 57 to 62 base pairs 5' to the start of transcription in addition to secondary HS consensus elements located further upstream. Comparative analysis of the deduced amino acid sequences of four soybean HS proteins illustrated that these proteins were greater than 90% homologous. Comparison of the amino acid sequence for soybean HS proteins with other organisms showed much lower homology (less than 20%). Hydropathy profiles for Drosophila, Xenopus, Caenorhabditis elegans, and G. max HS proteins showed a similarity of major hydrophilic and hydrophobic regions, which suggests conservation of functional domains for these proteins among widely dispersed organisms.

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Aleyakpo, Benjamin, Oghenetega Umukoro, Ryan Kavlie, Daniel C. Ranson, Andrew Thompsett, Olivia Corcoran, and Stefano O. Casalotti. "G-protein αq gene expression plays a role in alcohol tolerance in Drosophila melanogaster." Brain and Neuroscience Advances 3 (January 2019): 239821281988308. http://dx.doi.org/10.1177/2398212819883081.

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Ethanol is a psychoactive substance causing both short- and long-term behavioural changes in humans and animal models. We have used the fruit fly Drosophila melanogaster to investigate the effect of ethanol exposure on the expression of the Gαq protein subunit. Repetitive exposure to ethanol causes a reduction in sensitivity (tolerance) to ethanol, which we have measured as the time for 50% of a set of flies to become sedated after exposure to ethanol (ST50). We demonstrate that the same treatment that induces an increase in ST50 over consecutive days (tolerance) also causes a decrease in Gαq protein subunit expression at both the messenger RNA and protein level. To identify whether there may be a causal relationship between these two outcomes, we have developed strains of flies in which Gαq messenger RNA expression is suppressed in a time- and tissue-specific manner. In these flies, the sensitivity to ethanol and the development of tolerance are altered. This work further supports the value of Drosophila as a model to dissect the molecular mechanisms of the behavioural response to alcohol and identifies G proteins as potentially important regulatory targets for alcohol use disorders.

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Xu, Xian-Zhong Shawn, Atish Choudhury, Xiaoling Li, and Craig Montell. "Coordination of an Array of Signaling Proteins through Homo- and Heteromeric Interactions Between PDZ Domains and Target Proteins." Journal of Cell Biology 142, no. 2 (July 27, 1998): 545–55. http://dx.doi.org/10.1083/jcb.142.2.545.

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The rapid activation and feedback regulation of many G protein signaling cascades raises the possibility that the critical signaling proteins may be tightly coupled. Previous studies show that the PDZ domain containing protein INAD, which functions in Drosophila vision, coordinates a signaling complex by binding directly to the light-sensitive ion channel, TRP, and to phospholipase C (PLC). The INAD signaling complex also includes rhodopsin, protein kinase C (PKC), and calmodulin, though it is not known whether these proteins bind to INAD. In the current work, we show that rhodopsin, calmodulin, and PKC associate with the signaling complex by direct binding to INAD. We also found that a second ion channel, TRPL, bound to INAD. Thus, most of the proteins involved directly in phototransduction appear to bind to INAD. Furthermore, we found that INAD formed homopolymers and the homomultimerization occurred through two PDZ domains. Thus, we propose that the INAD supramolecular complex is a higher order signaling web consisting of an extended network of INAD molecules through which a G protein–coupled cascade is tethered.

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Matunis, M. J., E. L. Matunis, and G. Dreyfuss. "Isolation of hnRNP complexes from Drosophila melanogaster." Journal of Cell Biology 116, no. 2 (January 15, 1992): 245–55. http://dx.doi.org/10.1083/jcb.116.2.245.

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Nascent RNA polymerase II transcripts, heterogeneous nuclear RNAs (hnRNAs), become associated with nuclear proteins (hnRNP Proteins), and their processing into mRNAs takes place in these hnRNP complexes. hnRNP complexes have previously been purified from vertebrate cells. Here we report the isolation of hnRNP complexes from an invertebrate organism, the fruitfly Drosophila melanogaster. Candidate hnRNP proteins were purified from D. melanogaster embryos by ssDNA affinity chromatography, and mAbs were produced to many of the major proteins. Genuine hnRNP proteins were identified by several criteria, including nucleoplasmic localization, association with nascent transcripts, crosslinking to poly(A)-containing RNA in living cells, and amino acid sequence. In addition, mAbs that cross-react between the fruitfly and human hnRNP proteins were obtained. Most importantly, using hnRNP-specific mAbs we have purified the hnRNP complexes from D. melanogaster cells. These RNAase-sensitive complexes contain at least 10 major proteins designated hrps, the most abundant proteins having apparent molecular masses of 36, 38, 39, 40, 44, 48, 54, 62, 70, and 75 kD. cDNAs and complete sequences for several of these proteins have been obtained and are presented in the accompanying paper (Matunis, E. L., M. J. Matunis, and G. Dreyfuss. 1992. J. Cell Biol. 116:257-269). The purification of D. melanogaster hnRNP complexes will facilitate genetic and cytological studies on the function of hnRNA-binding proteins and on the posttranscriptional regulation of gene expression.

