Membrane association facilitates the correct processing of pp220 during production of the major matrix proteins of African swine fever virus.
The African swine fever (ASF) virus polyprotein pp220 is processed at Gly-Gly-X sites by a virally encoded SUMO-like protease to produce matrix proteins p150, p37, p34, and p14. Four Gly-Gly-X sites are used to produce the matrix proteins, but the polyprotein contains an additional 15 sites potentially recognized by the protease. This study shows that cleavage occurs at many, if not all, Gly-Gly-X sites, and at steady state, p150 and p34 are minor products of processing. Significantly, only the final structural proteins, p150 and p34, were found in mature virions, suggesting that there is a mechanism for excluding incorrectly processed forms. ASF virus is assembled on the cytoplasmic face of the endoplasmic reticulum, and the distribution of pp220 products between membranes and cytosol was studied. Incorrectly processed forms of p34 were recovered from both the cytosol and membrane fractions. Interestingly, p34 was only detected in the membrane fraction, and of the many processed forms bound to membranes, only p34 was protected from trypsin, suggesting envelopment. The majority of the incorrectly processed forms of p150 were recovered from the cytosol. Again, the correct product of processing, p150, was selectively recruited to membranes. Sucrose density centrifugation showed that membrane-associated forms of p34 and p150 assembled into large structures suggestive of a viral matrix, while cytosolic and/or incorrectly processed forms of pp220 did not. Taken together, these results suggest that association with cellular membranes is important for regulating the correct processing of pp220 and the packaging of matrix proteins into virions.
Heath
Colin M
CM
Institute for Animal Health, Pirbright Laboratories, Woking, Surrey GU24 0NF, United Kingdom.
Windsor
Miriam
M
Wileman
Thomas
T
D000358:African Swine Fever Virus / Q000502:physiology*; D000595:Amino Acid Sequence; D000818:Animals; D002462:Cell Membrane / Q000378:metabolism*; D002522:Chlorocebus aethiops; D008969:Molecular Sequence Data; D008970:Molecular Weight; D020815:Polyproteins / Q000378:metabolism*; D011498:Protein Precursors / Q000378:metabolism*; D014709:Vero Cells; D014763:Viral Matrix Proteins / Q000378:metabolism*; D014764:Viral Proteins / Q000378:metabolism*; D019065:Virus Assembly
D020815:Polyproteins; D011498:Protein Precursors; D014763:Viral Matrix Proteins; D014764:Viral Proteins