The Mycoplasma gallisepticum Virulence Factor Lipoprotein MslA Is a Novel Polynucleotide Binding Protein

Masukagami, Yumiko; Tivendale, Kelly A.; Mardani, Karim; Ben-Barak, Idan; Markham, Philip F.; Browning, Glenn F.

doi:10.1128/iai.00365-13

Cited by 25 publications

(22 citation statements)

References 29 publications

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“…MBOVPG45_0310 and its orthologues in most members of the M. hominis and M. pneumoniae phylogenic clusters lie within a putative nucleotide transporter operon (12,13,16,29,30), and the nuclease activity has been demonstrated for M. agalactiae MAG_5040 (16), M. gallisepticum MGA_0676 (13), M. hyopneumoniae mhp379 (12), M. pneumoniae MPN133 (15), and M. genitalium MG186 (14). It has recently been demonstrated that MslA, the MBOVPG45_0311 homologue in M. gallisepticum, binds single-and double-stranded DNA, suggesting that it may act in concert with these nucleases, with MslA binding and delivering oligonucleotides to the exonuclease, which then processes the oligonucleotides to generate individual nucleotides for transport into the cell via the ABC transporter (13).…”

Section: Discussionmentioning

confidence: 99%

“…It has recently been demonstrated that MslA, the MBOVPG45_0311 homologue in M. gallisepticum, binds single-and double-stranded DNA, suggesting that it may act in concert with these nucleases, with MslA binding and delivering oligonucleotides to the exonuclease, which then processes the oligonucleotides to generate individual nucleotides for transport into the cell via the ABC transporter (13).…”

Section: Discussionmentioning

confidence: 99%

“…The predicted molecular mass of the product of MBOVPG45_0215 is 44.13 kDa, that of the product of MBOVPG45_0310 is 41.56 kDa, and that of the product of MBOVPG45_0089 is 20.51 kDa. Mapping of a library of 319 transposon mutants of the M. bovis PG45 genome identified 1 mutant (the ⌬0310 mutant) with a disruption in MBOVPG45_0310, which codes for the nuclease of a putative nucleotide transporter operon (12,13,29,30). Another mutant (the ⌬0215 mutant) had a disruption in MBOVPG45_0215, which encodes a membrane nuclease, MnuA.…”

Section: Identification Of Disruptions In Nuclease Genesmentioning

confidence: 99%

“…Orthologues of the three annotated putative membrane nucleases in the genome of M. bovis strain PG45, MBOVPG45_0089, MBOVPG45_0215, and MBOVPG45_0310, were identified in a number of phylogenetically distant mycoplasmas belonging to the M. hominis and M. pneumoniae clusters and are listed in Table 1 Genomic location and features of membrane nucleases in M. bovis. MBOVPG45_0310 is part of a 7.6-kbp ABC transporter operon that is predicted to be involved in nucleotide transport (12,13,16,29,30). However, MBOVPG45_0089 and MBOVPG45_0215 do not appear to lie within operons.…”

Section: Identification Of Disruptions In Nuclease Genesmentioning

confidence: 99%

“…Nuclease activity has also been demonstrated in vitro following cloning, expression, and purification of predicted membrane nucleases from M. hyopneumoniae (12), M. gallisepticum (13), M. genitalium (14), M. pneumoniae (15), and M. agalactiae (16). However, most studies on membrane nucleases of mycoplasmas have been based on an examination of in vitro profiles of activity following protein purification (12)(13)(14)(15)(16). Genome sequencing has identified genes encoding many additional putative nucleases in mycoplasmas that are predicted to be located either on the cell surface or within the cytoplasm, but these studies cannot assess the relative significance of a particular nuclease in the cell.…”

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Disruption of the Membrane Nuclease Gene (MBOVPG45_0215) of Mycoplasma bovis Greatly Reduces Cellular Nuclease Activity

