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Generation and surface localization of intact M protein in Streptococcus pyogenes are dependent on sagA
1Department of Microbiology and Immunology, Emory University School of Medicine, Atlanta, Georgia 30322, USA.
Abstract:
The M protein is an important surface-located virulence factor of Streptococcus pyogenes, the group A streptococcus (GAS). Expression of M protein is primarily controlled by Mga, a transcriptional activator protein. A recent report suggested that the sag locus, which includes nine genes necessary and sufficient for production of streptolysin S, another GAS virulence factor, is also needed for transcription of emm, encoding the M protein (Z. Li, D. D. Sledjeski, B. Kreikemeyer, A. Podbielski, and M. D. Boyle, J. Bacteriol. 181:6019-6027, 1999). To investigate this in more detail, we constructed an insertion-deletion mutation in sagA, the first gene in the sag locus, in the M6 strain JRS4. The resulting strain, JRS470, produced no detectable streptolysin S and showed a drastic reduction in cell surface-associated M protein, as measured by cell aggregation and Western blot analysis. However, transcription of the emm gene was unaffected by the sagA mutation. Detailed analysis with monoclonal antibodies and an antipeptide antibody showed that the M protein in the sagA mutant strain was truncated so that it lacks the C-repeat region and the C-terminal domain required for anchoring it to the cell surface. This truncated M protein was largely found, as expected, in the culture supernatant. Lack of surface-located M protein made the sagA mutant strain susceptible to phagocytosis. Thus, although sagA does not affect transcription of the M6 protein gene, it is needed for the surface localization of this important virulence factor.
Insights
The sagA gene is crucial for Streptococcus pyogenes M protein surface localization, not its transcription. A mutation in sagA results in truncated M protein, increasing susceptibility to phagocytosis.
Area of Science:
- Microbiology
- Molecular Biology
- Immunology
Background:
- Streptococcus pyogenes (GAS) M protein is a key surface virulence factor.
- M protein expression is regulated by the Mga transcriptional activator.
- The sag locus, involved in streptolysin S production, was hypothesized to regulate M protein transcription.
Purpose of the Study:
- To investigate the role of the sag locus, specifically sagA, in M protein regulation.
- To determine if sagA affects M protein gene (emm) transcription or surface localization.
Main Methods:
- Constructed a sagA insertion-deletion mutant (JRS470) in GAS strain JRS4.
- Assessed streptolysin S production and M protein levels via cell aggregation and Western blot.
- Analyzed M protein transcription, structure, and localization using antibodies and Western blot.
- Evaluated phagocytosis susceptibility of the sagA mutant.
Main Results:
- The sagA mutant produced no streptolysin S and showed reduced cell surface M protein.
- emm gene transcription was unaffected by the sagA mutation.
- M protein in the sagA mutant was truncated, lacking the C-repeat and anchoring domains, and secreted into the supernatant.
- The sagA mutant lacking surface M protein was susceptible to phagocytosis.
Conclusions:
- The sagA gene is essential for the proper surface localization and anchoring of M protein in Streptococcus pyogenes.
- sagA does not regulate emm gene transcription but is critical for post-translational modification or trafficking of M protein.
- Disruption of M protein surface localization compromises GAS virulence by increasing susceptibility to phagocytosis.