scholarly journals DivIVA is essential in Deinococcus radiodurans and its C terminal domain regulates new septum orientation during cell division

2020 ◽  
Author(s):  
Reema Chaudhary ◽  
Swathi Kota ◽  
Hari S Misra
Keyword(s):  
Cell Division ◽  
Deinococcus Radiodurans ◽  
Wild Type ◽  
Orthogonal Axis ◽  
Septum Formation ◽  
Terminal Domain ◽  
Cell Position ◽  
Type Pattern ◽  
In Trans ◽  
Ftsz Assembly

AbstractFtsZ assembly at mid cell position in rod shaped bacteria is regulated by gradient of MinCDE complex across the poles. In round shaped bacteria, which lack predefined poles and the next plane of cell division is perpendicular to previous plane, the determination of site for FtsZ assembly is intriguing. Deinococcus radiodurans a coccus shaped bacterium, is characterized for its extraordinary resistance to DNA damage. Here we report that DivIVA a putative component of Min system in this bacterium (drDivIVA) interacts with cognate cell division and genome segregation proteins. The deletion of full length drDivIVA was found to be indispensable while its C-terminal deletion (ΔdivIVAC) was dispensable but produced distinguishable phenotypes like slow growth, altered plane for new septum formation and angular septum. Both wild type and mutant showed FtsZ foci formation and their gamma radiation responses were nearly identical. But unlike in wild type, the FtsZ localization in mutant cells was found to be away from orthogonal axis with respect to plane of previous septum. Notably, DivIVA-RFP localizes to membrane during cell division and then perpendicular to previous plane of cell division. In trans expression of drDivIVA in ΔdivIVAC background could restore the wild type pattern of septum formation perpendicular to previous septum. These results suggested that DivIVA is an essential protein in D. radiodurans and the C-terminal domain that contributes to its interaction with MinC determines the plane of new septum formation, possibly by controlling MinC oscillation through orthogonal axis in the cells.

scholarly journals DivIVA regulates its expression and the orientation of new septum growth during cell division in Deinococcus radiodurans.

2021 ◽  
Author(s):  
Reema Chaudhary ◽  
Swathi Kota ◽  
Hari S Misra
Keyword(s):  
Cell Division ◽  
Deinococcus Radiodurans ◽  
Gram Negative Bacteria ◽  
Wild Type ◽  
Gram Negative ◽  
Terminal Domain ◽  
Division Site ◽  
Correct Orientation ◽  
Cell Position ◽  
In Trans

In rod shaped Gram-negative bacteria, FtsZ localization at mid cell position is regulated by the gradient of MinCDE complex across the poles. In round shaped bacteria, which lack predefined poles, the next plane of cell division is perpendicular to previous plane and the determination of site for FtsZ assembly is still intriguing. Deinococcus radiodurans, a coccus bacterium, is characterized by its extraordinary resistance to DNA damage. DivIVA, a putative component of Min system in this bacterium, interacts with cognate cell division and genome segregation proteins. Here, we report that deletion of chromosomal copy of DivIVA was possible only when wild type copy of DivIVA was expressed in trans on the plasmid. However, the deletion of C-terminal domain of DivIVA (CTD mutant) was possible but produced distinguishable phenotypes like smaller cells, slower growth and tilted septum orientation in D. radiodurans. In trans expression of DivIVA in CTD mutant could restore these features of wild type. Interestingly, the overexpression of DivIVA led to delayed separation of tetrad from octet state in both trans-complemented divIVA mutant and wild type cells. The CTD mutant showed upregulation of yggS-divIVAN operon. Both wild type and CTD mutant formed FtsZ foci, however unlike wild type, the position of foci in the mutant cells was found to be away from conjectural mid-cell position in cocci. Notably, DivIVA-RFP localizes to septum during cell division at the new division site. These results suggested that DivIVA is an essential protein in D. radiodurans and its C-terminal domain plays an important role in the regulation of its expression and orientation of new septal growth in this bacterium. Importance: In rod-shaped Gram-negative bacteria, the mid-cell position for binary fission is relatively easy to model. In cocci that do not have predefined poles, the plane of next cell division is shown to be perpendicular to the previous plane. However, the molecular basis of perpendicularity is not known in cocci. The DivIVA protein of Deinococcus radiodurans, a coccus bacterium, physically interacts with septum and establishes macromolecular interactions with genome segregation proteins through its N-terminal domain and with MinC through C-terminal domain. Here, we have brought forth some evidence to suggest that DivIVA is essential for growth, plays an important role in cell-polarity determination and its C-terminal domain plays a crucial role in the growth of new septum in correct orientation as well as regulation of its expression.

