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IntroductionMrpC, a member of the CRP/Fnr transcription factor superfamily, is necessary to induce and control the multicellular developmental program of the bacterium,Myxococcus xanthus. During development, certain cells in the population first swarm into haystack-shaped aggregates and then differentiate into environmentally resistant spores to form mature fruiting bodies (a specialized biofilm).mrpCtranscriptional regulation is controlled by negative autoregulation (NAR). MethodsWild type and mutantmrpCpromoter regions were fused to a fluorescent reporter to examine effects onmrpCexpression in the population and in single cellsin situ. Phenotypic consequences of the mutantmrpCpromoter were assayed by deep convolution neural network analysis of developmental movies, sporulation efficiency assays, and anti-MrpC immunoblot. In situ analysis of single cell MrpC levels in distinct populations were assayed with an MrpC-mNeonGreen reporter. ResultsDisruption of MrpC binding sites within themrpCpromoter region led to increased and broadened distribution ofmrpCexpression levels between individual cells in the population. Expression ofmrpCfrom the mutant promoter led to a striking phenotype in which cells lose synchronized transition from aggregation to sporulation. Instead, some cells abruptly exit aggregation centers and remain locked in a cohesive swarming state we termed developmental swarms, while the remaining cells transition to spores inside residual fruiting bodies.In situexamination of a fluorescent reporter for MrpC levels in developmental subpopulations demonstrated cells locked in the developmental swarms contained MrpC levels that do not reach the levels observed in fruiting bodies. DiscussionIncreased cell-to-cell variation inmrpCexpression upon disruption of MrpC binding sites within its promoter is consistent with NAR motifs functioning to reducing noise. Noise reduction may be key to synchronized transition of cells in the aggregation state to the sporulation state. We hypothesize a novel subpopulation of cells trapped as developmental swarms arise from intermediate levels of MrpC that are sufficient to promote aggregation but insufficient to trigger sporulation. Failure to transition to higher levels of MrpC necessary to induce sporulation may indicate cells in developmental swarms lack an additional positive feedback signal required to boost MrpC levels.
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Cell-size regulation in budding yeast does not depend on linear accumulation of Whi5
Cells must couple cell-cycle progress to their growth rate to restrict the spread of cell sizes present throughout a population. Linear, rather than exponential, accumulation of Whi5, was proposed to provide this coordination by causing a higher Whi5 concentration in cells born at a smaller size. We tested this model using the inducibleGAL1promoter to make the Whi5 concentration independent of cell size. At an expression level that equalizes the mean cell size with that of wild-type cells, the size distributions of cells with galactose-induced Whi5 expression and wild-type cells are indistinguishable. Fluorescence microscopy confirms that the endogenous andGAL1promoters produce different relationships between Whi5 concentration and cell volume without diminishing size control in the G1 phase. We also expressed Cln3 from the GAL1 promoter, finding that the spread in cell sizes for an asynchronous population is unaffected by this perturbation. Our findings indicate that size control in budding yeast does not fundamentally originate from the linear accumulation of Whi5, contradicting a previous claim and demonstrating the need for further models of cell-cycle regulation to explain how cell size controls passage through Start.
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- Award ID(s):
- 1752024
- PAR ID:
- 10160413
- Publisher / Repository:
- Proceedings of the National Academy of Sciences
- Date Published:
- Journal Name:
- Proceedings of the National Academy of Sciences
- Volume:
- 117
- Issue:
- 25
- ISSN:
- 0027-8424
- Page Range / eLocation ID:
- p. 14243-14250
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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