Structures of partition protein ParA with nonspecific DNA and ParB effector reveal molecular insights into principles governing Walker-box DNA segregation

Zhang, Hengshan; Schumacher, Maria A.

doi:10.1101/gad.296319.117

Cited by 53 publications

(71 citation statements)

References 62 publications

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“…In C. crescentus ParB depletion or mutation quenches ParA dynamics, leading to ParA diffuse localisation (Ptacin et al ., ). It has been shown before that ParA‐DNA binding allows ParB movement (Schumacher and Funnell, ; Vecchiarelli et al ., ; Le Gall et al ., ; Zhang and Schumacher, ). Thus our results are consistent with general mechanism of ParA action in bacteria.…”

Section: Discussionmentioning

confidence: 97%

“…However, more recent findings suggest, that ParA dimers (or short filaments) translocate partition complexes on the surface of the nucleoid through diffusion-ratchet mechanism. Additionally, it has been shown that chromosome elasticity may contribute to the movement of segrosomes (Vecchiarelli et al, 2010(Vecchiarelli et al, , 2014Hwang et al, 2013;Lim et al, 2014;Sanchez et al, 2015;Zhang and Schumacher, 2017).…”

Section: Introductionmentioning

confidence: 99%

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The studies of ParA and ParB dynamics reveal asymmetry of chromosome segregation in mycobacteria

Ginda

Santi

Bousbaine

et al. 2017

Molecular Microbiology

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Active segregation of bacterial chromosomes usually involves the action of ParB proteins, which bind in proximity of chromosomal origin (oriC) regions forming nucleoprotein complexes - segrosomes. Newly duplicated segrosomes are moved either uni- or bidirectionally by the action of ATPases - ParA proteins. In Mycobacterium smegmatis the oriC region is located in an off-centred position and newly replicated segrosomes are segregated towards cell poles. The elimination of M. smegmatis ParA and/or ParB leads to chromosome segregation defects. Here, we took advantage of microfluidic time-lapse fluorescent microscopy to address the question of ParA and ParB dynamics in M. smegmatis and M. tuberculosis cells. Our results reveal that ParB complexes are segregated in an asymmetrical manner. The rapid movement of segrosomes is dependent on ParA that is transiently associated with the new pole. Remarkably in M. tuberculosis, the movement of the ParB complex is much slower than in M. smegmatis, but segregation as in M. smegmatis lasts approximately 10% of the cell cycle, which suggests a correlation between segregation dynamics and the growth rate. On the basis of our results, we propose a model for the asymmetric action of segregation machinery that reflects unequal division and growth of mycobacterial cells.

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Section: Discussionmentioning

confidence: 97%

Section: Introductionmentioning

confidence: 99%

The studies of ParA and ParB dynamics reveal asymmetry of chromosome segregation in mycobacteria

Ginda

Santi

Bousbaine

et al. 2017

Molecular Microbiology

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“…While previous studies have described the main components of the mycobacterial chromosome segregation machinery, the ParA and ParB proteins, and have reported ParA interaction with a polar growth determinant DivIVA homologue (named Wag31 in mycobacteria) (Jakimowicz et al , ; Ginda et al , ; ; Uhía et al ., ), the exact mechanism of asymmetric chromosome segregation and the biological role of the ParA‐DivIVA interaction have not been investigated. In mycobacteria, as in most studied bacterial species ( Bacillus subtilis, Caulobacter crescentus, Vibrio cholerae, Myxococcus xanthus, Corynebacterium glutamicum , and Streptomyces coelicolor , but not Escherichia coli and some other γ‐proteobacteria), origin of chromosomal replication ( oriC ) regions are organised into complexes named segrosomes by ParB binding to oriC ‐proximal parS sites (Chaudhuri and Dean, ; Wang et al , ; Graham et al , ; Zhang and Schumacher, ). Usually immediately (but in some species after some cohesion time) following the initiation of replication, ParB complexes duplicate, and then one or both complexes are moved to their target cell locations.…”

Section: Introductionmentioning

confidence: 99%

Competition between DivIVA and the nucleoid for ParA binding promotes segrosome separation and modulates mycobacterial cell elongation

Pióro

Małecki

Portas

et al. 2018

Molecular Microbiology

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Summary Although mycobacteria are rod shaped and divide by simple binary fission, their cell cycle exhibits unusual features: unequal cell division producing daughter cells that elongate with different velocities, as well as asymmetric chromosome segregation and positioning throughout the cell cycle. As in other bacteria, mycobacterial chromosomes are segregated by pair of proteins, ParA and ParB. ParA is an ATPase that interacts with nucleoprotein ParB complexes – segrosomes and non‐specifically binds the nucleoid. Uniquely in mycobacteria, ParA interacts with a polar protein DivIVA (Wag31), responsible for asymmetric cell elongation, however the biological role of this interaction remained unknown. We hypothesised that this interaction plays a critical role in coordinating chromosome segregation with cell elongation. Using a set of ParA mutants, we determined that disruption of ParA‐DNA binding enhanced the interaction between ParA and DivIVA, indicating a competition between the nucleoid and DivIVA for ParA binding. Having identified the ParA mutation that disrupts its recruitment to DivIVA, we found that it led to inefficient segrosomes separation and increased the cell elongation rate. Our results suggest that ParA modulates DivIVA activity. Thus, we demonstrate that the ParA‐DivIVA interaction facilitates chromosome segregation and modulates cell elongation.