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30

Hoey, T., R. Warrior, J. Manak, and M. Levine. "DNA-binding activities of the Drosophila melanogaster even-skipped protein are mediated by its homeo domain and influenced by protein context." Molecular and Cellular Biology 8, no. 11 (November 1988): 4598–607. http://dx.doi.org/10.1128/mcb.8.11.4598.

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The homeo box gene even-skipped (eve) encodes a 376-amino-acid protein that binds with high affinity to sequences located near the 5' termini of the eve and en genes. The 5' en sites are A + T rich and contain copies of the 10-base-pair (bp) consensus sequence T-C-A-A-T-T-A-A-A-T. In contrast, the 5' eve sites are G + C rich and contain the 9-bp sequence T-C-A-G-C-A-C-C-G. Among the five different homeo box proteins that have been tested for binding, eve is unique in that it shows virtually equal preference for the A + T-rich 5' en binding sites and the G + C-rich 5' eve sites. Most of the other proteins bind with a relatively higher affinity to the en sites than to the eve sites. In an effort to identify the regions of the eve protein that are responsible for its efficient binding to both classes of recognition sequences, we analyzed the DNA-binding properties of various mutant eve proteins. These studies suggest that the homeo domain of the eve protein is responsible for both binding activities. However, mutations in distant regions of the protein influenced the binding behavior of the eve homeo domain and caused a reduction in binding to the G + C class of recognition sites. We propose that the protein context of the homeo domain can influence its DNA-binding properties.

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31

Hoey, T., R. Warrior, J. Manak, and M. Levine. "DNA-binding activities of the Drosophila melanogaster even-skipped protein are mediated by its homeo domain and influenced by protein context." Molecular and Cellular Biology 8, no. 11 (November 1988): 4598–607. http://dx.doi.org/10.1128/mcb.8.11.4598-4607.1988.

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The homeo box gene even-skipped (eve) encodes a 376-amino-acid protein that binds with high affinity to sequences located near the 5' termini of the eve and en genes. The 5' en sites are A + T rich and contain copies of the 10-base-pair (bp) consensus sequence T-C-A-A-T-T-A-A-A-T. In contrast, the 5' eve sites are G + C rich and contain the 9-bp sequence T-C-A-G-C-A-C-C-G. Among the five different homeo box proteins that have been tested for binding, eve is unique in that it shows virtually equal preference for the A + T-rich 5' en binding sites and the G + C-rich 5' eve sites. Most of the other proteins bind with a relatively higher affinity to the en sites than to the eve sites. In an effort to identify the regions of the eve protein that are responsible for its efficient binding to both classes of recognition sequences, we analyzed the DNA-binding properties of various mutant eve proteins. These studies suggest that the homeo domain of the eve protein is responsible for both binding activities. However, mutations in distant regions of the protein influenced the binding behavior of the eve homeo domain and caused a reduction in binding to the G + C class of recognition sites. We propose that the protein context of the homeo domain can influence its DNA-binding properties.

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32

Martin, E. C., and P. N. Adler. "The Polycomb group gene Posterior Sex Combs encodes a chromosomal protein." Development 117, no. 2 (February 1, 1993): 641–55. http://dx.doi.org/10.1242/dev.117.2.641.

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The Posterior Sex Combs (Psc) gene of Drosophila has been studied at the molecular level both because it is a Polycomb group (Pc-G) gene and hence required for the maintenance of segmental determination, and because it is the Drosophila homolog of the murine bmi-1 oncogene. Although genetic interactions indicated that Psc functioned as a Pc-G gene, the zygotic mutant phenotype of Psc showed little evidence of segmental transformations. We have examined mutant embryos derived from a mutant maternal germ line and found a stronger mutant phenotype, indicating that the weak zygotic phenotype of Psc is due to maternal rescue. We have found that Psc RNA accumulates in developing oocytes and this maternal RNA is presumably responsible for the maternal rescue. We have studied the expression of the Psc gene at both the RNA and protein levels. On northern blots, we find evidence for two Psc mRNAs and, on western blots, we find evidence for two Psc proteins that are altered either in abundance or size in Psc mutants. The Psc protein accumulates in all regions of the embryo and also in many tissues in a variety of developmental stages. In all cases, it is nuclear, as is its mammalian homolog, the bmi-1 protein. On polytene chromosomes, we find Psc at 45 chromosomal loci where two other Pc-G proteins are present.