scholarly journals Analysis of MinC Reveals Two Independent Domains Involved in Interaction with MinD and FtsZ

2000 ◽  
Vol 182 (14) ◽  
pp. 3965-3971 ◽  
Author(s):  
Zonglin Hu ◽  
Joe Lutkenhaus
Keyword(s):  
Escherichia Coli ◽  
Cell Division ◽  
Functional Domains ◽  
Wild Type ◽  
Terminal Domain ◽  
Ring Assembly ◽  
Z Ring ◽  
Ftsz Assembly

ABSTRACT In Escherichia coli FtsZ assembles into a Z ring at midcell while assembly at polar sites is prevented by themin system. MinC, a component of this system, is an inhibitor of FtsZ assembly that is positioned within the cell by interaction with MinDE. In this study we found that MinC consists of two functional domains connected by a short linker. When fused to MalE the N-terminal domain is able to inhibit cell division and prevent FtsZ assembly in vitro. The C-terminal domain interacts with MinD, and expression in wild-type cells as a MalE fusion disrupts minfunction, resulting in a minicell phenotype. We also find that MinC is an oligomer, probably a dimer. Although the C-terminal domain is clearly sufficient for oligomerization, the N-terminal domain also promotes oligomerization. These results demonstrate that MinC consists of two independently functioning domains: an N-terminal domain capable of inhibiting FtsZ assembly and a C-terminal domain responsible for localization of MinC through interaction with MinD. The fusion of these two independent domains is required to achieve topological regulation of Z ring assembly.

scholarly journals Overlapping oriC and centromere-like functions in secondary genome replicons determine their maintenance independent of chromosome I in Deinococcus radiodurans

2020 ◽  
Author(s):  
Ganesh K Maurya ◽  
Hari S. Misra
Keyword(s):  
Radiation Resistance ◽  
Copy Number ◽  
Deinococcus Radiodurans ◽  
Specific Interactions ◽  
Wild Type ◽  
Γ Radiation ◽  
E Coli ◽  
Copy Numbers ◽  
Type Pattern ◽  
Chromosome I

AbstractThe Deinococcus radiodurans multipartite genome system (MGS) consists of chromosome I (ChrI) and secondary genome elements; Chr II and megaplasmid (MP). The sequences upstream to parAB operons in Chr II (cisII) and MP (cisMP) helped an E. coli plasmid maintenance in D. radiodurans and showed sequence specific interactions with DnaA and ParBs. The cells devoid of cisII (ΔcisII) or cisMP (ΔcisMP) showed reduced γ radiation resistance and copy number of Chr II and MP. Fluorescent Reporter-Operator System (FROS) developed for ChrI, ChrII and MP in ΔcisII or ΔcisMP mutants showed no change in wild type pattern of Chr I localization. However, the relative copy numbers of Chr II and MP had reduced while anucleate cells had increased in mutants. These results suggested that cisII and cisMP elements contain both ori and centromere-like functions, and like other MGS bacteria, the Chr I and secondary genome are maintained independently in D. radiodurans.

