The hemolytic and cytolytic activities of Serratia marcescensphospholipase A (PhlA) depend on lysophospholipid production by PhlA

Shimuta, Ken; Ohnishi, Makoto; Iyoda, Sunao; Gotoh, Naomasa; Koizumi, Nobuo; Watanabe, Haruo

doi:10.1186/1471-2180-9-261

Cited by 53 publications

(47 citation statements)

References 42 publications

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“…In vitro experiments previously demonstrated that purified PhlA could induce cytotoxicity in human bladder and cervical epithelial cells (3), and the Y. enterocolitica PhlA homolog has been proposed to have a role in pathogenesis during murine gastrointestinal infection (14). The partial loss of fitness for bacteria lacking PhlA observed in this work is intriguing, though not definitive.…”

Section: Discussionmentioning

confidence: 99%

Cysteine Biosynthesis Controls Serratia marcescens Phospholipase Activity

2017

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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 of S. 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, and fliP) that are involved in the flagellum-dependent PhlA secretion pathway. An additional phospholipase-deficient isolate harbored a transposon insertion in the cysE gene encoding a predicted serine O-acetyltransferase required for cysteine biosynthesis. The cysE requirement for extracellular phospholipase activity was confirmed using a fluorogenic phospholipase substrate. Phospholipase activity was restored to the cysE mutant by the addition of exogenous L-cysteine or O-acetylserine to the culture medium and by genetic complementation. Additionally, phlA transcript levels were decreased 6-fold in bacteria lacking cysE and were restored with added cysteine, indicating a role for cysteine-dependent transcriptional regulation of S. marcescens phospholipase activity. S. marcescens cysE mutants also exhibited a defect in swarming motility that was correlated with reduced levels of flhD and fliA 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 in S. 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 decreased phlA 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 of S. marcescens phospholipase and in bacterial motility.

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Section: Discussionmentioning

confidence: 99%

Cysteine Biosynthesis Controls Serratia marcescens Phospholipase Activity

2017

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“…1C and 3A). Indeed, Shimuta et al have indicated that S. marcescens utilizes phospholipase A to exploit hemolytic and cytotoxic effects and potentially to regulate hemolysin ShlA activity (56). Intriguingly, synthesis of phospholipase is positively regulated by FlhDC in S. marcescens (21,37), indicating that the higher hemolytic and cytotoxic activities of S. marcescens ⌬rssBA-shlBA might be due to upregulation of flhDC expression, which further induces phospholipase production.…”

Section: Discussionmentioning

confidence: 99%

RssAB-FlhDC-ShlBA as a Major Pathogenesis Pathway in Serratia marcescens

et al. 2010

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Serratia marcescens has long been recognized as an important opportunistic pathogen, but the underlying pathogenesis mechanism is not completely clear. Here, we report a key pathogenesis pathway in S. marcescens comprising the RssAB two-component system and its downstream elements, FlhDC and the dominant virulence factor hemolysin ShlBA. Expression of shlBA is under the positive control of FlhDC, which is repressed by RssAB signaling. At 37°C, functional RssAB inhibits swarming, represses hemolysin production, and promotes S. marcescens biofilm formation. In comparison, when rssBA is deleted, S. marcescens displays aberrant multicellularity favoring motile swarming with unbridled hemolysin production. Cellular and animal infection models further demonstrate that loss of rssBA transforms this opportunistic pathogen into hypervirulent phenotypes, leading to extensive inflammatory responses coupled with destructive and systemic infection. Hemolysin production is essential in this context. Collectively, a major virulence regulatory pathway is identified in S. marcescens.The Gram-negative bacterium Serratia marcescens is an important opportunistic pathogen that causes a wide range of diseases and clinical presentations with high morbidity (25). S. marcescens frequently causes outbreaks in intensive and neonatal care units, and the occurrence of multiple-antibioticresistant strains has further exacerbated clinical treatment difficulties (3, 39). Despite years of study, the mechanism of pathogenesis of S. marcescens and why it behaves as an opportunistic pathogen remain poorly understood. Unraveling the underlying mechanism of pathogenesis is thus very important for developing strategies to prevent and treat S. marcescens infection.The hemolysin ShlA was shown to be a dominant virulence factor in S. marcescens pathogenesis using a murine lung infection model (35). ShlA is responsible for the hemolytic and cytotoxic effects on erythrocytes and cultured cells, with the aid of an outer membrane protein, ShlB (28,29,47,53,54). ShlA also contributes to the release of inflammatory mediators, increases uropathogenicity, and triggers microtubule-dependent invasion of S. marcescens into epithelial cells (27,30,34,40). However, the mechanism by which the expression of shlA is regulated, especially in response to any bacterial signaling system control, remains uncharacterized. Only one reported study has indicated that iron is involved in the regulation of shlBA expression in S. marcescens (46). S. marcescens exhibits swarming, which is recognized as a highly coordinated multicellular surface migration behavior (24,51,62) that is correlated with virulence capability, antibiotic resistance, and hemolysin production in other bacteria (1,17,44). S. marcescens swarms on Luria-Bertani (LB) agar surfaces at 30°C, but not at 37°C (36). Our previous studies showed that activation of a bacterial two-component system, RssAB, comprising a sensor kinase, RssA, and a response regulator, RssB, inhibited swarming and reduced hemolysin productio...

