Cysteine Biosynthesis Controls Serratia marcescens Phospholipase Activity

Serratia marcescens causes health care-associated opportunistic infections that can be difficult to treat due to a high incidence of antibiotic resistance. One of the many secreted proteins ofS. marcescens is the PhlA phospholipase enzyme. Genes involved in the production and secretion of PhlA were identified by screening a transposon insertion library for phospholipase-deficient mutants on phosphatidylcholine-containing medium. Mutations were identified in four genes (cyaA ,crp ,fliJ , andfliP ) that are involved in the flagellum-dependent PhlA secretion pathway. An additional phospholipase-deficient isolate harbored a transposon insertion in thecysE gene encoding a predicted serineO -acetyltransferase required for cysteine biosynthesis. ThecysE requirement for extracellular phospholipase activity was confirmed using a fluorogenic phospholipase substrate. Phospholipase activity was restored to thecysE mutant by the addition of exogenousl -cysteine orO -acetylserine to the culture medium and by genetic complementation. Additionally,phlA transcript levels were decreased 6-fold in bacteria lackingcysE and were restored with added cysteine, indicating a role for cysteine-dependent transcriptional regulation ofS. marcescens phospholipase activity.S. marcescens cysE mutants also exhibited a defect in swarming motility that was correlated with reduced levels offlhD andfliA flagellar regulator gene transcription. Together, these findings suggest a model in which cysteine is required for the regulation of both extracellular phospholipase activity and surface motility inS. marcescens .IMPORTANCE Serratia marcescens is known to secrete multiple extracellular enzymes, but PhlA is unusual in that this protein is thought to be exported by the flagellar transport apparatus. In this study, we demonstrate that both extracellular phospholipase activity and flagellar function are dependent on the cysteine biosynthesis pathway. Furthermore, a disruption of cysteine biosynthesis results in decreasedphlA and flagellar gene transcription, which can be restored by supplying bacteria with exogenous cysteine. These results identify a previously unrecognized role for CysE and cysteine in the secretion ofS. marcescens phospholipase and in bacterial motility.