scholarly journals Cell Division Inhibition in Salmonella typhimuriumHistidine-Constitutive Strains: an ftsI-Like Defect in the Presence of Wild-Type Penicillin-Binding Protein 3 Levels

1998 ◽  
Vol 180 (19) ◽  
pp. 5231-5234 ◽  
Author(s):  
David A. Cano ◽  
Chakib Mouslim ◽  
Juan A. Ayala ◽  
Francisco García-del Portillo ◽  
Josep Casadesús
Keyword(s):  
Cell Division ◽  
Carbon Source ◽  
Salmonella Typhimurium ◽  
Binding Protein ◽  
Penicillin Binding Protein ◽  
Wild Type ◽  
Septum Formation ◽  
Phenotypic Suppression ◽  
High Concentrations ◽  
Cell Division Inhibition

ABSTRACT Histidine-constitutive (Hisc) strains ofSalmonella typhimurium undergo cell division inhibition in the presence of high concentrations of a metabolizable carbon source. Filaments formed by Hisc strains show constrictions and contain evenly spaced nucleoids, suggesting a defect in septum formation. Inhibitors of penicillin-binding protein 3 (PBP3) induce a filamentation pattern identical to that of Hisc strains. However, the Hisc septation defect is caused neither by reduced PBP3 synthesis nor by reduced PBP3 activity. Gross modifications of peptidoglycan composition are also ruled out.d-Cycloserine, an inhibitor of the soluble pathway producing peptidoglycan precursors, causes phenotypic suppression of filamentation, suggesting that the septation defect of Hiscstrains may be caused by scarcity of PBP3 substrate.

scholarly journals Phosphorylation of FtsZ and FtsA by a DNA Damage-Responsive Ser/Thr Protein Kinase Affects Their Functional Interactions in Deinococcus radiodurans

mSphere ◽  
2018 ◽  
Vol 3 (4) ◽  
pp. e00325-18 ◽  
Author(s):  
Ganesh K. Maurya ◽  
Kruti Modi ◽  
Manisha Banerjee ◽  
Reema Chaudhary ◽  
Yogendra S. Rajpurohit ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Cell Division ◽  
Protein Kinase ◽  
Deinococcus Radiodurans ◽  
Sos Response ◽  
Wild Type ◽  
Functional Interaction ◽  
Content Type ◽  
Dna Repair Proteins

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.

scholarly journals ssgA Is Essential for Sporulation ofStreptomyces coelicolor A3(2) and Affects Hyphal Development by Stimulating Septum Formation

2000 ◽  
Vol 182 (20) ◽  
pp. 5653-5662 ◽  
Author(s):  
Gilles P. van Wezel ◽  
Jannes van der Meulen ◽  
Shinichi Kawamoto ◽  
Ruud G. M. Luiten ◽  
Henk K. Koerten ◽  
...  
Keyword(s):  
Cell Division ◽  
Morphological Changes ◽  
Antibiotic Production ◽  
Streptomyces Coelicolor ◽  
Wild Type ◽  
Septum Formation ◽  
Transmission Electron ◽  
In The Wild ◽  
Regulator Genes

ABSTRACT The role of ssgA in cell division and development of streptomycetes was analyzed. An ssgA null mutant ofStreptomyces coelicolor produced aerial hyphae but failed to sporulate, and ssgA can therefore be regarded as a novelwhi gene. In addition to the morphological changes, antibiotic production was also disturbed, with strongly reduced actinorhodin production. These defects could be complemented by plasmid-borne ssgA. In the wild-type strain, transcription of ssgA was induced by nutritional shift-down and was shown to be linked to that of the upstream-located gene ssgR, which belongs to the family of iclR-type transcriptional regulator genes. Analysis of mycelium harvested from liquid-grown cultures by transmission electron microscopy showed that septum formation had strongly increased in ssgA-overexpressing strains in comparison to wild-type S. coelicolor and that spore-like compartments were produced at high frequency. Furthermore, the hyphae were significantly wider and contained irregular and often extremely thick septa. These data underline the important role forssgA in Streptomyces cell division.