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Kunwar, Prabhat S., Hiroko Sano, Andrew D. Renault, Vitor Barbosa, Naoyuki Fuse, and Ruth Lehmann. "Tre1 GPCR initiates germ cell transepithelial migration by regulating Drosophila melanogaster E-cadherin." Journal of Cell Biology 183, no. 1 (September 29, 2008): 157–68. http://dx.doi.org/10.1083/jcb.200807049.

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Despite significant progress in identifying the guidance pathways that control cell migration, how a cell starts to move within an intact organism, acquires motility, and loses contact with its neighbors is poorly understood. We show that activation of the G protein–coupled receptor (GPCR) trapped in endoderm 1 (Tre1) directs the redistribution of the G protein Gβ as well as adherens junction proteins and Rho guanosine triphosphatase from the cell periphery to the lagging tail of germ cells at the onset of Drosophila melanogaster germ cell migration. Subsequently, Tre1 activity triggers germ cell dispersal and orients them toward the midgut for directed transepithelial migration. A transition toward invasive migration is also a prerequisite for metastasis formation, which often correlates with down-regulation of adhesion proteins. We show that uniform down-regulation of E-cadherin causes germ cell dispersal but is not sufficient for transepithelial migration in the absence of Tre1. Our findings therefore suggest a new mechanism for GPCR function that links cell polarity, modulation of cell adhesion, and invasion.

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Ng, J., R. Li, K. Morgan, and J. Simon. "Evolutionary conservation and predicted structure of the Drosophila extra sex combs repressor protein." Molecular and Cellular Biology 17, no. 11 (November 1997): 6663–72. http://dx.doi.org/10.1128/mcb.17.11.6663.

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The Drosophila extra sex combs (esc) protein, a member of the Polycomb group (PcG), is a transcriptional repressor of homeotic genes. Genetic studies have shown that esc protein is required in early embryos at about the time that other PcG proteins become engaged in homeotic gene repression. The esc protein consists primarily of multiple copies of the WD repeat, a motif that has been implicated in protein-protein interaction. To further investigate the domain organization of esc protein, we have isolated and characterized esc homologs from divergent insect species. We report that esc protein is highly conserved in housefly (72% identical to Drosophila esc), butterfly (55% identical), and grasshopper (56% identical). We show that the butterfly homolog provides esc function in Drosophila, indicating that the sequence similarities reflect functional conservation. Homology modeling using the crystal structure of another WD repeat protein, the G-protein beta-subunit, predicts that esc protein adopts a beta-propeller structure. The sequence comparisons and modeling suggest that there are seven WD repeats in esc protein which together form a seven-bladed beta-propeller. We locate the conserved regions in esc protein with respect to this predicted structure. Site-directed mutagenesis of specific loops, predicted to extend from the propeller surface, identifies conserved parts of esc protein required for function in vivo. We suggest that these regions might mediate physical interaction with esc partner proteins.

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Miller, K. G., C. M. Field, and B. M. Alberts. "Actin-binding proteins from Drosophila embryos: a complex network of interacting proteins detected by F-actin affinity chromatography." Journal of Cell Biology 109, no. 6 (December 1, 1989): 2963–75. http://dx.doi.org/10.1083/jcb.109.6.2963.

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By using F-actin affinity chromatography columns to select proteins solely by their ability to bind to actin filaments, we have identified and partially purified greater than 40 proteins from early Drosophila embryos. These proteins represent approximately 0.5% of the total protein present in soluble cell extracts, and 2 mg are obtained by chromatography of an extract from 10 g of embryos. As judged by immunofluorescence of fixed embryos, 90% of the proteins that we have detected in F-actin column eluates are actin-associated in vivo (12 of 13 proteins tested). The distributions of antigens observed suggest that groups of these proteins cooperate in generating unique actin structures at different places in the cell. These structures change as cells progress through the cell cycle and as they undergo the specializations that accompany development. The variety of different spatial localizations that we have observed in a small subset of the total actin-binding proteins suggests that the actin cytoskeleton is a very complex network of interacting proteins.

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36

Quan, F., and M. A. Forte. "Two forms of Drosophila melanogaster Gs alpha are produced by alternate splicing involving an unusual splice site." Molecular and Cellular Biology 10, no. 3 (March 1990): 910–17. http://dx.doi.org/10.1128/mcb.10.3.910.

