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Mammalian myosin-18A, a highly divergent myosin.

ABSTRACT: The Mus musculus myosin-18A gene is expressed as two alternatively spliced isoforms, ? and ?, with reported roles in Golgi localization, in maintenance of cytoskeleton, and as receptors for immunological surfactant proteins. Both myosin-18A isoforms feature a myosin motor domain, a single predicted IQ motif, and a long coiled-coil reminiscent of myosin-2. The myosin-18A? isoform, additionally, has an N-terminal PDZ domain. Recombinant heavy meromyosin- and subfragment-1 (S1)-like constructs for both myosin-18A? and -18? species were purified from the baculovirus/Sf9 cell expression system. These constructs bound both essential and regulatory light chains, indicating an additional noncanonical light chain binding site in the neck. Myosin-18A?-S1 and -18A?-S1 molecules bound actin weakly with Kd values of 4.9 and 54 ?m, respectively. The actin binding data could be modeled by assuming an equilibrium between two myosin conformations, a competent and an incompetent form to bind actin. Actin binding was unchanged by presence of nucleotide. Both myosin-18A isoforms bound N-methylanthraniloyl-nucleotides, but the rate of ATP hydrolysis was very slow (<0.002 s(-1)) and not significantly enhanced by actin. Phosphorylation of the regulatory light chain had no effect on ATP hydrolysis, and neither did the addition of tropomyosin or of GOLPH3, a myosin-18A binding partner. Electron microscopy of myosin-18A-S1 showed that the lever is strongly angled with respect to the long axis of the motor domain, suggesting a pre-power stroke conformation regardless of the presence of ATP. These data lead us to conclude that myosin-18A does not operate as a traditional molecular motor in cells.

SUBMITTER: Guzik-Lendrum S

PROVIDER: S-EPMC3611021 | biostudies-literature | 2013 Mar

REPOSITORIES: biostudies-literature

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Mammalian myosin-18A, a highly divergent myosin.

Guzik-Lendrum Stephanie S Heissler Sarah M SM Billington Neil N Takagi Yasuharu Y Yang Yi Y Knight Peter J PJ Homsher Earl E Sellers James R JR

The Journal of biological chemistry 20130204 13

The Mus musculus myosin-18A gene is expressed as two alternatively spliced isoforms, α and β, with reported roles in Golgi localization, in maintenance of cytoskeleton, and as receptors for immunological surfactant proteins. Both myosin-18A isoforms feature a myosin motor domain, a single predicted IQ motif, and a long coiled-coil reminiscent of myosin-2. The myosin-18Aα isoform, additionally, has an N-terminal PDZ domain. Recombinant heavy meromyosin- and subfragment-1 (S1)-like constructs for ...[more]

PMID: 23382379

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Project description:UnlabelledThe hepatitis C virus (HCV; genus Hepacivirus) is a highly relevant human pathogen. Unique hepaciviruses (HV) were discovered recently in animal hosts. The direct ancestor of HCV has not been found, but the genetically most closely related animal HVs exist in horses. To investigate whether other peridomestic animals also carry HVs, we analyzed sera from Ghanaian cattle for HVs by reverse transcription-PCR (RT-PCR). Nine of 106 specimens from different sampling sites contained HV RNA (8.5%) at median viral loads of 1.6 × 10(5) copies/ml. Infection seemed unrelated to cattle age and gender. Near-full-genome sequencing of five representative viruses confirmed taxonomic classifications. Cattle HVs formed two distinct phylogenetic lineages that differed by up to 17.7% on the nucleotide level in the polyprotein-encoding region, suggesting cocirculation of different virus subtypes. A conserved microRNA122-binding site in the 5' internal ribosomal entry site suggested liver tropism of cattle HVs. Phylogenetic analyses suggested the circulation of HVs in cattle for several centuries. Cattle HVs were genetically highly divergent from all other HVs, including HCV. HVs from genetically related equine and bovine hosts were not monophyletic, corroborating host shifts during the evolution of the genus Hepacivirus. Similar to equine HVs, the genetic diversity of cattle HVs was low compared to that of HCV genotypes. This suggests an influence of the human-modified ecology of peridomestic animals on virus diversity. Further studies should investigate the occurrence of cattle HVs in other geographic areas and breeds, virus pathogenicity in cattle, and the potential exposure of human risk groups, such as farmers, butchers, and abattoir workers.ImportanceHCV (genus Hepacivirus) is a major human pathogen, causing liver failure and cancer. Unique hepaciviruses (HVs) were discovered over the last few years in animals, but the direct ancestor of HCV has not been found. The animal HV most closely related to HCV so far originated from horses, suggesting that other livestock animals also harbor HVs. Therefore, we investigated African cattle and discovered previously unknown HVs at high prevalence and viral loads. Because of the agricultural importance of cattle, it may be relevant to investigate HV pathogenicity. The frequent exposure of humans to cattle also may warrant investigations of the zoonotic potential of these viruses. Evolutionary analyses suggested that cattle HVs have existed for centuries. Despite the genetic relatedness of their animal hosts, HVs from cattle and horses were not phylogenetically related, corroborating frequent host shifts during the evolution of the genus Hepacivirus.

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Mammalian myosin-18A, a highly divergent myosin.

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Mammalian myosin-18A, a highly divergent myosin.

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