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1
Goehring, Nathan W., Ivana Petrovska, Dana Boyd, and Jon Beckwith. "Mutants, Suppressors, and Wrinkled Colonies: Mutant Alleles of the Cell Division Gene ftsQ Point to Functional Domains in FtsQ and a Role for Domain 1C of FtsA in Divisome Assembly." Journal of Bacteriology 189, no. 2 (2006): 633–45. http://dx.doi.org/10.1128/jb.00991-06.
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ABSTRACT Cell division in Escherichia coli requires the concerted action of at least 10 essential proteins. One of these proteins, FtsQ, is physically associated with multiple essential division proteins, including FtsK, FtsL, FtsB, FtsW, and FtsI. In this work we performed a genetic analysis of the ftsQ gene. Our studies identified C-terminal residues essential for FtsQ's interaction with two downstream proteins, FtsL and FtsB. Here we also describe a novel screen for cell division mutants based on a wrinkled-colony morphology, which yielded several new point mutations in ftsQ. Two of these mutations affect localization of FtsQ to midcell and together define a targeting role for FtsQ's α domain. Further characterization of one localization-defective mutant protein [FtsQ(V92D)] revealed an unexpected role in localization for the first 49 amino acids of FtsQ. Finally, we found a suppressor of FtsQ(V92D) that was due to a point mutation in domain 1C of FtsA, a domain previously implicated in the recruitment of divisome proteins. However, despite reports of a potential interaction between FtsA and FtsQ, suppression by FtsA(I143L) is not mediated via direct contact with FtsQ. Rather, this mutation acts as a general suppressor of division defects, which include deletions of the normally essential genes zipA and ftsK and mutations in FtsQ that affect both localization and recruitment. Together, these results reveal increasingly complex connections within the bacterial divisome.
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Hale, Cynthia A., and Piet A. J. de Boer. "ZipA Is Required for Recruitment of FtsK, FtsQ, FtsL, and FtsN to the Septal Ring in Escherichia coli." Journal of Bacteriology 184, no. 9 (2002): 2552–56. http://dx.doi.org/10.1128/jb.184.9.2552-2556.2002.
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ABSTRACT The septal ring in Escherichia coli consists of at least nine essential gene products whose order of assembly resembles a mostly linear dependency pathway: FtsA and ZipA directly bind FtsZ polymers at the prospective division site, followed by the sequential addition of FtsK, FtsQ, FtsL, FtsW, FtsI, and FtsN. Recruitment of FtsK and all downstream components requires the prior localization of FtsA. Here we show that recruitment of FtsK, FtsQ, FtsL, and FtsN equally requires ZipA. The results imply that association of both FtsA and ZipA with FtsZ polymers is needed for further maturation of the nascent organelle.
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Chen, Joseph C., David S. Weiss, Jean-Marc Ghigo, and Jon Beckwith. "Septal Localization of FtsQ, an Essential Cell Division Protein in Escherichia coli." Journal of Bacteriology 181, no. 2 (1999): 521–30. http://dx.doi.org/10.1128/jb.181.2.521-530.1999.
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ABSTRACT Septation in Escherichia coli requires several gene products. One of these, FtsQ, is a simple bitopic membrane protein with a short cytoplasmic N terminus, a membrane-spanning segment, and a periplasmic domain. We have constructed a merodiploid strain that expresses both FtsQ and the fusion protein green fluorescent protein (GFP)-FtsQ from single-copy chromosomal genes. The gfp-ftsQgene complements a null mutation in ftsQ. Fluorescence microscopy revealed that GFP-FtsQ localizes to the division site. Replacing the cytoplasmic and transmembrane domains of FtsQ with alternative membrane anchors did not prevent the localization of the GFP fusion protein, while replacing the periplasmic domain did, suggesting that the periplasmic domain is necessary and sufficient for septal targeting. GFP-FtsQ localization to the septum depended on the cell division proteins FtsZ and FtsA, which are cytoplasmic, but not on FtsL and FtsI, which are bitopic membrane proteins with comparatively large periplasmic domains. In addition, the septal localization of ZipA apparently did not require functional FtsQ. Our results indicate that FtsQ is an intermediate recruit to the division site.
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Eberhardt, Christian, Lars Kuerschner та David S. Weiss. "Probing the Catalytic Activity of a Cell Division-Specific Transpeptidase In Vivo with β-Lactams". Journal of Bacteriology 185, № 13 (2003): 3726–34. http://dx.doi.org/10.1128/jb.185.13.3726-3734.2003.
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ABSTRACT Penicillin-binding protein 3 (PBP3; also called FtsI) is a transpeptidase that catalyzes cross-linking of the peptidoglycan cell wall in the division septum of Escherichia coli. To determine whether the catalytic activity of PBP3 is activated during division, we assayed acylation of PBP3 with three β-lactams (cephalexin, aztreonam, and piperacillin) in growing cells. Acylation of PBP3 with cephalexin, but not aztreonam or piperacillin, appeared to be stimulated by cell division. Specifically, cephalexin acylated PBP3 about 50% faster in a population of dividing cells than in a population of filamentous cells in which division was inhibited by inactivation or depletion of FtsZ, FtsA, FtsQ, FtsW, or FtsN. However, in a simpler in vitro system using isolated membranes, acylation with cephalexin was not impaired by depletion of FtsW or FtsN. A conflicting previous report that the ftsA3(Ts) allele interferes with acylation of PBP3 was found to be due to the presence of a thermolabile PBP3 in the strain used in that study. The new findings presented here are discussed in light of the hypothesis that the catalytic activity of PBP3 is stimulated by interaction(s) with other division proteins. We suggest that there might be allosteric activation of substrate binding.
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Park, Kyung-Tae, Sebastien Pichoff, Shishen Du, and Joe Lutkenhaus. "FtsA acts through FtsW to promote cell wall synthesis during cell division in Escherichia coli." Proceedings of the National Academy of Sciences 118, no. 35 (2021): e2107210118. http://dx.doi.org/10.1073/pnas.2107210118.
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In Escherichia coli, FtsQLB is required to recruit the essential septal peptidoglycan (sPG) synthase FtsWI to FtsA, which tethers FtsZ filaments to the membrane. The arrival of FtsN switches FtsQLB in the periplasm and FtsA in the cytoplasm from a recruitment role to active forms that synergize to activate FtsWI. Genetic evidence indicates that the active form of FtsQLB has an altered conformation with an exposed domain of FtsL that acts on FtsI to activate FtsW. However, how FtsA contributes to the activation of FtsW is not clear, as it could promote the conformational change in FtsQLB or act directly on FtsW. Here, we show that the overexpression of an activated FtsA (FtsA*) bypasses FtsQ, indicating it can compensate for FtsQ’s recruitment function. Consistent with this, FtsA* also rescued FtsL and FtsB mutants deficient in FtsW recruitment. FtsA* also rescued an FtsL mutant unable to deliver the periplasmic signal from FtsN, consistent with FtsA* acting on FtsW. In support of this, an FtsW mutant was isolated that was rescued by an activated FtsQLB but not by FtsA*, indicating it was specifically defective in activation by FtsA. Our results suggest that in response to FtsN, the active form of FtsA acts on FtsW in the cytoplasm and synergizes with the active form of FtsQLB acting on FtsI in the periplasm to activate FtsWI to carry out sPG synthesis.
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NURZHAN, SHILMURZAYEV, TURSYNBAYEV ABYLAY, CHANDAN PAL SINGH, AN IGOR, and OMIRKUL BAISEYIT. "DEVELOPMENT OF ANALYSIS AND EVALUATION SKILLS IN A PHYSICS TEACHING THROUGH PRACTICAL WORK." International Journal Of Multidisciplinary Research And Studies 05, no. 06 (2022): 01–15. http://dx.doi.org/10.33826/ijmras/v05i06.3.
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This research paper addresses how to enhance analysis and evaluation skills in physics lessons using practical or lab work. Analysis and evaluation skills are very important for high school science students that can lead to developing critical thinking also, especially where English is a third-level language. Practicality plays a very important role in understanding complex and confusing topics, so integrating physics lessons with teaching practical work can improve a significant level of understanding. We have researched and analyzed data after applying for practical work in regular lessons and conclusions based on three factors, (a) performance on term-end examinations (b) Attitude towards physics/science (c) Awareness, and more enrolment in physics in grade 12. We have used the Form Two Students Attitude Questionnaire, and FTSAQ to get data to justify the research question. The study was performed on students in grade -11(NIS, Chem, and Bio, Shymkent, Kazakhstan. Respondents were divided into two groups: (1) the Control group and (2) the Experimental Group. Control group were taught in term 3 with all possible teaching methodologies (interactive lesson, Assessment, Inquiry-based learning) except practical work but in another group, practical work was included to analyze the effectiveness of integration. Results are exactly as assumed; a group with integration of practical work was high in results, attitude toward physics, and a more interesting subject with less confusion and more problem-solving skills. The approach to solving problems and connecting concepts with real life is increased significantly.