scholarly journals Requirement of the LtsA Protein for Formation of the Mycolic Acid-Containing Layer on the Cell Surface of Corynebacterium glutamicum

2021 ◽  
Vol 9 (2) ◽  
pp. 409
Author(s):  
Yutaro Kumagai ◽  
Takashi Hirasawa ◽  
Masaaki Wachi
Keyword(s):  
Cell Division ◽  
Corynebacterium Glutamicum ◽  
Fluorescent Dyes ◽  
Intracellular Localization ◽  
Mycolic Acid ◽  
Differential Staining ◽  
Wild Type ◽  
Terminal Domain ◽  
Mutant Cells ◽  
Increased Susceptibility

The ltsA gene of Corynebacterium glutamicum encodes a purF-type glutamine-dependent amidotransferase, and mutations in this gene result in increased susceptibility to lysozyme. Recently, it was shown that the LtsA protein catalyzes the amidation of diaminopimelate residues in the lipid intermediates of peptidoglycan biosynthesis. In this study, intracellular localization of wild-type and mutant LtsA proteins fused with green fluorescent protein (GFP) was investigated. The GFP-fused wild-type LtsA protein showed a peripheral localization pattern characteristic of membrane-associated proteins. The GFP-fusions with a mutation in the N-terminal domain of LtsA, which is necessary for the glutamine amido transfer reaction, exhibited a similar localization to the wild type, whereas those with a mutation or a truncation in the C-terminal domain, which is not conserved among the purF-type glutamine-dependent amidotransferases, did not. These results suggest that the C-terminal domain is required for peripheral localization. Differential staining of cell wall structures with fluorescent dyes revealed that formation of the mycolic acid-containing layer at the cell division planes was affected in the ltsA mutant cells. This was also confirmed by observation that bulge formation was induced at the cell division planes in the ltsA mutant cells upon lysozyme treatment. These results suggest that the LtsA protein function is required for the formation of a mycolic acid-containing layer at the cell division planes and that this impairment results in increased susceptibility to lysozyme.

Developing a Genetic System in Deinococcus radiodurans for Analyzing Mutations

Genetics ◽  
2004 ◽  
Vol 166 (2) ◽  
pp. 661-668
Author(s):  
Mandy Kim ◽  
Erika Wolff ◽  
Tiffany Huang ◽  
Lilit Garibyan ◽  
Ashlee M Earl ◽  
...  
Keyword(s):  
Dna Sequences ◽  
Deinococcus Radiodurans ◽  
Genetic System ◽  
Base Change ◽  
Base Substitution ◽  
Wild Type ◽  
Base Pairs ◽  
E Coli ◽  
Radiation Induced ◽  
Induced Mutagenesis

Abstract We have applied a genetic system for analyzing mutations in Escherichia coli to Deinococcus radiodurans, an extremeophile with an astonishingly high resistance to UV- and ionizing-radiation-induced mutagenesis. Taking advantage of the conservation of the β-subunit of RNA polymerase among most prokaryotes, we derived again in D. radiodurans the rpoB/Rif r system that we developed in E. coli to monitor base substitutions, defining 33 base change substitutions at 22 different base pairs. We sequenced >250 mutations leading to Rif r in D. radiodurans derived spontaneously in wild-type and uvrD (mismatch-repair-deficient) backgrounds and after treatment with N-methyl-N′-nitro-N-nitrosoguanidine (NTG) and 5-azacytidine (5AZ). The specificities of NTG and 5AZ in D. radiodurans are the same as those found for E. coli and other organisms. There are prominent base substitution hotspots in rpoB in both D. radiodurans and E. coli. In several cases these are at different points in each organism, even though the DNA sequences surrounding the hotspots and their corresponding sites are very similar in both D. radiodurans and E. coli. In one case the hotspots occur at the same site in both organisms.

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