et al. 2015

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Although the complete genome sequences of three strains of Mycoplasma bovis are available, few studies have examined gene function in this important pathogen. Mycoplasmas lack the biosynthetic machinery for the de novo synthesis of nucleic acid precursors, so nucleases are likely to be essential for them to acquire nucleotide precursors. Three putative membrane nucleases have been annotated in the genome of M. bovis strain PG45, MBOVPG45_0089 and MBOVPG45_0310, both of which have the thermonuclease (TNASE_3) functional domain, and MBOVPG45_0215 (mnuA), which has an exonuclease/endonuclease/phosphatase domain. While previous studies have demonstrated the function of TNASE_3 domain nucleases in several mycoplasmas, quantitative comparisons of the contributions of different nucleases to cellular nuclease activity have been lacking. Mapping of a library of 319 transposon mutants of M. bovis PG45 by direct genome sequencing identified mutants with insertions in MBOVPG45_0310 (the ⌬0310 mutant) and MBOVPG45_0215 (the ⌬0215 mutant). In this study, the detection of the product of MBOVPG45_0215 in the Triton X-114 fraction of M. bovis cell lysates, its cell surface exposure, and its predicted signal peptide suggested that it is a surface-exposed lipoprotein nuclease. Comparison of a ⌬mnuA mutant with wild-type M. bovis on native and denatured DNA gels and in digestion assays using double-stranded phage DNA and closed circular plasmid DNA demonstrated that inactivation of this gene abolishes most of the cellular exonuclease and endonuclease activity of M. bovis. This activity could be fully restored by complementation with the wild-type mnuA gene, demonstrating that MnuA is the major cellular nuclease of M. bovis. IMPORTANCENucleases are thought to be important contributors to virulence and crucial for the maintenance of a nutritional supply of nucleotides in mycoplasmas that are pathogenic in animals. This study demonstrates for the first time that of the three annotated cell surface nuclease genes in an important pathogenic mycoplasma, the homologue of the thermostable nuclease identified in Gram-positive bacteria is responsible for the majority of the nuclease activity detectable in vitro.A lthough the complete genomes of many Mycoplasma species are now available, the scarcity of tools for exploring their molecular biology has resulted in an understanding of the function of their genes more rudimentary than that which is available for many other prokaryotes. The genomes of three strains of the important bovine pathogen Mycoplasma bovis, the type strain PG45, the Hubei-1 strain, and the HB0801 strain, have been sequenced and published recently (1-3), but apart from several reports on phase switching in M. bovis (4-7), few studies have examined the molecular mechanisms underlying their pathogenicity and virulence or the metabolic functions of the products of their genes.Mycoplasmas evolved from Clostridium-like bacteria by a process of reductive evolution and are believed to have retained only the regulatory and ...

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Identification Of Disruptions In Nuclease Genesmentioning

confidence: 99%

Section: Identification Of Disruptions In Nuclease Genesmentioning

confidence: 99%

mentioning

confidence: 99%

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Disruption of the Membrane Nuclease Gene (MBOVPG45_0215) of Mycoplasma bovis Greatly Reduces Cellular Nuclease Activity

et al. 2015

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Mycoplasmosis

Ferguson‐Noel¹,

Armour²,

Noormohammadi³

et al. 2019

Diseases of Poultry

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Mycoplasma gallisepticum MGA_0676 is a membrane-associated cytotoxic nuclease with a staphylococcal nuclease region essential for nuclear translocation and apoptosis induction in chicken cells

Teng

Jiang

et al. 2014

Appl Microbiol Biotechnol

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Mycoplasma gallisepticum can infect a wide variety of birds including the commercial poultry. M. gallisepticum MGA_0676 is a putative lipoprotein, which is similar to bacterial thermostable nucleases. But the possible pathogenic effect of M. gallisepticum MGA_0676 has not been investigated so far. In the present study, we cloned the MGA_0676 gene after deletion of the amino-terminal signal sequence and mutagenesis of the Mycoplasma TGA tryptophan codons to TGG and expressed recombinant MGA_0676 protein in Escherichia coli. We identified and characterized MGA_0676 as a Ca(2+)-dependent cytotoxic nuclease of M. gallisepticum with a staphylococcal nuclease (SNc) region that displays the hallmarks of nucleases. Membrane protein immunoblot analysis and immunogold electron microscopy revealed that MGA_0676 locates on the membrane surface of M. gallisepticum. Furthermore, apoptosis assay using annexin V-FITC and propidium iodide (annexin V/PI) indicated that MGA_0676 played significant roles in apoptosis induction and pathological damages in chicken cells. Moreover, confocal microscopy showed that MGA_0676 localizes in the nuclei of host cells. Besides, after the SNc region was deleted, MGA_0676 lost its ability of nuclear localization, nuclease activity, and cytotoxicity, which revealed that the SNc region is essential for nuclear translocation and induction of apoptosis in chicken cells. The above results suggest that MGA_0676 is an important virulence factor in cellular pathology and may play a unique role in the life cycle events of M. gallisepticum.

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The Mycoplasma gallisepticum Virulence Factor Lipoprotein MslA Is a Novel Polynucleotide Binding Protein

Cited by 25 publications

References 29 publications

Disruption of the Membrane Nuclease Gene (MBOVPG45_0215) of Mycoplasma bovis Greatly Reduces Cellular Nuclease Activity

Disruption of the Membrane Nuclease Gene (MBOVPG45_0215) of Mycoplasma bovis Greatly Reduces Cellular Nuclease Activity

Mycoplasmosis

Mycoplasma gallisepticum MGA_0676 is a membrane-associated cytotoxic nuclease with a staphylococcal nuclease region essential for nuclear translocation and apoptosis induction in chicken cells

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