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Dr., SHILMURZAYEV NURZHAN, TURSYNBAYEV ABYLAY Dr., CHANDAN PAL SINGH Dr., AN IGOR Dr., and OMIRKUL BAISEYIT5 Dr. "DEVELOPMENT OF ANALYSIS AND EVALUATION SKILLS IN A PHYSICS TEACHING THROUGH PRACTICAL WORK." International Journal Of Multidisciplinary Research And Studies 05, no. 06 (2022): 19–33. https://doi.org/10.33826/ijmras/v05i06.3.
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This research paper addresses how to enhance analysis and evaluation skills in physics lessons using practical or lab work. Analysis and evaluation skills are very important for high school science students that can lead to developing critical thinking also, especially where English is a third-level language. Practicality plays a very important role in understanding complex and confusing topics, so integrating physics lessons with teaching practical work can improve a significant level of understanding. We have researched and analyzed data after applying for practical work in regular lessons and conclusions based on three factors, (a) performance on term-end examinations (b) Attitude towards physics/science (c) Awareness, and more enrolment in physics in grade 12. We have used the Form Two Students Attitude Questionnaire, and FTSAQ to get data to justify the research question. The study was performed on students in grade -11(NIS, Chem, and Bio, Shymkent, Kazakhstan. Respondents were divided into two groups: (1) the Control group and (2) the Experimental Group. Control group were taught in term 3 with all possible teaching methodologies (interactive lesson, Assessment, Inquiry-based learning) except practical work but in another group, practical work was included to analyze the effectiveness of integration. Results are exactly as assumed; the group with the integration of practical work was high in results, attitude towards physics, and more interesting subject with less confusion and more problem-solving skills. The approach to solving problems and connecting concepts with real life is increased significantly.
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Vinella, Daniel, Michael Cashel, and Richard D’Ari. "Selected Amplification of the Cell Division Genes ftsQ-ftsA-ftsZ in Escherichia coli." Genetics 156, no. 4 (2000): 1483–92. http://dx.doi.org/10.1093/genetics/156.4.1483.
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Abstract Rapidly growing Escherichia coli is unable to divide in the presence of the antibiotic mecillinam, whose direct target is penicillin-binding protein 2 (PBP2), responsible for the elongation of the cylindrical portion of the cell wall. Division can be restored in the absence of PBP2 activity by increasing the concentration of the cell division proteins FtsQ, FtsA, and FtsZ. We tried to identify regulators of the ftsQ-ftsA-ftsZ operon among mecillinam-resistant mutants, which include strains overexpressing these genes. By insertional mutagenesis with mini-Tn10 elements, we selected for insertions that conferred mecillinam resistance. Among 15 such mutants, 7 suppressed the thermosensitivity of the ftsZ84(Ts) mutant, strongly suggesting that they had increased FtsZ activity. In all 7 cases, however, the mutants resulted from a duplication of the ftsQAZ region. These duplications seemed to result from multiple events, suggesting that no simple insertional inactivation can result in a mutant with sufficiently amplified ftsQAZ expression to confer mecillinam resistance. The structure of the duplications suggests a general method for constructing directed duplications of precise sequences.
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Marbouty, Martial, Khalil Mazouni, Cyril Saguez, Corinne Cassier-Chauvat, and Franck Chauvat. "Characterization of the Synechocystis Strain PCC 6803 Penicillin-Binding Proteins and Cytokinetic Proteins FtsQ and FtsW and Their Network of Interactions with ZipN." Journal of Bacteriology 191, no. 16 (2009): 5123–33. http://dx.doi.org/10.1128/jb.00620-09.
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ABSTRACT Because very little is known about cell division in noncylindrical bacteria and cyanobacteria, we investigated 10 putative cytokinetic proteins in the unicellular spherical cyanobacterium Synechocystis strain PCC 6803. Concerning the eight penicillin-binding proteins (PBPs), which define three classes, we found that Synechocystis can survive in the absence of one but not two PBPs of either class A or class C, whereas the unique class B PBP (also termed FtsI) is indispensable. Furthermore, we showed that all three classes of PBPs are required for normal cell size. Similarly, the putative FtsQ and FtsW proteins appeared to be required for viability and normal cell size. We also used a suitable bacterial two-hybrid system to characterize the interaction web among the eight PBPs, FtsQ, and FtsW, as well as ZipN, the crucial FtsZ partner that occurs only in cyanobacteria and plant chloroplasts. We showed that FtsI, FtsQ, and ZipN are self-interacting proteins and that both FtsI and FtsQ interact with class A PBPs, as well as with ZipN. Collectively, these findings indicate that ZipN, in interacting with FtsZ and both FtsI and FtQ, plays a similar role to the Escherichia coli FtsA protein, which is missing in cyanobacteria and chloroplasts.
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Reddy, Manjula. "Role of FtsEX in Cell Division of Escherichia coli: Viability of ftsEX Mutants Is Dependent on Functional SufI or High Osmotic Strength." Journal of Bacteriology 189, no. 1 (2006): 98–108. http://dx.doi.org/10.1128/jb.01347-06.
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ABSTRACT In Escherichia coli, at least 12 proteins, FtsZ, ZipA, FtsA, FtsE/X, FtsK, FtsQ, FtsL, FtsB, FtsW, FtsI, FtsN, and AmiC, are known to localize to the septal ring in an interdependent and sequential pathway to coordinate the septum formation at the midcell. The FtsEX complex is the latest recruit of this pathway, and unlike other division proteins, it is shown to be essential only on low-salt media. In this study, it is shown that ftsEX null mutations are not only salt remedial but also osmoremedial, which suggests that FtsEX may not be involved in salt transport as previously thought. Increased coexpression of cell division proteins FtsQ-FtsA-FtsZ or FtsN alone restored the growth defects of ftsEX mutants. ftsEX deletion exacerbated the defects of most of the mutants affected in Z ring localization and septal assembly; however, the ftsZ84 allele was a weak suppressor of ftsEX. The viability of ftsEX mutants in high-osmolarity conditions was shown to be dependent on the presence of a periplasmic protein, SufI, a substrate of twin-arginine translocase. In addition, SufI in multiple copies could substitute for the functions of FtsEX. Taken together, these results suggest that FtsE and FtsX are absolutely required for the process of cell division in conditions of low osmotic strength for the stability of the septal ring assembly and that, during high-osmolarity conditions, the FtsEX and SufI functions are redundant for this essential process.
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Yi, Qing-Ming, Susan Rockenbach, John E. Ward, and Joe Lutkenhaus. "Structure and expression of the cell division genes ftsQ, ftsA and ftsZ." Journal of Molecular Biology 184, no. 3 (1985): 399–412. http://dx.doi.org/10.1016/0022-2836(85)90290-6.
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Goehring, Nathan W., Carine Robichon, and Jon Beckwith. "Role for the Nonessential N Terminus of FtsN in Divisome Assembly." Journal of Bacteriology 189, no. 2 (2006): 646–49. http://dx.doi.org/10.1128/jb.00992-06.
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ABSTRACT FtsN, the last essential protein in the cell division localization hierarchy in Escherichia coli, has several peculiar characteristics, suggesting that it has a unique role in the division process despite the fact that it is conserved in only a subset of bacteria. In addition to suppressing temperature-sensitive mutations in ftsA, ftsK, ftsQ, and ftsI, overexpression of FtsN can compensate for a complete lack of FtsK in the cell. We examined the requirements for this phenomenon. We found that the N-terminal terminal region (cytoplasmic and transmembrane domains) is critical for suppression, while the C-terminal murein-binding domain is dispensable. Our results further suggest that FtsN and FtsK act cooperatively to stabilize the divisome.
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Martin, Miriam E., Michael J. Trimble, and Yves V. Brun. "Cell cycle-dependent abundance, stability and localization of FtsA and FtsQ in Caulobacter crescentus." Molecular Microbiology 54, no. 1 (2004): 60–74. http://dx.doi.org/10.1111/j.1365-2958.2004.04251.x.
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Schmidt, Kari L., Nicholas D. Peterson, Ryan J. Kustusch, et al. "A Predicted ABC Transporter, FtsEX, Is Needed for Cell Division in Escherichia coli." Journal of Bacteriology 186, no. 3 (2004): 785–93. http://dx.doi.org/10.1128/jb.186.3.785-793.2004.
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ABSTRACT FtsE and FtsX have homology to the ABC transporter superfamily of proteins and appear to be widely conserved among bacteria. Early work implicated FtsEX in cell division in Escherichia coli, but this was subsequently challenged, in part because the division defects in ftsEX mutants are often salt remedial. Strain RG60 has an ftsE::kan null mutation that is polar onto ftsX. RG60 is mildly filamentous when grown in standard Luria-Bertani medium (LB), which contains 1% NaCl, but upon shift to LB with no NaCl growth and division stop. We found that FtsN localizes to potential division sites, albeit poorly, in RG60 grown in LB with 1% NaCl. We also found that in wild-type E. coli both FtsE and FtsX localize to the division site. Localization of FtsX was studied in detail and appeared to require FtsZ, FtsA, and ZipA, but not the downstream division proteins FtsK, FtsQ, FtsL, and FtsI. Consistent with this, in media lacking salt, FtsA and ZipA localized independently of FtsEX, but the downstream proteins did not. Finally, in the absence of salt, cells depleted of FtsEX stopped dividing before any change in growth rate (mass increase) was apparent. We conclude that FtsEX participates directly in the process of cell division and is important for assembly or stability of the septal ring, especially in salt-free media.
