A mutant CHO-K1 strain with resistance to Pseudomonas exotoxin A and alphaviruses fails to cleave Sindbis virus glycoprotein PE2.
RPE.40, a mutant CHO-K1 strain selected for resistance to Pseudomonas exotoxin A, is defective in the production of infectious alphaviruses, although viruses are taken in and processed normally (J. M. Moehring and T. J. Moehring, Infect. Immun. 41:998-1009, 1983). To determine the cause of this defect, the synthesis of Sindbis virus proteins was examined. RPE.40 cells produced and glycosylated structural glycoprotein precursors PE2 and immature E1 normally. Mature E1 was formed, but PE2 was not cleaved to E2 and E3. PE2 instead was modified to a higher-molecular-weight form (PE2') in which the high-mannose oligosaccharides were processed to the complex form without proteolytic cleavage. The data suggest that the cleavage which produces E2 occurs within the trans-Golgi or in post-Golgi elements and is closely associated with the addition of sialic acid residues to the asparagine-linked oligosaccharides. RPE.40 cells make and release noninfectious Sindbis virions that contain PE2' and no detectable E2. These virions can be converted to an infectious form by treatment with trypsin. A defect in an intracellular endopeptidase activity in RPE.40 cells is postulated. Comparison of two Sindbis virus strains showed that the requirement for E2 in the virion to ensure infectivity is strain specific.
Watson
D G
DG
Department of Microbiology and Molecular Genetics, College of Medicine, University of Vermont, Burlington 05405.
Moehring
J M
JM
Moehring
T J
TJ
D036002:ADP Ribose Transferases; D000524:Alphavirus / Q000502:physiology*; D000818:Animals; D001427:Bacterial Toxins / Q000494:pharmacology*; D015153:Blotting, Western; D002460:Cell Line / Q000378:metabolism / Q000382:microbiology*; D006224:Cricetinae; D004351:Drug Resistance / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D005098:Exotoxins / Q000494:pharmacology*; D005260:Female; D009154:Mutation; D010053:Ovary; D011498:Protein Precursors / Q000378:metabolism; D012845:Sindbis Virus / Q000378:metabolism*; D014357:Trypsin / Q000378:metabolism; D014759:Viral Envelope Proteins / Q000378:metabolism*; D037521:Virulence Factors; D000097668:Pseudomonas aeruginosa Exotoxin A
D036002:ADP Ribose Transferases; D001427:Bacterial Toxins; D005098:Exotoxins; D011498:Protein Precursors; D014357:Trypsin; D014759:Viral Envelope Proteins; D037521:Virulence Factors; D000097668:Pseudomonas aeruginosa Exotoxin A; C064950:glycoprotein E2, Sindbis virus