Lawrence Berkeley National Laboratory

The Environmental Stress Pathway Project (ESPP) focuses on the systems-biology and environmental activity of the sulfate-reducing bacterium (SRB) Desulfovibrio vulgaris Hildenborough (DvH). However, focusing on a single representative species of a genus offers little insight into the evolution, niche adaptation, and function of an entire microbial genus. Given the broad environmental significance of SRBs and their phylogenetic diversity, it is imperative that other representative species of the Desulfovibrio genus are investigated. Therefore, we are developing an integrated functional genomics platform in the SRB Desulfovibrio desulfuricans G20 (G20). The platform is compromised of phenotypic arrays, gene expression arrays, and a transposon mutant library. The ability of G20 to utilize carbon sources and electron acceptors for growth, survive stress conditions, and respond to growth inhibitors is measured on a high-throughput system to narrow the test conditions to be used in both gene expression and mutant profiling assays. Using high-density multiplex microarrays, we are monitoring gene expression under a number of diverse conditions in a global effort to elucidate G20 gene regulation. Our principle effort is the construction of a genome-wide, tagged transposon mutant library in G20. A hallmark of our transposon library is the ability to pool thousands of mutants for parallel phenotypic analysis. Profiling our transposon library under a large number of conditions will enable us to link a large fraction of the G20 genome to specific cellular phenotypes. Here we present our experimental approaches and preliminary data from each type of experiment. Functional genomic investigations into G20 will be used by the ESPP in a number of ways. First, G20 and DvH have a large number of orthologous genes. Therefore, functional insights made in G20 are a strong starting point for targeted experiments in DvH. Second, by expanding our experimental ?pan-genome? to include G20, we are now investigating more of the natural variation present in the genus Desulfovibrio. These data will be imperative for annotating and analyzing newly sequenced SRB genomes. Lastly, our integrated functional genomics data will be used to build a cellular model of G20 metabolism, regulation, and activity. This model can be used as a framework for the interpretation of many types of ESPP data generated in the laboratory and in the field.