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Di Lallo, G., M. Fagioli, D. Barionovi, P. Ghelardini, and L. Paolozzi. "Use of a two-hybrid assay to study the assembly of a complex multicomponent protein machinery: bacterial septosome differentiation." Microbiology 149, no. 12 (2003): 3353–59. http://dx.doi.org/10.1099/mic.0.26580-0.
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The ability of each of the nine Escherichia coli division proteins (FtsZ, FtsA, ZipA, FtsK, FtsQ, FtsL, FtsW, FtsI, FtsN) to interact with itself and with each of the remaining eight proteins was studied in 43 possible combinations of protein pairs by the two-hybrid system previously developed by the authors' group. Once the presumed interactions between the division proteins were determined, a model showing their temporal sequence of assembly was developed. This model agrees with that developed by other authors, based on the co-localization sequence in the septum of the division proteins fused with GFP. In addition, this paper shows that the authors' assay, which has already proved to be very versatile in the study of prokaryotic and eukaryotic protein interaction, is also a powerful instrument for an in vivo study of the interaction and assembly of proteins, as in the case of septum division formation.
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Mercer, Keri L. N., and David S. Weiss. "The Escherichia coli Cell Division Protein FtsW Is Required To Recruit Its Cognate Transpeptidase, FtsI (PBP3), to the Division Site." Journal of Bacteriology 184, no. 4 (2002): 904–12. http://dx.doi.org/10.1128/jb.184.4.904-912.2002.
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ABSTRACT The bacterial cell division protein FtsW has been suggested to perform two functions: stabilize the FtsZ cytokinetic ring, and facilitate septal peptidoglycan synthesis by the transpeptidase FtsI (penicillin-binding protein 3). We show here that depleting Escherichia coli cells of FtsW had little effect on the abundance of FtsZ rings but abrogated recruitment of FtsI to potential division sites. Analysis of FtsW localization confirmed and extended these results; septal localization of FtsW required FtsZ, FtsA, FtsQ, and FtsL but not FtsI. Thus, FtsW is a late recruit to the division site and is essential for subsequent recruitment of its cognate transpeptidase FtsI but not for stabilization of FtsZ rings. We suggest that a primary function of FtsW homologues—which are found in almost all bacteria and appear to work in conjunction with dedicated transpeptidases involved in division, elongation, or sporulation—is to recruit their cognate transpeptidases to the correct subcellular location.
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Descoteaux, A., and G. R. Drapeau. "Regulation of cell division in Escherichia coli K-12: probable interactions among proteins FtsQ, FtsA, and FtsZ." Journal of Bacteriology 169, no. 5 (1987): 1938–42. http://dx.doi.org/10.1128/jb.169.5.1938-1942.1987.
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Yeung, Leo W. Y., and Scott A. Mabury. "Are humans exposed to increasing amounts of unidentified organofluorine?" Environmental Chemistry 13, no. 1 (2016): 102. http://dx.doi.org/10.1071/en15041.
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Environmental context Polyfluorinated substances are anthropogenic chemicals that have been widely used in several industrial and commercial applications. Analysis of human plasma samples collected from Münster in Germany revealed, since the year 2000, increasing amounts and proportion of unidentified organofluorines. The increasing trend of unidentified organofluorines in plasma samples suggests that humans are being exposed to new and unidentified fluorinated products. Abstract Samples of human plasma (n=122) from two German cities (collected in 1982–2009, excluding 1994) and whole blood (n=47) from seven Chinese cities (collected in 2004) were analysed for 52 polyfluoroalkyl/perfluoroalkyl substances (PFASs) using LC-MS/MS. Quantifiable PFASs included some newly identified and commercially available chemicals PFPAs, PFPiAs, FTSAs, PAPs and di-SAmPAP, metabolites of fluorotelomer-based products (FTCAs/FTUCAs), PFCAs, PFSAs, FASAs and FOSAAs. The blood samples were also analysed for extractable organofluorine (EOF) using total organofluorine combustion ion chromatography (TOF-CIC). Seven more PFASs (C7 and C10 PFSAs, FOSAA, MeFOSAA, EtFOSAA, C13 PFCA and 8:2 FTSA) were detected in the Chinese samples than had been previously reported. For the German samples, PFHpS, FOSA, MeFOSA, EtFOSA, FTSAs (6:2, 8:2), PFPAs (C6, C8) and PFPiAs (C6/C6, C6/C8, C8/C8) were additional chemicals identified that were not measured in the earlier studies. Those newly identified and commercially available PFASs were either at trace levels (pg mL–1) or not detected. A mass balance of fluorine between quantifiable PFAS and EOF in the Chinese samples indicated quantifiable PFASs accounted for 31–86% of EOF. For the German samples, the quantifiable PFAS accounted for 52–100% and 57–100% of EOF in Münster and Halle samples respectively. After the year 2000, an increasing amount and proportion of unidentified organofluorine were observed in Münster samples. The increasing trend of unidentified organofluorine in plasma samples suggested humans are being exposed to new and unidentified fluorinated products.
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Weiss, David S., Joseph C. Chen, Jean-Marc Ghigo, Dana Boyd, and Jon Beckwith. "Localization of FtsI (PBP3) to the Septal Ring Requires Its Membrane Anchor, the Z Ring, FtsA, FtsQ, and FtsL." Journal of Bacteriology 181, no. 2 (1999): 508–20. http://dx.doi.org/10.1128/jb.181.2.508-520.1999.
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ABSTRACT Assembly of the division septum in bacteria is mediated by several proteins that localize to the division site. One of these, FtsI (also called penicillin-binding protein 3) of Escherichia coli, consists of a short cytoplasmic domain, a single membrane-spanning segment, and a large periplasmic domain that encodes a transpeptidase activity involved in synthesis of septal peptidoglycan. We have constructed a merodiploid strain with a wild-type copy offtsI at the normal chromosomal locus and a genetic fusion of ftsI to the green fluorescent protein (gfp) at the lambda attachment site. gfp-ftsI was expressed at physiologically appropriate levels under control of a regulatable promoter. Consistent with previous results based on immunofluorescence microscopy GFP-FtsI localized to the division site during the later stages of cell growth and throughout septation. Localization of GFP-FtsI to the cell pole(s) was not observed unless the protein was overproduced about 10-fold. Membrane anchor alterations shown previously to impair division but not membrane insertion or transpeptidase activity were found to interfere with localization of GFP-FtsI to the division site. In contrast, GFP-FtsI localized well in the presence of β-lactam antibiotics that inhibit the transpeptidase activity of FtsI. Septal localization depended upon every other division protein tested (FtsZ, FtsA, FtsQ, and FtsL). We conclude that FtsI is a late recruit to the division site, and that its localization depends on an intact membrane anchor.
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Thakur, Payal, Vinoj Gopalakrishnan, Priya Saxena, et al. "Influence of Copper on Oleidesulfovibrio alaskensis G20 Biofilm Formation." Microorganisms 12, no. 9 (2024): 1747. http://dx.doi.org/10.3390/microorganisms12091747.
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Copper is known to have toxic effects on bacterial growth. This study aimed to determine the influence of copper ions on Oleidesulfovibrio alaskensis G20 biofilm formation in a lactate-C medium supplemented with variable copper ion concentrations. OA G20, when grown in media supplemented with high copper ion concentrations of 5, 15, and 30 µM, exhibited inhibited growth in its planktonic state. Conversely, under similar copper concentrations, OA G20 demonstrated enhanced biofilm formation on glass coupons. Microscopic studies revealed that biofilms exposed to copper stress demonstrated a change in cellular morphology and more accumulation of carbohydrates and proteins than controls. Consistent with these findings, sulfur (dsrA, dsrB, sat, aprA) and electron transport (NiFeSe, NiFe, ldh, cyt3) genes, polysaccharide synthesis (poI), and genes involved in stress response (sodB) were significantly upregulated in copper-induced biofilms, while genes (ftsZ, ftsA, ftsQ) related to cellular division were negatively regulated compared to controls. These results indicate that the presence of copper ions triggers alterations in cellular morphology and gene expression levels in OA G20, impacting cell attachment and EPS production. This adaptation, characterized by increased biofilm formation, represents a crucial strategy employed by OA G20 to resist metal ion stress.
