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Minor pilin subunits are conserved in Vibrio cholerae type IV pili
Claudia Toma1, Hiromi Kuroki, Noboru Nakasone
1Department of Bacteriology, Faculty of Medicine, University of the Ryukyus, Okinawa 903-0215, Japan. k950417@med.u-ryukyu.ac.jp
Abstract:
The nucleotide sequences of five open reading frames within the Vibrio cholerae NAGV14 type IV pilus gene cluster were determined. The genes showed high homology to the mannose-sensitive hemagglutinin (MSHA) pilus genes mshB, mshC, mshD, mshO and mshP. PCR analysis showed that a MSHA-like gene cluster is highly conserved among different V. cholerae strains, with the exception of the previously reported major pilin subunit. Recombinant MshB and MshO proteins were purified and specific antiserum was raised to each of them. Western blotting analyses showed that these antisera reacted with purified NAGV14 and MSHA pili. The results suggested that MshB and MshO are minor components of the pilus fiber. Although there was no cross-reaction between the major pilin subunits of NAGV14 and MSHA pili, minor components seemed to be highly homologous and immunologically cross-reactive.
Insights
The Vibrio cholerae NAGV14 type IV pilus gene cluster shares homology with MSHA pilus genes. Minor pilus components MshB and MshO are conserved and immunologically cross-reactive between strains.
Area of Science:
- Microbiology
- Molecular Biology
- Bacterial Genetics
Background:
- Type IV pili are important virulence factors in many bacteria, including Vibrio cholerae.
- The mannose-sensitive hemagglutinin (MSHA) pilus system is well-characterized in V. cholerae, but its components and conservation require further investigation.
- Understanding the genetic basis and protein composition of V. cholerae pili is crucial for developing targeted interventions.
Purpose of the Study:
- To determine the nucleotide sequences of open reading frames within the V. cholerae NAGV14 type IV pilus gene cluster.
- To investigate the homology and conservation of this gene cluster with known MSHA pilus genes.
- To identify and characterize minor pilus components and assess their immunological cross-reactivity.
Main Methods:
- Nucleotide sequencing of five open reading frames in the V. cholerae NAGV14 pilus gene cluster.
- Polymerase Chain Reaction (PCR) analysis to assess gene cluster conservation across V. cholerae strains.
- Recombinant protein expression, purification, and antiserum generation for MshB and MshO.
- Western blotting to analyze protein-antiserum interactions.
Main Results:
- The determined nucleotide sequences showed high homology to known MSHA pilus genes (mshB, mshC, mshD, mshO, mshP).
- PCR analysis confirmed that the MSHA-like gene cluster is conserved among V. cholerae strains, except for the major pilin subunit.
- Western blotting demonstrated that MshB and MshO are minor pilus components, highly homologous and immunologically cross-reactive, unlike the major pilin subunits.
Conclusions:
- The V. cholerae NAGV14 type IV pilus gene cluster is homologous to the MSHA pilus system.
- Minor pilus components MshB and MshO are conserved and immunologically cross-reactive, suggesting a conserved function.
- While major pilin subunits differ, conserved minor components may play a significant role in pilus structure or function across V. cholerae strains.