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Updated: Aug 28, 2025
Optimized PCR-based Detection of Mycoplasma
Published on: June 20, 2011
Novel mycoplasma nucleomodulin MbovP475 decreased cell viability by regulating expression of CRYAB and MCF2L2
Gang Zhao1,2,3, Doukun Lu1,2, Shujuan Wang1,2
1National Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan, China.
Abstract:
Nucleomodulins are secreted bacterial proteins whose molecular targets are located in host cell nuclei. They are gaining attention as critical virulence factors that either modify the epigenetics of host cells or directly regulate host gene expression. Mycoplasma bovis is a major veterinary pathogen that secretes several potential virulence factors. The aim of this study was to determine whether any of their secreted proteins might function as nucleomodulins. After an initial in silico screening, the nuclear localization of the secreted putative lipoprotein MbovP475 of M. bovis was demonstrated in bovine macrophage cell line (BoMac) experimentally infected with M. bovis. Through combined application of ChIP-seq, Electrophoretic mobility shift assay (EMSA) and surface plasmon resonance (SPR) analysis, MbovP475 was determined to bind the promoter regions of the cell cycle central regulatory genes CRYAB and MCF2L2. MbovP475 has similar secondary structures with the transcription activator-like effectors (TALEs). Screening of various mutants affecting the potential DNA binding sites indicated that the residues 242NI243 within MbovP475 loop region of the helix-loop-helix domain were essential to its DNA binding activity, thereby contributing to decrease in BoMac cell viability. In conclusion, this is the first report to confirm M. bovis secretes a conserved TALE-like nucleomodulin that binds the promoters of CRYAB and MCF2L2 genes, and subsequently down-regulates their expression and decreases BoMac cell viability. Therefore, this study offers a new understanding of mycoplasma pathogenesis.
Insights
Mycoplasma bovis secretes a TALE-like nucleomodulin, MbovP475, that targets host cell nuclei. This protein binds specific gene promoters, reducing cell viability and offering new insights into mycoplasma pathogenesis.
Area of Science:
- Bacteriology
- Molecular Biology
- Veterinary Pathology
Background:
- Nucleomodulins are bacterial virulence factors targeting host cell nuclei.
- Mycoplasma bovis is a significant veterinary pathogen with unknown nucleomodulin activity.
Purpose of the Study:
- To investigate if Mycoplasma bovis secretes nucleomodulins.
- To characterize the function and targets of a potential nucleomodulin from M. bovis.
Main Methods:
- In silico screening and nuclear localization assays in bovine macrophages (BoMac).
- Chromatin immunoprecipitation sequencing (ChIP-seq), Electrophoretic mobility shift assay (EMSA), and Surface Plasmon Resonance (SPR).
- Mutagenesis studies to identify DNA binding residues.
Main Results:
- The secreted M. bovis protein MbovP475 localizes to the nucleus of infected BoMac cells.
- MbovP475 binds the promoter regions of cell cycle genes CRYAB and MCF2L2.
- Specific residues (242NI243) in MbovP475 are crucial for DNA binding, leading to decreased BoMac cell viability.
Conclusions:
- This study identifies a TALE-like nucleomodulin secreted by M. bovis.
- MbovP475 down-regulates CRYAB and MCF2L2 expression, impacting host cell viability.
- This finding advances understanding of Mycoplasma pathogenesis mechanisms.
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