Mechanisms and regulation of replication, the cell cycle, gene expression, and horizontal gene transfer in prokaryotes, focusing on Bacillus subtilis

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Our long term goals are to understand many of the molecular mechanisms and regulation underlying basic cellular processes in bacteria. We are particularly interested in the control of DNA replication, cellular responses to perturbations in replication, environmental sensing and signal transduction, and mechanisms controlling horizontal gene transfer. Our organism of choice for these studies is the Gram positive bacterium Bacillus subtilis. There is a wealth of information about B. subtilis. It is easy to grow and manipulate genetically and is related to several pathogens and environmental bacteria that are less tractable experimentally. Horizontal gene transfer (HGT) is the driving force in microbial evolution. It is largely mediated by mobile genetic elements, including viruses, conjugative plasmids, and integrative and conjugative elements (ICEs, also known as conjugative transposons). Conjugative elements are well known agents contributing to the spread of genes for antibiotic resistances, pathogenesis, symbiosis, metabolism, and more. Despite the prevalence and importance of ICEs, there are deficiencies in our understanding of these elements, especially in Gram positive bacteria. Our work will focus on the lifecycle of ICEBs1 in B. subtilis. The ability to experimentally induce ICEBs1 in >90% of cells in a population and achieve relatively high conjugation frequencies will allow us to answer previously difficult or unstudied problems fundamental to the ICE lifecycle. We will also identify and analyze host genes needed for ICE function and study interactions between ICEs and other mobile elements in cells. We will extend our analyses to Tn916, a widespread ICE involved in the spread of tetracycline resistance. Our findings should be relevant to the transfer of genes between bacteria growing in many different environments, including the human microbiome. Our work will also focus on several aspects of chromosome dynamics and gene expression. Cells have multiple mechanisms for regulating the initiation of replication. Cells also have regulatory responses to perturbations in replication, often called checkpoints, which control gene expression and cell cycle progression. Coupling gene expression and cell cycle progression to chromosome replication and integrity helps prevent the generation of cells with defective chromosomes. The coordination of genome duplication with cell cycle progression is important for cellular differentiation and preventing uncontrolled cell growth. Microbial pathogenesis often depends on normal bacterial replication and growth in the host. We will use a variety of approaches and methodologies, both in vivo and in vitro, to characterize: the control of replication initiation; regulators of the replication initiator and transcription factor DnaA; and genes controlled in response to perturbations in replication. Understanding these processes in B. subtilis will provide insights regarding similar processes and homologous proteins in a wide variety of organisms, including many Gram positive pathogens.