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“…While the ShlAB hemolysin of S. marcescens is contact dependent, an extracellular hemolysin was described in 1989 and was recently characterized (153,351). This hemolysin, PhlA, has phospholipase A activity (351).…”

Section: Enzymes Produced By Serratia Speciesmentioning

confidence: 99%

“…This hemolysin, PhlA, has phospholipase A activity (351). PhlA does not apparently have direct cytolytic activity; however, it acts upon phospholipid and produces lysophospholipid, which was cytolytic for human, horse, and sheep red blood cells and the HeLa and 5637 cell lines (351).…”

Section: Enzymes Produced By Serratia Speciesmentioning

confidence: 99%

Serratia Infections: from Military Experiments to Current Practice

2011

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SUMMARY Serratia species, in particular Serratia marcescens , are significant human pathogens. S. marcescens has a long and interesting taxonomic, medical experimentation, military experimentation, and human clinical infection history. The organisms in this genus, particularly S. marcescens , were long thought to be nonpathogenic. Because S. marcescens was thought to be a nonpathogen and is usually red pigmented, the U.S. military conducted experiments that attempted to ascertain the spread of this organism released over large areas. In the process, members of both the public and the military were exposed to S. marcescens , and this was uncovered by the press in the 1970s, leading to U.S. congressional hearings. S. marcescens was found to be a certain human pathogen by the mid-1960s. S. marcescens and S. liquefaciens have been isolated as causative agents of numerous outbreaks and opportunistic infections, and the association of these organisms with point sources such as medical devices and various solutions given to hospitalized patients is striking. Serratia species appear to be common environmental organisms, and this helps to explain the large number of nosocomial infections due to these bacteria. Since many nosocomial infections are caused by multiply antibiotic-resistant strains of S. marcescens , this increases the danger to hospitalized patients, and hospital personnel should be vigilant in preventing nosocomial outbreaks due to this organism. S. marcescens , and probably other species in the genus, carries several antibiotic resistance determinants and is also capable of acquiring resistance genes. S. marcescens and S. liquefaciens are usually identified well in the clinical laboratory, but the other species are rare enough that laboratory technologists may not recognize them. 16S rRNA gene sequencing may enable better identification of some of the less common Serratia species.

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The hemolytic and cytolytic activities of Serratia marcescensphospholipase A (PhlA) depend on lysophospholipid production by PhlA

Cited by 53 publications

References 42 publications

Cysteine Biosynthesis Controls Serratia marcescens Phospholipase Activity

Cysteine Biosynthesis Controls Serratia marcescens Phospholipase Activity

RssAB-FlhDC-ShlBA as a Major Pathogenesis Pathway in Serratia marcescens

Serratia Infections: from Military Experiments to Current Practice

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