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Buddelmeijer, Nienke, Mirjam E. G. Aarsman, Arend H. J. Kolk, Miguel Vicente, and Nanne Nanninga. "Localization of Cell Division Protein FtsQ by Immunofluorescence Microscopy in Dividing and Nondividing Cells ofEscherichia coli." Journal of Bacteriology 180, no. 23 (1998): 6107–16. http://dx.doi.org/10.1128/jb.180.23.6107-6116.1998.
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ABSTRACT The localization of cell division protein FtsQ in Escherichia coli wild-type cells was studied by immunofluorescence microscopy with specific monoclonal antibodies. FtsQ could be localized to the division site in constricting cells. FtsQ could also localize to the division site in ftsQ1(Ts) cells grown at the permissive temperature. A hybrid protein in which the cytoplasmic domain and the transmembrane domain were derived from the γ form of penicillin-binding protein 1B and the periplasmic domain was derived from FtsQ was also able to localize to the division site. This result indicates that the periplasmic domain of FtsQ determines the localization of FtsQ, as has also been concluded by others for the periplasmic domain of FtsN. Noncentral FtsQ foci were found in the area of the cell where the nucleoid resides and were therefore assumed to represent sites where the FtsQ protein is synthesized and simultaneously inserted into the cytoplasmic membrane.
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Scheffers, Dirk-Jan, Carine Robichon, Gert Jan Haan, et al. "Contribution of the FtsQ Transmembrane Segment to Localization to the Cell Division Site." Journal of Bacteriology 189, no. 20 (2007): 7273–80. http://dx.doi.org/10.1128/jb.00723-07.
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ABSTRACT The Escherichia coli cell division protein FtsQ is a central component of the divisome. FtsQ is a bitopic membrane protein with a large C-terminal periplasmic domain. In this work we investigated the role of the transmembrane segment (TMS) that anchors FtsQ in the cytoplasmic membrane. A set of TMS mutants was made and analyzed for the ability to complement an ftsQ mutant. Study of the various steps involved in FtsQ biogenesis revealed that one mutant (L29/32R;V38P) failed to functionally insert into the membrane, whereas another mutant (L29/32R) was correctly assembled and interacted with FtsB and FtsL but failed to localize efficiently to the cell division site. Our results indicate that the FtsQ TMS plays a role in FtsQ localization to the division site.
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Corbin, Brian D., Brett Geissler, Mahalakshmi Sadasivam, and William Margolin. "Z-Ring-Independent Interaction between a Subdomain of FtsA and Late Septation Proteins as Revealed by a Polar Recruitment Assay." Journal of Bacteriology 186, no. 22 (2004): 7736–44. http://dx.doi.org/10.1128/jb.186.22.7736-7744.2004.
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ABSTRACT FtsA, a member of the ATPase superfamily that includes actin and bacterial actin homologs, is essential for cell division of Escherichia coli and is recruited to the Z ring. In turn, recruitment of later essential division proteins to the Z ring is dependent on FtsA. In a polar recruitment assay, we found that FtsA can recruit at least two late proteins, FtsI and FtsN, to the cell poles independently of Z rings. Moreover, a unique structural domain of FtsA, subdomain 1c, which is divergent in the other ATPase superfamily members, is sufficient for this recruitment but not required for the ability of FtsA to localize to Z rings. Surprisingly, targeting the 1c subdomain to the Z ring by fusing it to FtsZ could partially suppress a thermosensitive ftsA mutation. These results suggest that subdomain 1c of FtsA is a completely independent functional domain with an important role in interacting with a septation protein subassembly.
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Trespidi, Gabriele, Viola Camilla Scoffone, Giulia Barbieri, Giovanna Riccardi, Edda De Rossi, and Silvia Buroni. "Molecular Characterization of the Burkholderia cenocepacia dcw Operon and FtsZ Interactors as New Targets for Novel Antimicrobial Design." Antibiotics 9, no. 12 (2020): 841. http://dx.doi.org/10.3390/antibiotics9120841.
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The worldwide spread of antimicrobial resistance highlights the need of new druggable cellular targets. The increasing knowledge of bacterial cell division suggested the potentiality of this pathway as a pool of alternative drug targets, mainly based on the essentiality of these proteins, as well as on the divergence from their eukaryotic counterparts. People suffering from cystic fibrosis are particularly challenged by the lack of antibiotic alternatives. Among the opportunistic pathogens that colonize the lungs of these patients, Burkholderia cenocepacia is a well-known multi-drug resistant bacterium, particularly difficult to treat. Here we describe the organization of its division cell wall (dcw) cluster: we found that 15 genes of the dcw operon can be transcribed as a polycistronic mRNA from mraZ to ftsZ and that its transcription is under the control of a strong promoter regulated by MraZ. B. cenocepacia J2315 FtsZ was also shown to interact with the other components of the divisome machinery, with a few differences respect to other bacteria, such as the direct interaction with FtsQ. Using an in vitro sedimentation assay, we validated the role of SulA as FtsZ inhibitor, and the roles of FtsA and ZipA as tethers of FtsZ polymers. Together our results pave the way for future antimicrobial design based on the divisome as pool of antibiotic cellular targets.
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Karimova, Gouzel, Carine Robichon, and Daniel Ladant. "Characterization of YmgF, a 72-Residue Inner Membrane Protein That Associates with the Escherichia coli Cell Division Machinery." Journal of Bacteriology 191, no. 1 (2008): 333–46. http://dx.doi.org/10.1128/jb.00331-08.
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ABSTRACT Formation of the Escherichia coli division septum is catalyzed by a number of essential proteins (named Fts) that assemble into a ring-like structure at the future division site. Many of these Fts proteins are intrinsic transmembrane proteins whose functions are largely unknown. In the present study, we attempted to identify a novel putative component(s) of the E. coli cell division machinery by searching for proteins that could interact with known Fts proteins. To do that, we used a bacterial two-hybrid system based on interaction-mediated reconstitution of a cyclic AMP (cAMP) signaling cascade to perform a library screening in order to find putative partners of E. coli cell division protein FtsL. Here we report the characterization of YmgF, a 72-residue integral membrane protein of unknown function that was found to associate with many E. coli cell division proteins and to localize to the E. coli division septum in an FtsZ-, FtsA-, FtsQ-, and FtsN-dependent manner. Although YmgF was previously shown to be not essential for cell viability, we found that when overexpressed, YmgF was able to overcome the thermosensitive phenotype of the ftsQ1(Ts) mutation and restore its viability under low-osmolarity conditions. Our results suggest that YmgF might be a novel component of the E. coli cell division machinery.
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Uehara, Tsuyoshi, та James T. Park. "Role of the Murein Precursor UDP-N-Acetylmuramyl-l-Ala-γ-d-Glu- meso-Diaminopimelic Acid-d-Ala-d-Ala in Repression of β-Lactamase Induction in Cell Division Mutants". Journal of Bacteriology 184, № 15 (2002): 4233–39. http://dx.doi.org/10.1128/jb.184.15.4233-4239.2002.
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ABSTRACT Certain β-lactam antibiotics induce the chromosomal ampC β-lactamase of many gram-negative bacteria. The natural inducer, though not yet unequivocally identified, is a cell wall breakdown product which enters the cell via the AmpG permease component of the murein recycling pathway. Surprisingly, it has been reported that β-lactamase is not induced by cefoxitin in the absence of FtsZ, which is required for cell division, or in the absence of penicillin-binding protein 2 (PBP2), which is required for cell elongation. Since these results remain unexplained, we examined an ftsZ mutant and other cell division mutants (ftsA, ftsQ, and ftsI) and a PBP2 mutant for induction of β-lactamase. In all mutants, β-lactamase was not induced by cefoxitin, which confirms the initial reports. The murein precursor, UDP-N-acetylmuramyl-l-Ala-γ-d-Glu-meso-diaminopimelic acid-d-Ala-d-Ala (UDP-MurNAc-pentapeptide), has been shown to serve as a corepressor with AmpR to repress β-lactamase expression in vitro. Our results suggest that β-lactamase is not induced because the fts mutants contain a greatly increased amount of corepressor which the inducer cannot displace. In the PBP2(Ts) mutant, in addition to accumulation of corepressor, cell wall turnover and recycling were greatly reduced so that little or no inducer was available. Hence, in both cases, a high ratio of repressor to inducer presumably prevents induction.
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Du, Shishen, Sebastien Pichoff, and Joe Lutkenhaus. "FtsEX acts on FtsA to regulate divisome assembly and activity." Proceedings of the National Academy of Sciences 113, no. 34 (2016): E5052—E5061. http://dx.doi.org/10.1073/pnas.1606656113.