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G proteins are responsible for modulating the activity of intracellular effector systems in response to receptor activation. The stimulatory G protein Gs is responsible for activation of adenylate cyclase in response to a variety of hormonal signals. In this report, we describe the structure of the gene for the alpha subunit of Drosophila melanogaster Gs. The gene is approximately 4.5 kilobases long and is divided into nine exons. The exon-intron structure of the Drosophila gene shows substantial similarity to that of the human gene for Gs alpha. Alternate splicing of intron 7, involving either use of an unusual TG or consensus AG 3' splice site, results in transcripts which code for either a long (DGs alpha L) or short (DGs alpha S) form of Gs alpha. These subunits differ by inclusion or deletion of three amino acids and substitution of a Ser for a Gly. The two forms of Drosophila Gs alpha differ in a region where no variation in the primary sequence of vertebrate Gs alpha subunits has been observed. In vitro translation experiments demonstrated that the Drosophila subunits migrate anomalously on sodium dodecyl sulfate-polyacrylamide gels with apparent molecular weights of 51,000 and 48,000. Additional Gs alpha transcript heterogeneity reflects the use of multiple polyadenylation sites.

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Quan, F., and M. A. Forte. "Two forms of Drosophila melanogaster Gs alpha are produced by alternate splicing involving an unusual splice site." Molecular and Cellular Biology 10, no. 3 (March 1990): 910–17. http://dx.doi.org/10.1128/mcb.10.3.910-917.1990.

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G proteins are responsible for modulating the activity of intracellular effector systems in response to receptor activation. The stimulatory G protein Gs is responsible for activation of adenylate cyclase in response to a variety of hormonal signals. In this report, we describe the structure of the gene for the alpha subunit of Drosophila melanogaster Gs. The gene is approximately 4.5 kilobases long and is divided into nine exons. The exon-intron structure of the Drosophila gene shows substantial similarity to that of the human gene for Gs alpha. Alternate splicing of intron 7, involving either use of an unusual TG or consensus AG 3' splice site, results in transcripts which code for either a long (DGs alpha L) or short (DGs alpha S) form of Gs alpha. These subunits differ by inclusion or deletion of three amino acids and substitution of a Ser for a Gly. The two forms of Drosophila Gs alpha differ in a region where no variation in the primary sequence of vertebrate Gs alpha subunits has been observed. In vitro translation experiments demonstrated that the Drosophila subunits migrate anomalously on sodium dodecyl sulfate-polyacrylamide gels with apparent molecular weights of 51,000 and 48,000. Additional Gs alpha transcript heterogeneity reflects the use of multiple polyadenylation sites.

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Schaefer, Matthias, Mark Petronczki, Daniela Dorner, Michael Forte, and Juergen A. Knoblich. "Heterotrimeric G Proteins Direct Two Modes of Asymmetric Cell Division in the Drosophila Nervous System." Cell 107, no. 2 (October 2001): 183–94. http://dx.doi.org/10.1016/s0092-8674(01)00521-9.

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Peters, Kimberly A., and Stephen L. Rogers. "Drosophila Ric-8 interacts with the Gα12/13 subunit, Concertina, during activation of the Folded gastrulation pathway." Molecular Biology of the Cell 24, no. 21 (November 2013): 3460–71. http://dx.doi.org/10.1091/mbc.e12-11-0813.

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Heterotrimeric G proteins, composed of α, β, and γ subunits, are activated by exchange of GDP for GTP on the Gα subunit. Canonically, Gα is stimulated by the guanine-nucleotide exchange factor (GEF) activity of ligand-bound G protein–coupled receptors. However, Gα subunits may also be activated in a noncanonical manner by members of the Ric-8 family, cytoplasmic proteins that also act as GEFs for Gα subunits. We used a signaling pathway active during Drosophila gastrulation as a model system to study Ric-8/Gα interactions. A component of this pathway, the Drosophila Gα12/13 subunit, Concertina (Cta), is necessary to trigger actomyosin contractility during gastrulation events. Ric-8 mutants exhibit similar gastrulation defects to Cta mutants. Here we use a novel tissue culture system to study a signaling pathway that controls cytoskeletal rearrangements necessary for cellular morphogenesis. We show that Ric-8 regulates this pathway through physical interaction with Cta and preferentially interacts with inactive Cta and directs its localization within the cell. We also use this system to conduct a structure–function analysis of Ric-8 and identify key residues required for both Cta interaction and cellular contractility.

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Tata, Fred, and David A. Hartley. "The role of the Enhancer of split complex during cell fate determination in Drosophila." Development 119, Supplement (December 1, 1993): 139–48. http://dx.doi.org/10.1242/dev.119.supplement.139.

