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Transcriptional regulation of the Yersinia pseudotuberculosis pH6 antigen adhesin by two envelope-associated
1Department of Molecular Biology and Microbiology, Tufts University School of medicine, Boston, Massachusetts 02111, USA.
Abstract:
The Yersinia pseudotuberculosis pH6 antigen mediates haemagglutination and adhesion to cultured mammalian cells. The synthesis of pH6 antigen requires the products of the psaEFABC genes in both Yersinia pseudotuberculosis and Escherichia coli. In-frame deletion mutations of psaE and psaF caused defective haemagglutination. In contrast, we showed that the psaABC genes were sufficient for haemagglutination if they were expressed by a heterologous promoter. Environmental regulation of pH6 antigen by temperature and pH occurs via regulation of the major pilus protein PsaA at the transcriptional level. Northern blot analyses indicate that the psaA transcript was absent in either psaE or psaF mutant strains. Primer extension analyses indicate that, in Y. pseudotuberculosis, the transcription of the psaE and psaF genes is constitutive. Alkaline phosphatase fusion studies confirm the topology prediction that PsaE and PsaF are both inner-membrane-associated proteins. PsaE consists of an N-terminal cytoplasmic domain, containing sequence similarity to transcriptional regulators found in two-component systems as well as to the Salmonella typhimurium HIIA protein, with a C-terminal domain that is periplasmically localized. PsaF is predicted to be oriented with most of the protein in the periplasm, the hydrophobic N-terminus being either integrated in the inner membrane or cleaved as a signal peptide.
Insights
The Yersinia pseudotuberculosis pH6 antigen, crucial for bacterial adhesion and hemagglutination, requires psaEFABC genes. PsaABC genes alone can mediate hemagglutination, with environmental factors regulating PsaA transcription.
Area of Science:
- Microbiology
- Bacterial Pathogenesis
- Molecular Biology
Background:
- The pH6 antigen of Yersinia pseudotuberculosis facilitates bacterial adhesion and hemagglutination.
- Synthesis of the pH6 antigen is dependent on the psaEFABC gene products in both Y. pseudotuberculosis and Escherichia coli.
Purpose of the Study:
- To investigate the roles of psaEFABC genes in pH6 antigen synthesis and function.
- To elucidate the environmental regulation of pH6 antigen expression.
Main Methods:
- In-frame deletion mutagenesis to create psaE and psaF mutants.
- Expression of psaABC genes in a heterologous system.
- Northern blot analysis to assess psaA transcript levels.
- Primer extension analysis to determine transcription start sites.
- Alkaline phosphatase fusion studies for protein topology prediction.
Main Results:
- Deletion mutations in psaE and psaF impaired hemagglutination.
- The psaABC genes were sufficient for hemagglutination when expressed from a heterologous promoter.
- Environmental factors (temperature, pH) regulate pH6 antigen via transcriptional control of the major pilus protein PsaA.
- The psaA transcript was absent in psaE or psaF mutants.
- PsaE and PsaF are inner-membrane-associated proteins, with PsaE having a cytoplasmic N-terminal domain and periplasmic C-terminal domain, and PsaF primarily localized to the periplasm.
Conclusions:
- The psaABC genes are essential and sufficient for pH6 antigen-mediated hemagglutination.
- PsaE and PsaF play regulatory roles in pH6 antigen synthesis, likely at the transcriptional level of PsaA.
- Environmental cues modulate bacterial adhesion through the regulation of PsaA expression.