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Bacterial cell division is driven by the divisome, a ring-shaped protein complex organized by the bacterial tubulin homolog FtsZ. Although most of the division proteins inEscherichia colihave been identified, how they assemble into the divisome and synthesize the septum remains poorly understood. Recent studies suggest that the bacterial actin homolog FtsA plays a critical role in divisome assembly and acts synergistically with the FtsQLB complex to regulate the activity of the divisome. FtsEX, an ATP-binding cassette transporter-like complex, is also necessary for divisome assembly and inhibits division when its ATPase activity is inactivated. However, its role in division is not clear. Here, we find that FtsEX acts on FtsA to regulate both divisome assembly and activity. FtsX interacts with FtsA and this interaction is required for divisome assembly and inhibition of divisome function by ATPase mutants of FtsEX. Our results suggest that FtsEX antagonizes FtsA polymerization to promote divisome assembly and the ATPase mutants of FtsEX block divisome activity by locking FtsA in the inactive form or preventing FtsA from communicating with other divisome proteins. Because FtsEX is known to govern cell wall hydrolysis at the septum, our findings indicate that FtsEX acts on FtsA to promote divisome assembly and to coordinate cell wall synthesis and hydrolysis at the septum. Furthermore, our study provides evidence that FtsA mutants impaired for self-interaction are favored for division, and FtsW plays a critical role in divisome activation in addition to the FtsQLB complex.
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Glas, Marjolein, Eiso AB, Johan Hollander, Gregg Siegal, Joen Luirink, and Iwan de Esch. "Interrogating the Essential Bacterial Cell Division Protein FtsQ with Fragments Using Target Immobilized NMR Screening (TINS)." International Journal of Molecular Sciences 20, no. 15 (2019): 3684. http://dx.doi.org/10.3390/ijms20153684.
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The divisome is a large protein complex that regulates bacterial cell division and therefore represents an attractive target for novel antibacterial drugs. In this study, we report on the ligandability of FtsQ, which is considered a key component of the divisome. For this, the soluble periplasmic domain of Escherichia coli FtsQ was immobilized and used to screen a library of 1501 low molecular weight (< 300 Da), synthetic compounds for those that interact with the protein. A primary screen was performed using target immobilized NMR screening (TINS) and yielded 72 hits. Subsequently, these hits were validated in an orthogonal assay. At first, we aimed to do this using surface plasmon resonance (SPR), but the lack of positive control hampered optimization of the experiment. Alternatively, a two-dimensional heteronuclear single quantum coherence (HSQC) NMR spectrum of FtsQ was obtained and used to validate these hits by chemical shift perturbation (CSP) experiments. This resulted in the identification of three fragments with weak affinity for the periplasmic domain of FtsQ, arguing that the ligandability of FtsQ is low. While this indicates that developing high affinity ligands for FtsQ is far from straightforward, the identified hit fragments can help to further interrogate FtsQ interactions.
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Yao, Xuanli, Xiangfeng Wang, and Xin Xiang. "FHIP and FTS proteins are critical for dynein-mediated transport of early endosomes in Aspergillus." Molecular Biology of the Cell 25, no. 14 (2014): 2181–89. http://dx.doi.org/10.1091/mbc.e14-04-0873.
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The minus end–directed microtubule motor cytoplasmic dynein transports various cellular cargoes, including early endosomes, but how dynein binds to its cargo remains unclear. Recently fungal Hook homologues were found to link dynein to early endosomes for their transport. Here we identified FhipA in Aspergillus nidulans as a key player for HookA (A. nidulans Hook) function via a genome-wide screen for mutants defective in early-endosome distribution. The human homologue of FhipA, FHIP, is a protein in the previously discovered FTS/Hook/FHIP (FHF) complex, which contains, besides FHIP and Hook proteins, Fused Toes (FTS). Although this complex was not previously shown to be involved in dynein-mediated transport, we show here that loss of either FhipA or FtsA (A. nidulans FTS homologue) disrupts HookA–early endosome association and inhibits early endosome movement. Both FhipA and FtsA associate with early endosomes, and interestingly, while FtsA–early endosome association requires FhipA and HookA, FhipA–early endosome association is independent of HookA and FtsA. Thus FhipA is more directly linked to early endosomes than HookA and FtsA. However, in the absence of HookA or FtsA, FhipA protein level is significantly reduced. Our results indicate that all three proteins in the FtsA/HookA/FhipA complex are important for dynein-mediated early endosome movement.
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Ottolenghi, A. C., and J. A. Ayala. "Induction of a class I beta-lactamase from Citrobacter freundii in Escherichia coli requires active ftsZ but not ftsA or ftsQ products." Antimicrobial Agents and Chemotherapy 35, no. 11 (1991): 2359–65. http://dx.doi.org/10.1128/aac.35.11.2359.
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Jensen, S. O., L. S. Thompson, and E. J. Harry. "Cell Division in Bacillus subtilis: FtsZ and FtsA Association Is Z-Ring Independent, and FtsA Is Required for Efficient Midcell Z-Ring Assembly." Journal of Bacteriology 187, no. 18 (2005): 6536–44. http://dx.doi.org/10.1128/jb.187.18.6536-6544.2005.
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ABSTRACT The earliest stage in cell division in bacteria is the assembly of a Z ring at the division site at midcell. Other division proteins are also recruited to this site to orchestrate the septation process. FtsA is a cytosolic division protein that interacts directly with FtsZ. Its function remains unknown. It is generally believed that FtsA localization to the division site occurs immediately after Z-ring formation or concomitantly with it and that FtsA is responsible for recruiting the later-assembling membrane-bound division proteins to the division site. Here, we report the development of an in vivo chemical cross-linking assay to examine the association between FtsZ and FtsA in Bacillus subtilis cells. We subsequently use this assay in a synchronous cell cycle to show that these two proteins can interact prior to Z-ring formation. We further show that in a B. subtilis strain containing an ftsA deletion, FtsZ localized at regular intervals along the filament but the majority of Z rings were abnormal. FtsA in this organism is therefore critical for the efficient formation of functional Z rings. This is the first report of abnormal Z-ring formation resulting from the loss of a single septation protein. These results suggest that in this organism, and perhaps others, FtsA ensures recruitment of the membrane-bound division proteins by ensuring correct formation of the Z ring.
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Weiner, Barbara, Leo W. Y. Yeung, Erin B. Marchington, Lisa A. D'Agostino, and Scott A. Mabury. "Organic fluorine content in aqueous film forming foams (AFFFs) and biodegradation of the foam component 6 : 2 fluorotelomermercaptoalkylamido sulfonate (6 : 2 FTSAS)." Environmental Chemistry 10, no. 6 (2013): 486. http://dx.doi.org/10.1071/en13128.
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Environmental context Total organofluorine and known fluorosurfactants were quantified in 11 aqueous film forming foams (AFFFs) used to extinguish fires in Ontario, Canada, and one commercial AFFF product. By comparing the concentrations of known fluorosurfactants with the total organofluorine, less than 10% of the fluorosurfactants were identified in half of the samples. Our biodegradation experiment with one of the fluorosurfactants using waste-water treatment plant sludge showed that it was a potential source of perfluoroalkyl carboxylates, which are persistent in the environment. Abstract Eleven aqueous film forming foam (AFFF) samples that were used to extinguish fires in Ontario, Canada, and one commercial product, were analysed using a variety of analytical techniques to obtain structural information and quantities of organofluorine and known perfluoroalkyl and polyfluoroalkyl substances (PFASs). The NMR spectra of the foams distinguished the fluorosurfactants that were synthesised by either electrochemical fluorination or telomerisation. Total organofluorine content was quantified using total organofluorine–combustion ion chromatography (TOF-CIC), which revealed that the samples contained from 475 to 18 000µgFmL–1. The common AFFF component 6 : 2 fluorotelomermercaptoalkylamido sulfonate (FTSAS) was quantified by liquid chromatography tandem mass spectrometry (LC-MS/MS) together with perfluoroalkane sulfonates (PFSAs), perfluoroalkyl carboxylates (PFCAs) and fluorotelomer sulfonates (FTSAs); in five samples, 6 : 2 FTSAS was present in concentrations greater than 1000µgmL–1. By comparing the concentrations of these quantifiable fluorochemicals with the total organofluorine content, it was evident that in half of the AFFF samples, less than 10% of the fluorochemicals were identified; in two of the samples, perfluorooctane sulfonate (PFOS) accounted for ~50% of the total organofluorine content. Our degradation experiment with 6 : 2 FTSAS using waste-water treatment plant sludge showed that 6 : 2 FTSAS was a potential source of FTSAs, fluorotelomer alcohols and PFCAs in the environment.
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Fujiwara, Kei, and Hideki Taguchi. "Filamentous Morphology in GroE-Depleted Escherichia coli Induced by Impaired Folding of FtsE." Journal of Bacteriology 189, no. 16 (2007): 5860–66. http://dx.doi.org/10.1128/jb.00493-07.