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Molecular and genetic data predict that the Enhancer of split locus functions at the end of a pathway dictating appropriate cell fate determination in a number of developmental contexts. We have sought to dissect the role individual member genes of the complex play through a molecular analysis. Of the two principle class of genes, the first, members of the basic helix-loop-helix (bHLH) class of proteins are expressed in specific regions of the embryo in subtle, overlapping patterns in cells that will differentiate as epidermis. The second, groucho, a member of the WD40 class of proteins, is expressed more generally. Immunoprecipitation experiments do not implicate groucho in G protein mediated signal transduction, a known function of many WD40 type proteins. Instead, the nuclear localisation of the protein suggests a relationship to the bHLH members of the complex. Differences in expression of the bHLH genes between neurogenic mutants implies two pathways to their activation during epidermal determination.

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Gu, Qiuxiang, Jinglin Wu, Yao Tian, Shanshan Cheng, Zi Chao Zhang, and Junhai Han. "Gαq splice variants mediate phototransduction, rhodopsin synthesis, and retinal integrity in Drosophila." Journal of Biological Chemistry 295, no. 17 (March 20, 2020): 5554–63. http://dx.doi.org/10.1074/jbc.ra120.012764.

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Heterotrimeric G proteins mediate a variety of signaling processes by coupling G protein–coupled receptors to intracellular effector molecules. In Drosophila, the Gαq gene encodes several Gαq splice variants, with the Gαq1 isoform protein playing a major role in fly phototransduction. However, Gαq1 null mutant flies still exhibit a residual light response, indicating that other Gαq splice variants or additional Gq α subunits are involved in phototransduction. Here, we isolated a mutant fly with no detectable light responses, decreased rhodopsin (Rh) levels, and rapid retinal degeneration. Using electrophysiological and genetic studies, biochemical assays, immunoblotting, real-time RT-PCR, and EM analysis, we found that mutations in the Gαq gene disrupt light responses and demonstrate that the Gαq3 isoform protein is responsible for the residual light response in Gαq1 null mutants. Moreover, we report that Gαq3 mediates rhodopsin synthesis. Depletion of all Gαq splice variants led to rapid light-dependent retinal degeneration, due to the formation stable Rh1-arrestin 2 (Arr2) complexes. Our findings clarify essential roles of several different Gαq splice variants in phototransduction and retinal integrity in Drosophila and reveal that Gαq3 functions in rhodopsin synthesis.

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Shiel, M. J., and M. J. Caplan. "Developmental regulation of membrane protein sorting in Drosophila embryos." American Journal of Physiology-Cell Physiology 269, no. 1 (July 1, 1995): C207—C216. http://dx.doi.org/10.1152/ajpcell.1995.269.1.c207.

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We have examined the process of membrane protein targeting in the polarized cells of the developing Drosophila melanogaster embryo. Human placental alkaline phosphatase (PLAP) is a glycosylphosphatidyl inositol-linked protein that accumulates at the apical membranes of mammalian epithelial cells. A chimeric construct composed of the transmembrane and cytosolic portions of the vesicular stomatitis virus G protein coupled to the ectodomain of PLAP, termed PLAPG, has been found to behave as a basolateral protein (D. A. Brown, B. Crise, and J. K. Rose. Science Wash. DC 232: 34-47, 1989). The subcellular distributions of these proteins were examined in the epithelial and neuronal tissues of transgenic Drosophila embryos. In the surface ectoderm, both PLAP and PLAPG were restricted to the basolateral membranes throughout development. Internal epithelia derived from the surface ectoderm accumulated PLAP at their apical surfaces, whereas PLAPG retained its basolateral distribution. The redistribution of PLAP from the basolateral to the apical plasma membrane was found to be coincident with the invagination of the surface epithelium to form internal structures, suggesting that the sorting pathways that function in the epithelium of the Drosophila embryo are developmentally regulated.

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Chu, Brian, Che-Hsiung Liu, Sukanya Sengupta, Amit Gupta, Padinjat Raghu, and Roger C. Hardie. "Common mechanisms regulating dark noise and quantum bump amplification in Drosophila photoreceptors." Journal of Neurophysiology 109, no. 8 (April 15, 2013): 2044–55. http://dx.doi.org/10.1152/jn.00001.2013.

