Streptomyces sporulation - Genes and regulators involved in bacterial cell differentiation

Sammanfattning: Streptomycetes are Gram-positive bacteria with a complex developmental life cycle. They form spores on specialized cells called aerial hyphae, and this sporulation involves alterations in growth, morphogenesis and cell cycle processes like cell division and chromosome segregation. Understanding the developmental mechanisms that streptomycetes have evolved for regulating for example cell division is of general interest in bacterial cell biology. It can also be valuable in the design of new drugs against bacterial pathogens. Very few sporulation genes have been found with an impact on cell cycle-related processes. Finding of such genes is important for a clarification of the molecular mechanisms that are underlying the developmental control of fundamental cellular processes in Streptomyces. The work of this thesis has led to the identification of genes previously unknown to be developmentally regulated in Streptomyces. By comparing the transcriptome of the wildtype S. coelicolor strain M145 to two developmental mutants, whiA and whiH, which specifically affect sporulation processes, it was possible to identify differentially expressed genes. Genes that so far have been characterized proved to have important roles during sporulation, affecting spore maturation, chromosome condensation and cell division. WhiH is a central regulator in the early sporulation process and required for the developmentally controlled form of cell division in S. coelicolor. In this thesis the role of WhiH as a transcription factor has been established and WhiH was found to bind to a specific site in its own promoter and function as an autoregulator. A whiHp-mCherry reporter fusion was used to monitor cell type-specific activity of the whiH promoter in aerial hyphae, and showed that it is active before delimitation of the sporogenic cell in which multiple developmentally controlled cell divisions will be triggered. A new Streptomyces model organism, S. venezuelae, was finally exploited to identify target genes for control by WhiH. This organism sporulates efficiently in liquid culture and is well suited for global transcriptomic approaches. In this study, microarray analysis of whiH-dependent gene expression was used to find putative targets for WhiH. Combined with chromatin-immunoprecipitation (ChIP-chip) and protein-DNA binding assays this identified a group of genes that are directly repressed by WhiH during a late stage of sporulation, and also some candidate genes that could be activated by WhiH at an earlier stage. Future analyses should shed light on the functions of these genes and their potential roles in developmental and cell cycle-related processes.