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ABSTRACT The chaperonin GroE (GroEL and the cochaperonin GroES) is the only chaperone system that is essential for the viability of Escherichia coli. It is known that GroE-depleted cells exhibit a filamentous morphology, suggesting that GroE is required for the folding of proteins involved in cell division. Although previous studies, including proteome-wide analyses of GroE substrates, have suggested several targets of GroE in cell division, there is no direct in vivo evidence to identify which substrates exhibit obligate dependence on GroE for folding. Among the candidate substrates, we found that prior excess production of FtsE, a protein engaged in cell division, completely suppressed the filamentation of GroE-depleted E. coli. The GroE depletion led to a drastic decrease in FtsE, and the cells exhibited a known phenotype associated with impaired FtsE function. In the GroE-depleted filamentous cells, the localizations of FtsA and ZipA, both of which assemble with the FtsZ septal ring before FtsE, were normal, whereas FtsX, the interaction partner of FtsE, and FtsQ, which is recruited after FtsE, did not localize to the ring, suggesting that the decrease in FtsE is a cause of the filamentous morphology. Finally, a reconstituted cell-free translation system revealed that the folding of newly translated FtsE was stringently dependent on GroEL/GroES. Based on these findings, we concluded that FtsE is a target substrate of the GroE system in E. coli cell division.
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Zhang, Jing, Zongming Ren, and Meng Chen. "Immunotoxicity and Transcriptome Analyses of Zebrafish (Danio rerio) Embryos Exposed to 6:2 FTSA." Toxics 11, no. 5 (2023): 459. http://dx.doi.org/10.3390/toxics11050459.
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As a new alternative to perfluorooctane sulfonic acid (PFOS), 6:2 fluorotelomer sulfonic acid (6:2 FTSA) has been widely produced and used in recent years, and its concentration and frequency of detection in the aquatic environment and aquatic organisms are increasing. However, studies of its toxicity in aquatic biological systems are alarmingly scarce, and the relevant toxicological information needs to be improved. In this study, we investigated AB wild-type zebrafish (Danio rerio) embryos subjected to acute 6:2 FTSA exposure for immunotoxicity using immunoassays and transcriptomics. Immune indexes showed significant decreases in SOD and LZM activities, but no significant change in NO content. Other indexes (TNOS, iNOS, ACP, AKP activities, and MDA, IL-1β, TNF-α, NF-κB, TLR4 content) all showed significant increases. These results indicated that 6:2 FTSA induced oxidative stress and inflammatory responses in zebrafish embryos and exhibited immunotoxicity. Consistently, transcriptomics showed that genes involved in the MAPK, TLR and NOD-like receptor signaling pathways (hsp70, hsp701, stat1b, irf3, cxcl8b, map3k8, il1b, tnfa and nfkb) were significantly upregulated after 6:2 FTSA exposure, suggesting that 6:2 FTSA might induce immunotoxicity in zebrafish embryos through the TLR/NOD-MAPK pathway. The results of this study indicate that the safety of 6:2 FTSA should be examined further.
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Demesa-Castañeda, Alba V., David J. Pérez, César Millán-Pacheco, Armando Hernández-Mendoza, and Rodrigo Said Razo-Hernández. "Searching for New Antibacterial Compounds Against Staphylococcus aureus: A Computational Study on the Binding Between FtsZ and FtsA." Drugs and Drug Candidates 3, no. 4 (2024): 751–73. http://dx.doi.org/10.3390/ddc3040043.
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Background: Staphylococcus aureus is a pathogen that has become resistant to different antibiotics, which makes it a threat to human health. Although the first penicillin-resistant strain appeared in 1945, nowadays, there are just a few alternatives to fight it. To circumvent this issue, novel approaches to develop drugs to target proteins of the bacteria cytoskeleton, essential for bacteria’s binary fission, are being developed. FtsZ and FtsA are two proteins that are key for the initial stages of binary fission. On one side, FtsZ forms a polymeric circular structure called the Z ring; meanwhile, FtsA binds to the cell membrane and then anchors to the Z ring. According to the literature, this interaction occurs within the C-terminus domain of FtsZ, which is mainly disordered. Objective: In this work, we studied the binding of FtsZ to FtsA using computational chemistry tools to identify the interactions between the two proteins to further use this information for the search of potential protein-protein binding inhibitors (PPBIs). Methods: We made a bioinformatic analysis to obtain a representative sequence of FtsZ and FtsA of Staphylococcus aureus. With this information, we built homology models of the FtsZ to carry out the molecular docking with the FtsA. Furthermore, alanine scanning was conducted to identify the key residues forming the FtsZ–FtsA complex. Finally, we used this information to generate a pharmacophore model to carry out a virtual screening approach. Results: We identified the key residues forming the FtsZ-FtsA complex as well as five molecules with high potential as PPBIs.
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Taschner, P. E., P. G. Huls, E. Pas, and C. L. Woldringh. "Division behavior and shape changes in isogenic ftsZ, ftsQ, ftsA, pbpB, and ftsE cell division mutants of Escherichia coli during temperature shift experiments." Journal of Bacteriology 170, no. 4 (1988): 1533–40. http://dx.doi.org/10.1128/jb.170.4.1533-1540.1988.
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Chen, Joseph C., Michael Minev, and Jon Beckwith. "Analysis of ftsQ Mutant Alleles in Escherichia coli: Complementation, Septal Localization, and Recruitment of Downstream Cell Division Proteins." Journal of Bacteriology 184, no. 3 (2002): 695–705. http://dx.doi.org/10.1128/jb.184.3.695-705.2002.
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ABSTRACT FtsQ, a 276-amino-acid, bitopic membrane protein, is one of the nine proteins known to be essential for cell division in gram-negative bacterium Escherichia coli. To define residues in FtsQ critical for function, we performed random mutagenesis on the ftsQ gene and identified four alleles (ftsQ2, ftsQ6, ftsQ15, and ftsQ65) that fail to complement the ftsQ1(Ts) mutation at the restrictive temperature. Two of the mutant proteins, FtsQ6 and FtsQ15, are functional at lower temperatures but are unable to localize to the division site unless wild-type FtsQ is depleted, suggesting that they compete poorly with the wild-type protein for septal targeting. The other two mutants, FtsQ2 and FtsQ65, are nonfunctional at all temperatures tested and have dominant-negative effects when expressed in an ftsQ1(Ts) strain at the permissive temperature. FtsQ2 and FtsQ65 localize to the division site in the presence or absence of endogenous FtsQ, but they cannot recruit downstream cell division proteins, such as FtsL, to the septum. These results suggest that FtsQ2 and FtsQ65 compete efficiently for septal targeting but fail to promote the further assembly of the cell division machinery. Thus, we have separated the localization ability of FtsQ from its other functions, including recruitment of downstream cell division proteins, and are beginning to define regions of the protein responsible for these distinct capabilities.
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Bottos, Eric M., Ebtihal Y. AL-shabib, Dayton M. J. Shaw, et al. "Transcriptomic response of Gordonia sp. strain NB4-1Y when provided with 6:2 fluorotelomer sulfonamidoalkyl betaine or 6:2 fluorotelomer sulfonate as sole sulfur source." Biodegradation 31, no. 4-6 (2020): 407–22. http://dx.doi.org/10.1007/s10532-020-09917-8.
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Abstract Perfluoroalkyl and polyfluoroalkyl substances (PFAS) are environmental contaminants of concern. We previously described biodegradation of two PFAS that represent components and transformation products of aqueous film-forming foams (AFFF), 6:2 fluorotelomer sulfonamidoalkyl betaine (6:2 FTAB) and 6:2 fluorotelomer sulfonate (6:2 FTSA), by Gordonia sp. strain NB4-1Y. To identify genes involved in the breakdown of these compounds, the transcriptomic response of NB4-1Y was examined when grown on 6:2 FTAB, 6:2 FTSA, a non-fluorinated analog of 6:2 FTSA (1-octanesulfonate), or MgSO4, as sole sulfur source. Differentially expressed genes were identified as those with ± 1.5 log2-fold-differences (± 1.5 log2FD) in transcript abundances in pairwise comparisons. Transcriptomes of cells grown on 6:2 FTAB and 6:2 FTSA were most similar (7.9% of genes expressed ± 1.5 log2FD); however, several genes that were expressed in greater abundance in 6:2 FTAB treated cells compared to 6:2 FTSA treated cells were noted for their potential role in carbon–nitrogen bond cleavage in 6:2 FTAB. Responses to sulfur limitation were observed in 6:2 FTAB, 6:2 FTSA, and 1-octanesulfonate treatments, as 20 genes relating to global sulfate stress response were more highly expressed under these conditions compared to the MgSO4 treatment. More highly expressed oxygenase genes in 6:2 FTAB, 6:2 FTSA, and 1-octanesulfonate treatments were found to code for proteins with lower percent sulfur-containing amino acids compared to both the total proteome and to oxygenases showing decreased expression. This work identifies genetic targets for further characterization and will inform studies aimed at evaluating the biodegradation potential of environmental samples through applied genomics. Graphic Abstract
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Pilhofer, Martin, Kristina Rappl, Christina Eckl, et al. "Characterization and Evolution of Cell Division and Cell Wall Synthesis Genes in the Bacterial Phyla Verrucomicrobia, Lentisphaerae, Chlamydiae, and Planctomycetes and Phylogenetic Comparison with rRNA Genes." Journal of Bacteriology 190, no. 9 (2008): 3192–202. http://dx.doi.org/10.1128/jb.01797-07.