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Absolute visual thresholds are limited by “dark noise,” which in Drosophila photoreceptors is dominated by brief (∼10 ms), small (∼2 pA) inward current events, occurring at ∼2/s, believed to reflect spontaneous G protein activations. These dark events were increased in rate and amplitude by a point mutation in myosin III (NINAC), which disrupts its interaction with the scaffolding protein, INAD. This phenotype mimics that previously described in null mutants of ninaC (no inactivation no afterpotential; encoding myosin III) and an associated protein, retinophilin ( rtp). Dark noise was similarly increased in heterozygote mutants of diacylglycerol kinase ( rdgA/+). Dark noise in ninaC, rtp, and rdgA/+ mutants was greatly suppressed by mutations of the Gq α-subunit ( Gα q) and the major light-sensitive channel ( trp) but not rhodopsin. ninaC, rtp, and rdgA/+ mutations also all facilitated residual light responses in Gα q and PLC hypomorphs. Raising cytosolic Ca2+ in the submicromolar range increased dark noise, facilitated activation of transient receptor potential (TRP) channels by exogenous agonist, and again facilitated light responses in Gα q hypomorphs. Our results indicate that RTP, NINAC, INAD, and diacylglycerol kinase, together with a Ca2+-dependent threshold, share common roles in suppressing dark noise and regulating quantum bump generation; consequently, most spontaneous G protein activations fail to generate dark events under normal conditions. By contrast, quantum bump generation is reliable but delayed until sufficient G proteins and PLC are activated to overcome threshold, thereby ensuring generation of full-size bumps with high quantum efficiency.

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Horgan, A. M., M. T. Lagrange, and P. F. Copenhaver. "Developmental expression of G proteins in a migratory population of embryonic neurons." Development 120, no. 4 (April 1, 1994): 729–42. http://dx.doi.org/10.1242/dev.120.4.729.

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Directed neuronal migration contributes to the formation of many developing systems, but the molecular mechanisms that control the migratory process are still poorly understood. We have examined the role of heterotrimeric G proteins (guanyl nucleotide binding proteins) in regulating the migratory behavior of embryonic neurons in the enteric nervous system of the moth, Manduca sexta. During the formation of the enteric nervous system, a group of approx. 300 enteric neurons (the EP cells) participate in a precise migratory sequence, during which the undifferentiated cells populate a branching nerve plexus that lies superficially on the visceral musculature. Once migration is complete, the cells then acquire a variety of position-specific neuronal phenotypes. Using affinity-purified antisera against different G protein subtypes, we found no apparent staining for any G protein in the EP cells prior to their migration. Coincident with the onset of migration, however, the EP cells commenced the expression of one particular G protein, Go alpha. The intensity of immunostaining continued to increase as migration progressed, with Go alpha immunoreactivity being detectable in the leading processes of the neurons as well as their somata. The identity of the Go alpha-related proteins was confirmed by protein immunoblot analysis and by comparison with previously described forms of Go alpha from Drosophila. When cultured embryos were treated briefly with aluminium fluoride, a compound known to stimulate the activity of heterotrimeric G proteins, both EP cell migration and process outgrowth were inhibited. The effects of aluminium fluoride were potentiated by alpha toxin, a pore-forming compound that by itself caused no significant perturbations of migration. In preliminary experiments, intracellular injections of the non-hydrolyzable nucleotide GTP gamma-S also inhibited the migration of individual EP cells, supporting the hypothesis that G proteins play a key role in the control of neuronal motility in this system. In addition, once migration was complete, the expression of Go alpha-related proteins in the EP cells underwent a subsequent phase of regulation, so that only certain phenotypic classes among the differentiated EP cells retained detectable levels of Go alpha immunoreactivity. Thus Go may perform multiple functions within the same population of migratory neurons in the course of embryonic development.

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Abry, Muna F., Kelvin M. Kimenyi, Daniel K. Masiga, and Benard W. Kulohoma. "Comparative genomics identifies male accessory gland proteins in five Glossina species." Wellcome Open Research 2 (August 30, 2017): 73. http://dx.doi.org/10.12688/wellcomeopenres.12445.1.

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Accessory gland proteins (ACPs) are important reproductive proteins produced by the male accessory glands (MAGs) of most insect species. These proteins are essential for male insect fertility, and are transferred alongside semen to females during copulation. ACPs are poorly characterized in Glossina species (tsetse fly), the principal vector of the parasite that causes life-threatening Human African Trypanosomiasis and Animal trypanosomiasis in endemic regions in Africa. The tsetse fly has a peculiar reproductive cycle because of the absence of oviposition. Females mate once and store sperm in a spermathecal, and produce a single fully developed larva at a time that pupates within minutes of exiting their uterus. This slow reproductive cycle, compared to other insects, significantly restricts reproduction to only 3 to 6 larvae per female lifespan. This unique reproductive cycle is an attractive vector control strategy entry point. We exploit comparative genomics approaches to explore the diversity of ACPs in the recently available whole genome sequence data from five tsetse fly species (Glossina morsitans, G. austeni, G. brevipalpis, G. pallidipes and G. fuscipes). We used previously described ACPs in Drosophila melanogaster and Anopheles gambiae as reference sequences. We identified 36, 27, 31, 29 and 33 diverse ACP orthologous genes in G. austeni, G. brevipalpis, G. fuscipes, G. pallidipes and G. morsitans genomes respectively, which we classified into 21 functional classes. Our findings provide genetic evidence of MAG proteins in five recently sequenced Glossina genomes. It provides new avenues for molecular studies that evaluate potential field control strategies of these important vectors of human and animal disease.