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“…Our knowledge of the determinants mediating the DNA-binding activity of P-loop ATPases is largely based on studies of canonical ParA homologs. For this group of proteins, two distinct modes of DNA binding, mediated by different regions of the dimer surface, have been reported (37,38). Moreover, the residues identified in previous studies are not universally conserved among DNA-binding members of the ParA-like ATPase family.…”

Section: Identification Of Dna-binding Residues On the Mipz Surfacementioning

confidence: 95%

“…Residues homologous to these arginines were later also implicated in the DNA-binding activity of PpfA from Rhodobacter sphaeroides and PomZ from Myxococcus xanthus (17,20). More evidence for the importance of positively charged amino acids has recently come from the crystal structures of ParA-DNA complexes formed by ParA homologs from Helicobacter pylori (HpParA) (37) and Sulfolobus solfataricus pNOB8 (38), which each identified several surface-exposed lysine residues that are in direct contact with the DNA ligand. However, the binding interface pNOB8 ParA was different from that of its H. pylori homolog or R. capsulatus PpfA (38), suggesting that ParA proteins could have evolved different modes of DNA binding.…”

Section: Introductionmentioning

confidence: 99%

Molecular architecture of the DNA‐binding sites of the P‐loop ATPases MipZ and ParA fromCaulobacter crescentus

Refes

Corrales-Guerrero

et al. 2019

Preprint

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Two related P-loop ATPases, ParA and MipZ, mediate the spatiotemporal regulation of chromosome segregation and cell division in Caulobacter crescentus. Both of these proteins share the ability to form dynamic concentration gradients that control the positioning of regulatory targets within the cell. Their proper localization relies on their nucleotide-dependent cycling between a monomeric and a dimeric state, driven by interaction with the chromosome partitioning protein ParB, and on the ability of the dimeric species to associate non-specifically with the nucleoid. In this study, we use a combination of genetic screening, biochemical analysis and hydrogen/deuterium exchange mass spectrometry to identify the residues mediating the interaction of MipZ with DNA. Our results show that the DNA-binding activity of MipZ relies on a series of positively charged and hydrophobic residues lining both sides of the dimer interface. MipZ thus appears to associate with DNA in a sequence-independent manner through electrostatic interactions with the DNA phosphate backbone. In support of this hypothesis, chromatin immunoprecipitation analyses did not reveal any specific target sites in vivo. When extending our analysis to ParA, we found that the architectures of the MipZ and ParA DNA-binding sites are markedly different, although their relative positions on the dimer surface and their mode of DNA binding are conserved. Importantly, bioinformatic analysis suggests that the same principles apply to other members of the P-loop ATPase family. ParA-like ATPases thus share common mechanistic features, although their modes of action have diverged considerably during the course of evolution. SIGNIFICANCEParA-like P-loop ATPases are involved in a variety of cellular processes in bacteria, including chromosome and plasmid segregation, chemoreceptor and carboxysome positioning, and division site placement. Many members of this large protein family depend on the ability to bind non-specific DNA for proper function.Although previous studies have yielded insights in the DNA-binding properties of some ParA-like ATPases, a comprehensive view of the underlying mechanisms is still lacking. Here, we combine state-of-the-art cell biological, biochemical and biophysical approaches to localize the DNA-binding regions of the ParA-like ATPases MipZ and ParA from Caulobacter crescentus. We show that the two proteins use the same interface and mode of action to associate with DNA, suggesting that the mechanistic basis of DNA binding may be conserved in the ParA-like ATPase family.

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Structures of partition protein ParA with nonspecific DNA and ParB effector reveal molecular insights into principles governing Walker-box DNA segregation

Cited by 53 publications

References 62 publications

The studies of ParA and ParB dynamics reveal asymmetry of chromosome segregation in mycobacteria

The studies of ParA and ParB dynamics reveal asymmetry of chromosome segregation in mycobacteria

Competition between DivIVA and the nucleoid for ParA binding promotes segrosome separation and modulates mycobacterial cell elongation

Molecular architecture of the DNA‐binding sites of the P‐loop ATPases MipZ and ParA fromCaulobacter crescentus

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