Editors' ChoiceMicrobiology

Defining the daughter swarmer cell

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Sci. Signal.  11 Oct 2016:
Vol. 9, Issue 449, pp. ec239
DOI: 10.1126/scisignal.aal1591

The freshwater bacterium Caulobacter crescentus has a complex cell cycle involving two different cell types: an adhesive stalked cell capable of division and a swarmer cell capable of motility and differentiation into the stalked form. Stalked cells divide asymmetrically to form one stalked and one swarmer cell. Ricci et al. determined that the essential nucleoid-associated protein (NAP) called growth-associated A/T-binding protein (GapR) is necessary for this asymmetrical division. On the basis of its sequence, GapR was predicted to associate with DNA. Depletion (by expression of a form that could be induced to undergo proteolysis) or overexpression of GapR resulted in abnormal cell morphology and aberrant cell division, with the appearance of some mini-cell daughters that did not completely separate from the mother cell. Chromatin immunoprecipitation with DNA sequencing (ChIP-seq) of FLAG-tagged GapR revealed that the protein binds to AT-rich intergenic regions, particularly in the promoter regions of numerous genes with expression that varied during the cell cycle, such as those encoding proteins specific to swarmer cells and cell cycle regulatory factors. Most of the GapR-binding sites in promoters overlapped with the binding sites of master regulator protein MucR1. However, GapR depletion or overexpression had little effect on the transcription of these genes, suggesting that GapR does not directly regulate gene expression. Before the cells divided, fluorescently tagged GapR localized to part of the cell that would become the swarmer daughter cell, a localization that contrasted with that of an unrelated, non-essential AT-binding NAP (H-NS) from Escherichia coli, which distributed symmetrically when expressed in C. crescentus. Overexpression of either an asymmetrically (GapR) or symmetrically (H-NS) localized DNA-binding protein disrupted cell division. Most genomes of free-living alphaproteobacteria (which include both symmetrically and asymmetrically dividing bacteria), but not other clades of bacteria, contained homologs of gapR. A tagged GapR homolog from Rhodobacter capsulatus (a bacterium that does not differentiate into different cell types) distributed asymmetrically into the swarmer daughter when expressed in C. crescentus, indicating that the asymmetric distribution may be related to other proteins or molecules in C. crescentus rather than an intrinsic property of all GapR homologs. Similar to overexpressing gapR, individually overexpressing homologs from several different species (including one from a bacteriophage) in C. crescentus resulted in defects in asymmetric cell division. When expressed in E. coli, which divide symmetrically, C. crescentus GapR localized symmetrically, similarly to H-NS, but did not perform a transcriptional repression function of H-NS (silencing the bgl operon). Thus, correctly regulated GapR performs essential roles in asymmetric cell division and binds to AT-rich DNA in a manner distinct from that of previously studied NAPs, yet does not appear to regulate gene expression. These findings provide clues to the mechanisms used by bacteria to control 3D genomic organization, cell division, and differentiation in the absence of histones and other methods of chromosomal organization found in eukaryotes.

D. P. Ricci, M. D. Melfi, K. Lasker, D. L. Dill, H. H. McAdams, L. Shapiro, Cell cycle progression in Caulobacter requires a nucleoid-associated protein with high AT sequence recognition. Proc. Nat. Acad. Sci. USA 113, E5952–E5961 (2016). [PubMed]