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Abry, Muna F., Kelvin M. Kimenyi, Daniel K. Masiga, and Benard W. Kulohoma. "Comparative genomics identifies male accessory gland proteins in five Glossina species." Wellcome Open Research 2 (November 22, 2017): 73. http://dx.doi.org/10.12688/wellcomeopenres.12445.2.

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Accessory gland proteins (ACPs) are important reproductive proteins produced by the male accessory glands (MAGs) of most insect species. These proteins are essential for male insect fertility, and are transferred alongside semen to females during copulation. ACPs are poorly characterized in Glossina species (tsetse fly), the principal vector of the parasite that causes life-threatening Human African Trypanosomiasis and Animal trypanosomiasis in endemic regions in Africa. The tsetse fly has a peculiar reproductive cycle because of the absence of oviposition. Females mate once and store sperm in a spermathecal, and produce a single fully developed larva at a time that pupates within minutes of exiting their uterus. This slow reproductive cycle, compared to other insects, significantly restricts reproduction to only 3 to 6 larvae per female lifespan. This unique reproductive cycle is an attractive vector control strategy entry point. We exploit comparative genomics approaches to explore the diversity of ACPs in the recently available whole genome sequence data from five tsetse fly species ( Glossina morsitans, G. austeni, G. brevipalpis, G. pallidipes and G. fuscipes). We used previously described ACPs in Drosophila melanogaster and Anopheles gambiae as reference sequences. We identified 36, 27, 31, 29 and 33 diverse ACP orthologous genes in G. austeni, G. brevipalpis, G. fuscipes, G. pallidipes and G. morsitans genomes respectively, which we classified into 21 functional classes. Our findings provide genetic evidence of MAG proteins in five recently sequenced Glossina genomes. It highlights new avenues for molecular studies that evaluate potential field control strategies of these important vectors of human and animal disease.

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Lee, B. A., and D. J. Donoghue. "Membrane-anchored form of v-sis/PDGF-B induces mitogenesis without detectable PDGF receptor autophosphorylation." Journal of Cell Biology 113, no. 2 (April 15, 1991): 361–70. http://dx.doi.org/10.1083/jcb.113.2.361.

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The v-sis protein is structurally and functionally related to PDGF. Forms of the v-sis protein which are anchored to the cell membrane via the transmembrane domain of the vesicular stomatitis virus G protein have been previously described (Hannink, M., and D.J. Donoghue. 1986. J. Cell Biol. 103:2311-2322). Several of these fusion proteins were shown to interact productively with the PDGF receptor (PDGFR) based on their ability to transform NIH 3T3 cells. In this report, we further characterized one of these membrane-anchored v-sis proteins, designated v-sis239-G. The gene encoding v-sis239-G was placed under control of the Drosophila melanogaster hsp70 promotor and synthesis of this protein was shown to induce a mitogenic response in NIH 3T3 cells. Unexpectedly, v-sis239-G did not induce detectable autophosphorylation of the PDGFR, in contrast to a similarly expressed secreted form of the v-sis protein. Thus, it appears that a PDGFR-mediated mitogenic response may be dissociated from detectable receptor autophosphorylation. Furthermore, induced synthesis of v-sis239-G was shown to lead to c-fos expression even in the absence of detectable receptor autophosphorylation. Interestingly, a nonmitogenic membrane-anchored form of the v-sis protein, designated v-sis239-G338, also induced c-fos without receptor autophosphorylation. These results raise interesting questions regarding the roles of autophosphorylation and c-fos induction in PDGFR-mediated signal transduction and suggest the possibility of an autophosphorylation-independent signal transduction pathway.

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Zhang, Yuhan, Weiwei Liu, Ronghong Li, Jiaqi Gu, Ping Wu, Chao Peng, Jinbiao Ma, Ligang Wu, Yang Yu, and Ying Huang. "Structural insights into the sequence-specific recognition of Piwi by Drosophila Papi." Proceedings of the National Academy of Sciences 115, no. 13 (March 12, 2018): 3374–79. http://dx.doi.org/10.1073/pnas.1717116115.

