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Conjugative mobilization of the cloned M6 protein gene from Streptococcus pneumoniae to Streptococcus pyogenes
1Institute of Microbiology, University of Verona, Italy.
Abstract:
The host-vector system omega 6001-pDP36 was used to transfer the M6 protein gene (emm-6.1) of Streptococcus pyogenes to other S. pyogenes strains, isogenic and nonisogenic to D471, the strain from which emm-6.1 was originally cloned. The first step was to subclone emm-6.1 into the insertion vector pDP36. The resulting plasmid, pRMB20, was used as donor in transformation to insert emm-6.1 into the conjugative transposon omega 6001. Streptococcus pneumoniae DP1322, carrying omega 6001 integrated into the chromosome, was the recipient in the transformation experiment. omega 6001 containing emm-6.1 was then transferred by conjugation from S. pneumoniae to the chromosomes of M+ and M- S. pyogenes strains. S. pyogenes transconjugants contained one intact copy of emm-6.1 integrated into the chromosome, but no expression of M6 protein could be detected by Western blot analysis. We found no evidence of the positive transacting regulation of emm gene expression postulated by other authors. In fact, the cloned emm-6.1 was not expressed in three strains expressing their own M proteins (M5, M17 and a shorter M6). In these partial diploids M protein genes were expressed only when present in the original chromosomal locus.
Insights
Transferring the M6 protein gene (emm-6.1) of Streptococcus pyogenes did not result in M6 protein expression in recipient strains. This suggests M6 protein gene expression is regulated by its original chromosomal locus.
Area of Science:
- Microbiology
- Molecular Biology
- Genetics
Background:
- Streptococcus pyogenes possesses surface M proteins crucial for virulence.
- Understanding the regulation of M protein gene expression is vital for controlling S. pyogenes infections.
Purpose of the Study:
- To investigate the transferability and expression of the M6 protein gene (emm-6.1) from Streptococcus pyogenes.
- To determine if emm-6.1 expression is regulated by trans-acting factors or its native chromosomal location.
Main Methods:
- Subcloning the emm-6.1 gene into insertion and conjugative transposon vectors.
- Transformation and conjugation experiments to transfer emm-6.1 into various S. pyogenes strains.
- Western blot analysis to detect M6 protein expression.
Main Results:
- The emm-6.1 gene was successfully integrated into the chromosomes of recipient S. pyogenes strains.
- No M6 protein expression was detected in any transconjugant strains via Western blot.
- The cloned emm-6.1 gene was not expressed in strains already expressing other M proteins or a different M6 variant.
Conclusions:
- The expression of the emm-6.1 gene in Streptococcus pyogenes is not driven by positive trans-acting regulation.
- M protein gene expression appears to be dependent on its original chromosomal locus, suggesting cis-acting regulatory mechanisms.