Characterization of the role of MrpC in Myxococcus xanthus developmental cell fate determination
Myxococcus xanthus is an excellent model system for multicellular prokaryotic behaviour and Gram-negative differentiation. Under nutrient-limited conditions, the population enters a complex multicellular developmental program wherein cells undergo at least three distinct known cell fates: sporulatio...
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|Zusammenfassung:||Myxococcus xanthus is an excellent model system for multicellular prokaryotic behaviour and Gram-negative differentiation. Under nutrient-limited conditions, the population enters a complex multicellular developmental program wherein cells undergo at least three distinct known cell fates: sporulation within multicellular fruiting bodies; differentiation into persister-like state termed peripheral rods and cell lysis. A fourth distinct, relatively less understood cell type, called the cell clusters is also thought to exist. This starvation-induced developmental program is tightly regulated by the temporal and spatial expression of specific genes. One of them is mrpC, which codes for an important developmental transcriptional regulator. We hypothesized that MrpC, which is necessary for inducing aggregation and sporulation and was implicated in mediating developmental cell lysis, may act as a master cell fate regulator in M. xanthus. MrpC has been shown to accumulate heterogeneously in developmental subpopulations and its misaccumulation results in perturbed cell fate segregation. MrpC is known to be highly regulated. It has previously been proposed that MrpC positively regulates its own expression. The transcriptional activity of MrpC is thought to be regulated- MrpC is inactivated by phosphorylation and activated by proteolytic processing to the shorter isoform MrpC2. In the presented thesis, using a combination of genetic and fluorescent techniques, I characterized the regulation of the transcription factor MrpC and my data suggests that MrpC negatively regulates its own expression. My analysis further revealed that the MrpC protein heterogeneity in developmental cell subpopulations is not due to transcriptional or translational differences but likely regulated by a protein turnover in the aggregated cell population. Using genetic and biochemical techniques, I addressed the two known activity states of MrpC. My detailed in vivo analysis revealed that the previously published in vitro data needs to be re-evaluated in order to assign a biological role to an isoform of MrpC. Using fluorescent reporters of promoter activity, I analyzed single-cell expression of various genes in developmental subpopulations. It was revealed that targets of MrpC do not follow the differential accumulation of MrpC. Moreover, cell subpopulations cannot be distinguished by means of cell-fate specific transcriptional markers or chromosome status. This study emphasizes on the implications of regulation of the key developmental regulator MrpC being different than previously proposed and provides the framework to re-investigate the proposed models of MrpC regulation in order to aid an appropriate understanding of how this complicated system functions to enable cells to adopt distinct cell fates.|