Högskolan i Skövde

The global regulator CsrA (carbon storage regulator) is an RNA binding protein that coordinates centralcarbon metabolism, activates flagellum biosynthesis and motility, and represses biofilm formation in Escherichiacoli. CsrA activity is antagonized by the untranslated RNA CsrB, to which it binds and forms a globularribonucleoprotein complex. CsrA indirectly activates csrB transcription, in an apparent autoregulatory mechanism.In the present study, we elucidate the intermediate regulatory circuitry of this system. Mutationsaffecting the BarA/UvrY two-component signal transduction system decreased csrB transcription but did notaffect csrA-lacZ expression. The uvrY defect was severalfold more severe than that of barA. Both csrA and uvrYwere required for optimal barA expression. The latter observation suggests an autoregulatory loop for UvrY.Ectopic expression of uvrY suppressed the csrB-lacZ expression defects caused by uvrY, csrA, or barA mutations;csrA suppressed csrA or barA defects; and barA complemented only the barA mutation. Purified UvrY proteinstimulated csrB-lacZ expression approximately sixfold in S-30 transcription-translation reactions, revealing adirect effect of UvrY on csrB transcription. Disruption of sdiA, which encodes a LuxR homologue, decreased theexpression of uvrY-lacZ and csrB-lacZ fusions but did not affect csrA-lacZ. The BarA/UvrY system activatedbiofilm formation. Ectopic expression of uvrY stimulated biofilm formation by a csrB-null mutant, indicative ofa CsrB-independent role for UvrY in biofilm development. Collectively, these results demonstrate that uvrYresides downstream from csrA in a signaling pathway for csrB and that CsrA stimulates UvrY-dependentactivation of csrB expression by BarA-dependent and -independent mechanisms.