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The Tudor domain-containing (Tdrd) family proteins play a critical role in transposon silencing in animal gonads by recognizing the symmetrically dimethylated arginine (sDMA) on the (G/A)R motif of the N-terminal of PIWI family proteins via the eTud domains. Papi, also known as “Tdrd2,” is involved in Zucchini-mediated PIWI-interacting RNA (piRNA) 3′-end maturation. Intriguingly, a recent study showed that, in papi mutant flies, only Piwi-bound piRNAs increased in length, and not Ago3-bound or Aub-bound piRNAs. However, the molecular and structural basis of the Papi–Piwi complex is still not fully understood, which limits mechanistic understanding of the function of Papi in piRNA biogenesis. In the present study, we determined the crystal structures of Papi-eTud in the apo form and in complex with a peptide containing unmethylated or dimethylated R10 residues. Structural and biochemical analysis showed that the Papi interaction region on the Drosophila Piwi contains an RGRRR motif (R7–R11) distinct from the consensus (G/A)R motif recognized by canonical eTud. Mass spectrometry results indicated that Piwi is the major binding partner of Papi in vivo. The papi mutant flies suffered from both fertility and transposon-silencing defects, supporting the important role conferred to Papi in piRNA 3′ processing through direct interaction with Piwi proteins.

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Kellerman, K. A., and K. G. Miller. "An unconventional myosin heavy chain gene from Drosophila melanogaster." Journal of Cell Biology 119, no. 4 (November 15, 1992): 823–34. http://dx.doi.org/10.1083/jcb.119.4.823.

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As part of a study of cytoskeletal proteins involved in Drosophila embryonic development, we have undertaken the molecular analysis of a 140-kD ATP-sensitive actin-binding protein (Miller, K. G., C. M. Field, and B. M. Alberts. 1989. J. Cell Biol. 109:2963-2975). Analysis of cDNA clones encoding this protein revealed that it represents a new class of unconventional myosin heavy chains. The amino-terminal two thirds of the protein comprises a head domain that is 29-33% identical (60-65% similar) to other myosin heads, and contains ATP-binding, actin-binding and calmodulin/myosin light chain-binding motifs. The carboxy-terminal tail has no significant similarity to other known myosin tails, but does contain a approximately 100-amino acid region that is predicted to form an alpha-helical coiled-coil. Since the unique gene that encodes this protein maps to the polytene map position 95F, we have named the new gene Drosophila 95F myosin heavy chain (95F MHC). The expression profile of the 95F MHC gene is complex. Examination of multiple cDNAs reveals that transcripts are alternatively spliced and encode at least three protein isoforms; in addition, a fourth isoform is detected on Western blots. Developmental Northern and Western blots show that transcripts and protein are present throughout the life cycle, with peak expression occurring during mid-embryogenesis and adulthood. Immunolocalization in early embryos demonstrates that the protein is primarily located in a punctate pattern throughout the peripheral cytoplasm. Most cells maintain a low level of protein expression throughout embryogenesis, but specific tissues appear to contain more protein. We speculate that the 95F MHC protein isoforms are involved in multiple dynamic processes during Drosophila development.

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Belenkaya, Tatiana, Alexey Soldatov, Elena Nabirochkina, Inna Birjukova, Sofia Georgieva, and Pavel Georgiev. "P-Element Insertion at the polyhomeotic Gene Leads to Formation of a Novel Chimeric Protein That Negatively Regulates yellow Gene Expression in P-Element-Induced Alleles of Drosophila melanogaster." Genetics 150, no. 2 (October 1, 1998): 687–97. http://dx.doi.org/10.1093/genetics/150.2.687.

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Abstract Polyhomeotic is a member of the Polycomb group (Pc-G) of homeotic repressors. The proteins encoded by the Pc-G genes form repressive complexes on the polycomb group response element sites. The phP1 mutation was induced by insertion of a 1.2-kb P element into the 5′ transcribed nontranslated region of the proximal polyhomeotic gene. The phP1 allele confers no mutant phenotype, but represses transcription of P-element-induced alleles at the yellow locus. The phP1 allele encodes a chimeric P-PH protein, consisting of the DNA-binding domain of the P element and the PH protein lacking 12 amino-terminal amino acids. The P-PH, Polycomb (PC), and Posterior sex combs (PSC) proteins were immunohistochemically detected on polytene chromosomes in the regions of P-element insertions.

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Journal articles on the topic 'G proteins ; Drosophila'

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