This proposal is focused on understanding fundamental properties of bacterial chromosome dynamics: How is the bacterial chromosome remodeled and segregated during the cell cycle and differentiation? How is chromosome organization and segregation linked to DNA replication? What are the roles of the cis-acting elements and trans-acting factors implicated in these processes? How is mis-segregated DNA translocated into the appropriate daughter after cytokinesis? These most basic biological questions have proven remarkably intractable, in part, due to functional redundancy and the lack of quantitative assays. We propose to study the organization and segregation of the chromosome in the model gram-positive bacterium Bacillus subtilis. The ease with which genetic, biochemical and cytological analysis can be carried out in this organism makes it an ideal system to study chromosome dynamics. We will address these questions using the next generation of molecular and cell biological tools and quantitative assays that we have developed to study the chromosome during spore-formation and the vegetative cell cycle. As a conceptual and experimental framework, the process of chromosome segregation in bacteria can be divided into three steps. i) The newly replicated origins are re-positioned towards the cell poles. ii) The sister chromosomes are remodeled and segregated followed by cell division. iii) Unsegregated DNA present at the division plane at the time of cytokinesis is translocated across the septum into the appropriate daughter cell. This dynamic process is executed in every cell cycle with extremely high fidelity. In this proposal we will investigate the molecular underpinnings of all three steps. We propose to: 1) Determine how the chromosomal partitioning protein ParB bound to its cognate parS sites recruits the SMC condensation complex to the origin and how origin-localized SMC compacts and organizes the chromosome. 2) Determine how the SpoIIIE translocase functions at the division to septum to transport unsegregated DNA. 3) Investigate how replication initiation and origin segregation are linked upon entry into sporulation.

Public Health Relevance

The faithful segregation of chromosomes to daughter cells is an essential process. Understanding the molecular mechanisms that govern this process in bacteria could lead to the discovery of new targets appropriate for antimicrobial therapeutics. In this proposal we investigate chromosome dynamics during growth and sporulation in Bacillus subtilis. Accordingly, insights will be particularly relevant to gram-positive pathogens and spore-formers of the Clostridiales and B. anthracis.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM086466-03
Application #
8098890
Study Section
Prokaryotic Cell and Molecular Biology Study Section (PCMB)
Program Officer
Janes, Daniel E
Project Start
2009-08-01
Project End
2013-06-30
Budget Start
2011-07-01
Budget End
2012-06-30
Support Year
3
Fiscal Year
2011
Total Cost
$329,808
Indirect Cost
Name
Harvard University
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
047006379
City
Boston
State
MA
Country
United States
Zip Code
02115
Ramírez-Guadiana, Fernando H; Meeske, Alexander J; Wang, Xindan et al. (2017) The Bacillus subtilis germinant receptor GerA triggers premature germination in response to morphological defects during sporulation. Mol Microbiol 105:689-704
Wang, Xindan; Brandão, Hugo B; Le, Tung B K et al. (2017) Bacillus subtilis SMC complexes juxtapose chromosome arms as they travel from origin to terminus. Science 355:524-527
Rodrigues, Christopher D A; Henry, Xavier; Neumann, Emmanuelle et al. (2016) A ring-shaped conduit connects the mother cell and forespore during sporulation in Bacillus subtilis. Proc Natl Acad Sci U S A 113:11585-11590
Widderich, Nils; Rodrigues, Christopher D A; Commichau, Fabian M et al. (2016) Salt-sensitivity of ?(H) and Spo0A prevents sporulation of Bacillus subtilis at high osmolarity avoiding death during cellular differentiation. Mol Microbiol 100:108-24
Wang, Xindan; Le, Tung B K; Lajoie, Bryan R et al. (2015) Condensin promotes the juxtaposition of DNA flanking its loading site in Bacillus subtilis. Genes Dev 29:1661-75
Meeske, Alexander J; Sham, Lok-To; Kimsey, Harvey et al. (2015) MurJ and a novel lipid II flippase are required for cell wall biogenesis in Bacillus subtilis. Proc Natl Acad Sci U S A 112:6437-42
Wang, Xindan; Tang, Olive W; Riley, Eammon P et al. (2014) The SMC condensin complex is required for origin segregation in Bacillus subtilis. Curr Biol 24:287-92
Graham, Thomas G W; Wang, Xindan; Song, Dan et al. (2014) ParB spreading requires DNA bridging. Genes Dev 28:1228-38
Wang, Xindan; Montero Llopis, Paula; Rudner, David Z (2014) Bacillus subtilis chromosome organization oscillates between two distinct patterns. Proc Natl Acad Sci U S A 111:12877-82
Wang, Xindan; Rudner, David Z (2014) Spatial organization of bacterial chromosomes. Curr Opin Microbiol 22:66-72

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