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.
Mohandas
D V
DV
Department of Microbiology and Immunology, University of Western Ontario, London, Canada.
Dales
S
S
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