The Effect of CsrA on Biofilm Development in Escherichia coli

Jackson, Debra W., The Effect of CsrA on Biofilm Development in Escherichia coli. Doctor of Philosophy (Biomedical Sciences), May 2001, 127 pp., 2 tables, 15 illustrations, bibliography, 138 titles. CsrA, carbon storage regulator, is a small RNA-binding protein that acts as a global regulator and modulates specific mRNA stability in Escherichia coli. CsrA regulates central carbon metabolism in addition to flagella biogenesis. In this study, the phylogenetic distribution of csrA and its role in Escherichia coli biofilm development were examined. CsrA homologs were examined using Southern hybridization experiment and by analyzing existing sequencing data and was found to be widespread among eubacteria. CsrA was shown to be capable of acting as a genetic switch for biofilm formation and dispersal. A csrA mutant of E. coli was shown to increase biofilm formation and exhibit apparent pillars and channels characteristic of a mature biofilm. Over-expression of csrA completely inhibited biofilm formation in E. coli K-12 and decreased biofilm formation in related enteric pathogens. Induction of csrA expression from a multicopy plasmid caused dispersal of a pre-formed biofilm. Gene expression studies revealed that csrA expression is dynamically regulated during biofilm formation. Several outer-membrane factors and global regulators that have been implicated in biofilm formation were examined for effects on biofilm formation in a csrA mutant. Crystal violet adherence assays revealed that flagella and type I pili affect biofilm formation in a scrA mutant strain; however, colonic acid and curli fimbriae did not exhibit quantitative effects on biofilm formation in the csrA mutant, but the stationary phase sigma factor, RpoS, had no quantitative effect on csrA mutant biofilm formation. Therefore, a csrA mutant will form a biofilm in the absence of each of these outer-membrane factors and global regulatory factors of biofilm formation. The effects of csrA on biofilm formation were found to be mediated in part through its effects on intracellular glycogen metabolism. Thus the redirection of carbon flux, in response to environmental and/or physiological cues, is important for biofilm development.