Suppression of host mRNA in human smooth muscle cells by a virion competent factor in herpes simplex virus type 1.
Herpes simplex virus type 1 produces an early decrease in protein synthesis in cultured smooth muscle cells isolated from human umbilical artery. To confirm that suppression of host protein synthesis occurs before virus protein expression, cultures were treated with actinomycin D (5 micrograms/ml) continuously from the beginning of infection. In the presence of actinomycin D, by 3.5 hours postinfection, virus-infected cultures showed much less incorporation of [35S]methionine into sodium dodecyl sulfate-polyacrylamide electrophoresis-separated products than did identically treated mock-infected cultures. In the absence of actinomycin D, there was incorporation of radiolabel into viral proteins as early as 4.5 hours postinfection. Hybridization of slot-blotted total RNA demonstrated that virus-infected cultures treated with and without actinomycin D showed a marked decrease in hybridizable RNA by 2 hours PI when compared with mock-infected controls. This decrease was seen with cDNA probes for the mRNA for secreted extracellular matrix macromolecules (fibronectin, thrombospondin and collagen chains alpha 2(I), alpha 1(III] as well as with cDNA probes for RNA of intracellular macromolecules actin and beta-tubulin. In vitro translation of total RNA isolated from infected human smooth muscle cells at 2 and 5 hours postinfection resulted in far less [35S]methionine incorporation into sodium dodecyl sulfate-polyacrylamide electrophoresis-separated products than would be predicted from the amount of hybridizable RNA available. The results suggest that host message is rapidly rendered nonfunctional as well as degraded by means of a virion-competent mechanism(s).
London
F S
FS
Department of Medicine, University of Pennsylvania, Philadelphia.
Brinker
J M
JM
Ziaie
Z
Z
Kefalides
N A
NA
D003609:Dactinomycin / Q000378:metabolism; D006801:Humans; D006824:Hybridization, Genetic; D009124:Muscle Proteins / Q000378:metabolism*; D009131:Muscle, Smooth, Vascular / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis* / Q000302:isolation & purification; D018139:Simplexvirus / Q000235:genetics* / Q000378:metabolism; D013997:Time Factors; D014158:Transcription, Genetic; D014469:Umbilical Arteries / Q000378:metabolism; D014764:Viral Proteins / Q000378:metabolism*; D014771:Virion / Q000378:metabolism*
D009124:Muscle Proteins; D012333:RNA, Messenger; D014764:Viral Proteins; D003609:Dactinomycin