Global transcriptional analysis ofGeobacter sulfurreducens gsu1771mutant biofilm grown on two different support structures

Electroactive biofilms formation by the metal-reducing bacterium Geobacter sulfurreducens is a crucial for bioelectricity generation and bioremediation. The transcriptional regulator GSU1771 controls the expression of essential genes involved in electron transfer and biofilm formation in G. sulfurreducens , with GSU1771-deficient producing thicker and more electroactive biofilms. Here, RNA-seq analyses were conducted to compare the global gene expression patterns of wild-type and Δ gsu1771 mutant biofilms grown on non-conductive (glass) and conductive (graphite electrode) materials. The Δ gsu1771 biofilm grown on the glass surface exhibited 467 differentially expressed (DE) genes (167 upregulated and 300 downregulated) versus the wild-type biofilm. In contrast, the Δ gsu1771 biofilm grown on the graphite electrode exhibited 119 DE genes (79 upregulated and 40 downregulated) versus the wild-type biofilm. Among these DE genes, 67 were also differentially expressed in the Δ gsu1771 biofilm grown on glass (56 with the same regulation and 11 exhibiting counter-regulation). Among the upregulated genes in the Δ gsu1771 biofilms, we identified potential target genes involved in exopolysaccharide synthesis ( gsu1961-63 , gsu1959 , gsu1972-73 , gsu1976-77 ). RT-qPCR analyses were then conducted to confirm the differential expression of a selection of genes of interest. DNA-protein binding assays demonstrated the direct binding of the GSU1771 regulator to the promoter region of pgcA , pulF , relA , and gsu3356 . Furthermore, heme-staining and western blotting revealed an increase in c -type cytochromes including OmcS and OmcZ in Δ gsu1771 biofilms. Collectively, our findings demonstrated that GSU1771 is a global regulator that controls extracellular electron transfer and exopolysaccharide synthesis in G. sulfurreducens , which is crucial for electroconductive biofilm development. Importance Biofilm formation is a multi-stage process that is finely coordinated by signal transduction and complex gene regulation mechanisms. Given the importance of biofilms and their biotechnological applications, understanding these processes in G. sulfurreducens is of great significance. Here, we studied the transcriptional profile of the Δ gsu1771 strain biofilms formed on two different supporting materials: (1) glass, a non-conductive surface and (2) a graphite electrode-based microbial fuel cell (MFC), which enabled us to assess the transcriptional responses of this strain during current production. By analyzing these two conditions, our study elucidated genes of interest that could be essential for biofilm production and extracellular electron transfer (EET) and provides new insights into the mechanisms that control these complex processes in G. sulfurreducens .