Endosomal association of a protein phosphatase with high dephosphorylating activity against a coronavirus nucleocapsid protein.
On the assumption that dephosphorylation of the neurotropic coronavirus JHM (JHMV) nucleocapsid protein (N) may be connected with initiation of the infectious cycle we searched for a relevant host enzyme activity. Analysis of subcellular fractions from L-2 murine fibroblasts, separated by dual Percoll density gradients, revealed the presence of a phosphoprotein phosphatase (PPPase), co-sedimenting with the endososomal/prelysosomal material, which possesses high activity against N. With purified [32P]N as substrate it was demonstrated that this PPPase, distinguishable from acid and alkaline phosphatases, acts optimally at neutral pH in the presence of Mn2+ following treatment with a detergent. Complete inhibition with okadaic acid at 0.9-4.5 microM but not at 1-10 nM relegates this PPPase to a type 1 protein phosphatase. Similar PPPase activity for N was present in the endosome fraction of a rat Roc-1 astrocytoma-oligodendrocyte cell line and in homogenates of brain and cultured oligodendrocytes. Our data suggest that the phosphorylated N of the inoculum may be modified by the endosomal PPPase in host cells, including those from the CNS so as to facilitate the JHMV infectious process.
Department of Microbiology and Immunology, University of Western Ontario, London, Canada.
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Mohandas
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Dales
D000135:Acid Phosphatase / Q000378:metabolism; D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D001921:Brain / Q000201:enzymology; D002213:Capsid / Q000378:metabolism*; D002413:Cations, Divalent / Q000494:pharmacology; D002451:Cell Compartmentation; D011992:Endosomes / Q000201:enzymology*; D004988:Ethers, Cyclic / Q000494:pharmacology; D066298:In Vitro Techniques; D051379:Mice; D006517:Murine hepatitis virus / Q000378:metabolism*; D009474:Neurons / Q000201:enzymology; D019319:Okadaic Acid; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D012969:Sodium Fluoride / Q000494:pharmacology; D013347:Subcellular Fractions / Q000201:enzymology; D014758:Viral Core Proteins / Q000378:metabolism*
D002413:Cations, Divalent; D004988:Ethers, Cyclic; D014758:Viral Core Proteins; D019319:Okadaic Acid; D012969:Sodium Fluoride; D000469:Alkaline Phosphatase; D010749:Phosphoprotein Phosphatases; D000135:Acid Phosphatase