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ABSTRACT In the past, studies on the relationships of the bacterial phyla Planctomycetes, Chlamydiae, Lentisphaerae, and Verrucomicrobia using different phylogenetic markers have been controversial. Investigations based on 16S rRNA sequence analyses suggested a relationship of the four phyla, showing the branching order Planctomycetes, Chlamydiae, Verrucomicrobia/Lentisphaerae. Phylogenetic analyses of 23S rRNA genes in this study also support a monophyletic grouping and their branching order—this grouping is significant for understanding cell division, since the major bacterial cell division protein FtsZ is absent from members of two of the phyla Chlamydiae and Planctomycetes. In Verrucomicrobia, knowledge about cell division is mainly restricted to the recent report of ftsZ in the closely related genera Prosthecobacter and Verrucomicrobium. In this study, genes of the conserved division and cell wall (dcw) cluster (ddl, ftsQ, ftsA, and ftsZ) were characterized in all verrucomicrobial subdivisions (1 to 4) with cultivable representatives (1 to 4). Sequence analyses and transcriptional analyses in Verrucomicrobia and genome data analyses in Lentisphaerae suggested that cell division is based on FtsZ in all verrucomicrobial subdivisions and possibly also in the sister phylum Lentisphaerae. Comprehensive sequence analyses of available genome data for representatives of Verrucomicrobia, Lentisphaerae, Chlamydiae, and Planctomycetes strongly indicate that their last common ancestor possessed a conserved, ancestral type of dcw gene cluster and an FtsZ-based cell division mechanism. This implies that Planctomycetes and Chlamydiae may have shifted independently to a non-FtsZ-based cell division mechanism after their separate branchings from their last common ancestor with Verrucomicrobia.
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Pichoff, Sebastien, Shishen Du, and Joe Lutkenhaus. "Disruption of divisome assembly rescued by FtsN–FtsA interaction inEscherichia coli." Proceedings of the National Academy of Sciences 115, no. 29 (2018): E6855—E6862. http://dx.doi.org/10.1073/pnas.1806450115.
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Cell division requires the assembly of a protein complex called the divisome. The divisome assembles in a hierarchical manner, with FtsA functioning as a hub to connect the Z-ring with the rest of the divisome and FtsN arriving last to activate the machine to synthesize peptidoglycan. FtsEX arrives as the Z-ring forms and acts on FtsA to initiate recruitment of the other divisome components. In the absence of FtsEX, recruitment is blocked; however, a multitude of conditions allow FtsEX to be bypassed. Here, we find that all such FtsEX bypass conditions, as well as the bypass of FtsK, depend upon the interaction of FtsN with FtsA, which promotes the back-recruitment of the late components of the divisome. Furthermore, our results suggest that these bypass conditions enhance the weak interaction of FtsN with FtsA and its periplasmic partners so that the divisome proteins are brought to the Z-ring when the normal hierarchical pathway is disrupted.
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Berezuk, Alison M., Elyse J. Roach, Laura Seidel, Reggie Y. Lo, and Cezar M. Khursigara. "FtsA G50E mutant suppresses the essential requirement for FtsK during bacterial cell division in Escherichia coli." Canadian Journal of Microbiology 66, no. 4 (2020): 313–27. http://dx.doi.org/10.1139/cjm-2019-0493.
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In Escherichia coli, the N-terminal domain of the essential protein FtsK (FtsKN) is proposed to modulate septum formation through the formation of dynamic and essential protein interactions with both the Z-ring and late-stage division machinery. Using genomic mutagenesis, complementation analysis, and in vitro pull-down assays, we aimed to identify protein interaction partners of FtsK essential to its function during division. Here, we identified the cytoplasmic Z-ring membrane anchoring protein FtsA as a direct protein–protein interaction partner of FtsK. Random genomic mutagenesis of an ftsK temperature-sensitive strain of E. coli revealed an FtsA point mutation (G50E) that is able to fully restore normal cell growth and morphology, and further targeted site-directed mutagenesis of FtsA revealed several other point mutations capable of fully suppressing the essential requirement for functional FtsK. Together, this provides insight into a potential novel co-complex formed between these components during division and suggests FtsA may directly impact FtsK function.
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Kushwah, Vinita Chauhan, Ritika Chauhan, and Ram Kumar Dhaked. "Fluorescence Thermal Shift Assay based High Through-put Screening in Small Molecule Drug Discovery: A Brief Overview." Journal of Modern Biology and Drug Discovery 3 (May 28, 2024): 3. http://dx.doi.org/10.53964/jmbdd.2024003.
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Background: The Early drug discovery process was majorly phenotypic, lacking target specificity, and mechanism of action causing short and/or long-term toxicological impacts on the patients leading to several drug withdrawals from the market. Biochemical procedures and methods used in drug discovery and development make it lengthy in terms of time requiring approx. 15 years with the investment of billions of dollars for a new drug molecule. Technological advancement leading to the identification of protein structures through crystallography paved a new path toward targeted drug discovery. Objective: This paper provides a brief overview of the use of the fluorescence thermal shift assay (FTSA) as a high throughput (HTS) screening technique in small molecule drug discovery. Methods: The article discusses the principles of thermal denaturation techniques and how they are applied in drug discovery. It highlights the advantages of FTSA over other biophysical methods, including its label-free, cost-effective, and high HTS nature. The review also presents a comparative analysis of FTSA with techniques such as isothermal titration calorimetry (ITC), mass spectrometry (MS), nuclear magnetic resonance (NMR), surface plasmon resonance (SPR), X-ray crystallography(X-RD), circular dichroism (CD), and differential scanning calorimetry (DSC). Results: The FTSA technique is described as a powerful tool that addresses the limitations of larger protein requirements, specific assay development, and low HTS associated with other thermal denaturation methods. Unlike biochemical and cellular assays that measure secondary effects, FTSA directly estimates the binding of small molecules to target proteins. Conclusion: The review concludes that FTSA is the only biophysical method that satisfies the three key prerequisites of being label-free, cost-effective, and suitable for high HTS screening in small molecule drug discovery.
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Masters, Millicent, Trevor Paterson, Andrew G. Popplewell, Thomas Owen-Hughes, J. H. Pringle, and Kenneth J. Begg. "The effect of DnaA protein levels and the rate of initiation at oriC on transcription originating in the ftsQ and ftsA genes: In vivo experiments." Molecular and General Genetics MGG 216, no. 2-3 (1989): 475–83. http://dx.doi.org/10.1007/bf00334393.
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Krupka, Marcin, and William Margolin. "Unite to divide: Oligomerization of tubulin and actin homologs regulates initiation of bacterial cell division." F1000Research 7 (February 28, 2018): 235. http://dx.doi.org/10.12688/f1000research.13504.1.
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To generate two cells from one, bacteria such asEscherichia coliuse a complex of membrane-embedded proteins called the divisome that synthesize the division septum. The initial stage of cytokinesis requires a tubulin homolog, FtsZ, which forms polymers that treadmill around the cell circumference. The attachment of these polymers to the cytoplasmic membrane requires an actin homolog, FtsA, which also forms dynamic polymers that directly bind to FtsZ. Recent evidence indicates that FtsA and FtsZ regulate each other’s oligomeric state inE. colito control the progression of cytokinesis, including the recruitment of septum synthesis proteins. In this review, we focus on recent advances in our understanding of protein-protein association between FtsZ and FtsA in the initial stages of divisome function, mainly in the well-characterizedE. colisystem.
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Hale, Cynthia A., and Piet A. J. de Boer. "Recruitment of ZipA to the Septal Ring ofEscherichia coli Is Dependent on FtsZ and Independent of FtsA." Journal of Bacteriology 181, no. 1 (1999): 167–76. http://dx.doi.org/10.1128/jb.181.1.167-176.1999.
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ABSTRACT Cell division in prokaryotes is mediated by the septal ring. InEscherichia coli, this organelle consists of several essential division proteins, including FtsZ, FtsA, and ZipA. To gain more insight into how the structure is assembled, we studied the interdependence of FtsZ, FtsA, and ZipA localization using both immunofluorescence and Gfp tagging techniques. To this end, we constructed a set of strains allowing us to determine the cellular location of each of these three proteins in cells from which one of the other two had been specifically depleted. Our results show that ZipA fails to accumulate in a ring shape in the absence of FtsZ. Conversely, depletion of ZipA does not abolish formation of FtsZ rings but leads to a significant reduction in the number of rings per unit of cell mass. In addition, ZipA does not appear to require FtsA for assembly into the septal ring and vice versa. It is suggested that septal ring formation starts by assembly of the FtsZ ring, after which ZipA and FtsA join this structure in a mutually independent fashion through direct interactions with the FtsZ protein.
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El Najjar, Nina, Jihad El Andari, Christine Kaimer, Georg Fritz, Thomas C. Rösch, and Peter L. Graumann. "Single-Molecule Tracking of DNA Translocases inBacillus subtilisReveals Strikingly Different Dynamics of SftA, SpoIIIE, and FtsA." Applied and Environmental Microbiology 84, no. 8 (2018): e02610-17. http://dx.doi.org/10.1128/aem.02610-17.
