CHARACTERIZATION OF THE REGULATION MECHANISMS OF ENTEROBACTERIAL COMMON ANTIGEN (ECA) BIOSYNTHESIS IN Serratia marcescens.

MEDIAVILLA GABRIELA; STEPANENKO, TATIANA; CASTELLI, MARÍA EUGENIA; GARCÍA VESCCOVI ELEONORA

Congreso; IX Congreso de Microbiología general; 2013

Serratia is a gram negative enteric bacterium that presents a highly conserved enterobacterial common antigen (ECA) and an RcsF-RcsCDB phosphorelay that were shown to be implicated in regulation of flagellar synthesis by our group. Our results showed that the deficiency in the enterobacterial common antigen (ECA) exopolysaccharide promotes the activity of RcsCDB, thereby controlling flagellar biogenesis and modulating the swimming and swarming motile phenotypes of Serratia. By bioinformatic screening we found an RcsB binding consensus motif in the promoter region of wec cluster genes. The analysis of wec cluster mutant strains indicated that the absence of the periplasmic ECA cyclic structure could constitute a signal detected by the Serratia Rcs phosphorelay. Adding to this, osmotic stress was established to be a signal modulating the activity of the Rcs system, suggesting that ECAcyc might play a role in the osmotic balance of this cellular compartment. To further investigate the relationship of ECA biosynthetic pathway and the Rcs system in Serratia, we analyzed the transcriptional expression from the wecA promoter in an rcsB background and determined ECAcyc levels by LC-MS analysis in wzzE, wecD, rcsB mutants grown in liquid media containing different NaCl concentrations or in agar plates. As a control of Rcs activity in different NaCl concentrations we determined flagellin levels by western blot. Activity of the wecA promoter was decreased as NaCl concentrations raised in wild type (Wt) strain and it was decreased and not osmotically regulated in rcsB mutant. As expected, wzzE and wecD mutants did not contain detectable ECAcyc. Instead, ECAcyc levels were not subjected to osmoregulation in the rcsB mutant strain while Wt Serratia presented higher levels when grown without NaCl compared to LB, similarly to what was observed for the wecA promoter activity. Unsurprisingly, ECAcyc levels correlated with the wecA promoter activity. These results indicate that Rcs system modulates ECA biosynthesis in an osmoregulated pattern, and provide a new insight in the role of Rcs in the control of the expression of essential envelop components of the bacterial cell.