Novel mycoplasma nucleomodulin MbovP475 decreased cell viability by regulating expression of CRYAB and MCF2L2

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 242 NI 243 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.

MbovP0725, a secreted serine/threonine phosphatase, inhibits the host inflammatory response and affects metabolism in Mycoplasma bovis

“…Several secreted proteins have been confirmed to have virulence-related activities. For example, MbovP0145 has been shown to induce bovine lung epithelial cells (EBL) to produce IL-8 by interacting with β-actin ( 21 ), MbovP280 induced apoptosis of bovine macrophages ( 22 ), and MbovP475 with a conserved transcription activator-like effector decreased cell viability ( 23 ). Among these proteins, a differentially expressed protein MbovP0725 attracted attention due to its higher expression in a wild-type (WT) strain than the attenuated strain, indicating that it might play a role in the pathogenesis of M. bovis .…”

Intelligent-Responsive Enrofloxacin-Loaded Chitosan Oligosaccharide–Sodium Alginate Composite Core-Shell Nanogels for On-Demand Release in the Intestine

“…Subsequently, the E. coli ATCC 25922 was inoculated into each tube to achieve a final inoculum of 1 × 10 6 CFU/mL. The minimum inhibitory concentration (MIC) is the lowest concentration required for inhibiting the visible bacterial growth after 24 h incubation of the bacterial cultures at 37 °C [ 15 , 16 , 17 ].…”

Mycoplasma bovis mastitis in dairy cattle

“…Nucleomodulins are effector proteins secreted by bacteria that can interact with the host DNA and serve to regulate gene transcription to favor the pathogenesis of the bacteria ( 41 ). The MbovP475 lipoprotein is secreted by M. bovis and binds the promoters of the cell cycle central regulatory genes, CRYAB and MCF2L2 genes and downregulates their expression in bovine macrophage cell line ( 42 ) resulting in decreases in bovine macrophage cell line viability.…”

“…Screening of original herd before purchasing new cows is worthwhile as well as quarantine of new cows which adds extra layer of security. In addition, isolation and culling of infected cows are necessary measures to effectively control the disease and minimize outbreaks ( 42 ). However, advances in rapid and accessible tests to detect M. bovis on dairy farms are necessary to control and prevent outbreaks from occurring more effectively.…”