The role of CsrA in controls the extracellular electron transfers in Geobacter sulfurreducens
CsrA is a post-transcriptional regulator that controls biofilm formation, virulence, carbon metabolism, and motility, among other phenotypes in bacteria. Although CsrA has been extensively studied in ?-proteobacteria and firmicute, it is unknown which cellular processes are targeted for regulation in d-proteobacteria. In this work, we constructed and characterized the ?csrA mutant strain in Geobacter sulfurreducens to determine the involvement of the CsrA protein in the regulation of biofilm and extracellular electron transfer. The ?csrA mutant strain grows on acetate-fumarate and reduces soluble Fe(III) similarly to the wild type strain, but has higher rates of insoluble Fe(III) reduction than the wild type. Quantification and characterization of biofilms by violet staining and confocal laser scanning microscopy showed that the ?csrA strain produces up to twice as much biofilm as the wild type strain, containing more than 95% viable cells. Transcriptome analysis by RNA-seq shows that in ?csrA biofilms developed on an inert support, 244 (103 upregulated and 141 downregulated) genes changed their expression, including those related to extracellular electron transfer, exopolysaccharide synthesis, c-di-GMP synthesis and degradation. To validate the transcriptome data, RT-qPCR confirmed the differential expression of several selected genes in the ?csrA strain. Current production in microbial fuel cells was tested and the ?csrA strain was found to produce 45-50% more current than the wild type. To verify the genes that changed expression in the graphite electrodes in the ?csrA strain, a transcriptome analysis was performed. 181 genes changed their expression in the ?csrA biofilms of which 113 genes showed differentially expression only in MFC and 68 genes changed their expression as well as transcriptome from biofilms grown on inert support. In silico analysis of the 5'-UTR regions revealed that 76 genes that changed expression in the RNA-seq analysis have a consensus sequence for CsrA binding. This is the first report describing the involvement of CsrA in the regulation of extracellular electron transfer and biofilm in a member of the ?-proteobacteria. Overall design: Comparative gene expression profiling analysis of RNA-seq data for Geobacter sulfurreducens biofilm from wild type and csrA-mutant strains grown on a glass support and graphite electrode. From each sample, experimental duplicates were used.