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ABSTRACTLike many bacteria,Bacillus subtilispossesses two DNA translocases that affect chromosome segregation at different steps. Prior to septum closure, nonsegregated DNA is moved into opposite cell halves by SftA, while septum-entrapped DNA is rescued by SpoIIIE. We have used single-molecule fluorescence microscopy and tracking (SMT) experiments to describe the dynamics of the two different DNA translocases, the cell division protein FtsA and the glycolytic enzyme phosphofructokinase (PfkA), in real time. SMT revealed that about 30% of SftA molecules move through the cytosol, while a fraction of 70% is septum bound and static. In contrast, only 35% of FtsA molecules are static at midcell, while SpoIIIE molecules diffuse within the membrane and show no enrichment at the septum. Several lines of evidence suggest that FtsA plays a role in septal recruitment of SftA: anftsAdeletion results in a significant reduction in septal SftA recruitment and a decrease in the average dwell time of SftA molecules. FtsA can recruit SftA to the membrane in a heterologous eukaryotic system, suggesting that SftA may be partially recruited via FtsA. Therefore, SftA is a component of the division machinery, while SpoIIIE is not, and it is otherwise a freely diffusive cytosolic enzymein vivo. Our developed SMT script is a powerful technique to determine if low-abundance proteins are membrane bound or cytosolic, to detect differences in populations of complex-bound and unbound/diffusive proteins, and to visualize the subcellular localization of slow- and fast-moving molecules in live cells.IMPORTANCEDNA translocases couple the late events of chromosome segregation to cell division and thereby play an important role in the bacterial cell cycle. The proteins fall into one of two categories, integral membrane translocases or nonintegral translocases. We show that the membrane-bound translocase SpoIIIE moves slowly throughout the cell membrane inB. subtilisand does not show a clear association with the division septum, in agreement with the idea that it binds membrane-bound DNA, which can occur through cell division across nonsegregated chromosomes. In contrast, SftA behaves like a soluble protein and is recruited to the division septum as a component of the division machinery. We show that FtsA contributes to the recruitment of SftA, revealing a dual role of FtsA at the division machinery, but it is not the only factor that binds SftA. Our work represents a detailedin vivostudy of DNA translocases at the single-molecule level.
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Maurya, Ganesh K., Kruti Modi, Manisha Banerjee, Reema Chaudhary, Yogendra S. Rajpurohit, and Hari S. Misra. "Phosphorylation of FtsZ and FtsA by a DNA Damage-Responsive Ser/Thr Protein Kinase Affects Their Functional Interactions in Deinococcus radiodurans." mSphere 3, no. 4 (2018): e00325-18. http://dx.doi.org/10.1128/msphere.00325-18.
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ABSTRACT Deinococcus radiodurans, a highly radioresistant bacterium, does not show LexA-dependent regulation of recA expression in response to DNA damage. On the other hand, phosphorylation of DNA repair proteins such as PprA and RecA by a DNA damage-responsive Ser/Thr protein kinase (STPK) (RqkA) could improve their DNA metabolic activities as well as their roles in the radioresistance of D. radiodurans. Here we report RqkA-mediated phosphorylation of cell division proteins FtsZ and FtsA in vitro and in surrogate Escherichia coli bacteria expressing RqkA. Mass spectrometric analysis mapped serine 235 and serine 335 in FtsZ and threonine 272, serine 370, and serine 386 in FtsA as potential phosphorylation sites. Although the levels of FtsZ did not change during postirradiation recovery (PIR), phosphorylation of both FtsZ and FtsA showed a kinetic change during PIR. However, in an rqkA mutant of D. radiodurans, though FtsZ underwent phosphorylation, no kinetic change in phosphorylation was observed. Further, RqkA adversely affected FtsA interaction with FtsZ, and phosphorylated FtsZ showed higher GTPase activity than unphosphorylated FtsZ. These results suggest that both FtsZ and FtsA are phosphoproteins in D. radiodurans. The increased phosphorylation of FtsZ in response to radiation damage in the wild-type strain but not in an rqkA mutant seems to be regulating the functional interaction of FtsZ with FtsA. For the first time, we demonstrate the role of a DNA damage-responsive STPK (RqkA) in the regulation of functional interaction of cell division proteins in this bacterium. IMPORTANCE The LexA/RecA-type SOS response is the only characterized mechanism of DNA damage response in bacteria. It regulates cell cycle by attenuating the functions of cell division protein FtsZ and inducing the expression of DNA repair proteins. There are bacteria, including Deinococcus radiodurans, that do not show this classical SOS response. D. radiodurans is known for its extraordinary resistance to gamma radiation, and a DNA damage-responsive Ser/Thr protein kinase (RqkA) has been characterized for its role in radioresistance. RqkA phosphorylates a large number of proteins in solution. The phosphorylation of RecA and PprA by RqkA enhanced their activities. FtsZ phosphorylation is inducible by gamma radiation in wild-type D. radiodurans but not in an rqkA mutant. Phosphorylation affected the interaction of FtsZ and FtsA in this bacterium. This study, therefore, brought forth some findings that might lead to the discovery of a new mechanism regulating the bacterial cell cycle in response to DNA damage.
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Chon, Younghae, and Randall Gayda. "Studies with FtsA-LacZ protein fusions reveal FtsA located inner-outer membrane junctions." Biochemical and Biophysical Research Communications 152, no. 3 (1988): 1023–30. http://dx.doi.org/10.1016/s0006-291x(88)80386-3.
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Yim, Lucía, Guy Vandenbussche, Jesús Mingorance, et al. "Role of the Carboxy Terminus of Escherichia coli FtsA in Self-Interaction and Cell Division." Journal of Bacteriology 182, no. 22 (2000): 6366–73. http://dx.doi.org/10.1128/jb.182.22.6366-6373.2000.
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ABSTRACT The role of the carboxy terminus of the Escherichia coli cell division protein FtsA in bacterial division has been studied by making a series of short sequential deletions spanning from residue 394 to 420. Deletions as short as 5 residues destroy the biological function of the protein. Residue W415 is essential for the localization of the protein into septal rings. Overexpression of theftsA alleles harboring these deletions caused a coiled cell phenotype previously described for another carboxy-terminal mutation (Gayda et al., J. Bacteriol. 174:5362–5370, 1992), suggesting that an interaction of FtsA with itself might play a role in its function. The existence of such an interaction was demonstrated using the yeast two-hybrid system and a protein overlay assay. Even these short deletions are sufficient for impairing the interaction of the truncated FtsA forms with the wild-type protein in the yeast two-hybrid system. The existence of additional interactions between FtsA molecules, involving other domains, can be postulated from the interaction properties shown by the FtsA deletion mutant forms, because although unable to interact with the wild-type and with FtsAΔ1, they can interact with themselves and cross-interact with each other. The secondary structures of an extensive deletion, FtsAΔ27, and the wild-type protein are indistinguishable when analyzed by Fourier transform infrared spectroscopy, and moreover, FtsAΔ27 retains the ability to bind ATP. These results indicate that deletion of the carboxy-terminal 27 residues does not alter substantially the structure of the protein and suggest that the loss of biological function of the carboxy-terminal deletion mutants might be related to the modification of their interacting properties.
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Tamura, Masaru, Kangseok Lee, Christine A. Miller, et al. "RNase E Maintenance of Proper FtsZ/FtsA Ratio Required for Nonfilamentous Growth of Escherichia coli Cells but Not for Colony-Forming Ability." Journal of Bacteriology 188, no. 14 (2006): 5145–52. http://dx.doi.org/10.1128/jb.00367-06.
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ABSTRACT Inactivation or deletion of the RNase E-encoding rne gene of Escherichia coli results in the growth of bacterial cells as filamentous chains in liquid culture (K. Goldblum and D. Apirion, J. Bacteriol. 146:128-132, 1981) and the loss of colony-forming ability (CFA) on solid media. RNase E dysfunction is also associated with abnormal processing of ftsQAZ transcripts (K. Cam, G. Rome, H. M. Krisch, and J.-P. Bouché, Nucleic Acids Res. 24:3065-3070, 1996), which encode proteins having a central role in septum formation during cell division. We show here that RNase E regulates the relative abundances of FtsZ and FtsA proteins and that RNase E depletion results in decreased FtsZ, increased FtsA, and consequently an altered FtsZ/FtsA ratio. However, while restoration of the level of FtsZ to normal in rne null mutant bacteria reverses the filamentation phenotype, it does not restore CFA. Conversely, overexpression of a related RNase, RNase G, in rne-deleted bacteria restores CFA, as previously reported, without affecting FtsZ abundance. Our results demonstrate that RNase E activity is required to maintain a proper cellular ratio of the FtsZ and FtsA proteins in E. coli but that FtsZ deficiency does not account for the nonviability of cells lacking RNase E.