Characterization of Herpes Simplex Virus Type 1 RNA Present in the Absence of De Novo Protein Synthesis

Abstract

We used herpes simplex virus type 1 (HSV-1) DNA and restriction fragments of HSV-1 DNA covalently coupled to cellulose as a reagent to isolate for further characterization the major and minor HSV-1 immediate-early mRNA species in HeLa cells infected and maintained in the absence of de novo protein synthesis. Five major and several minor immediate-early mRNA species were characterized. One major species was a 4.2-kilobase mRNA mapping in the TR S /IR S region with its 3′ end distal to the U S region; this mRNA encoded a 170,000-dalton polypeptide in vitro. A 2.8-kilobase mRNA, encoding a 120,000-dalton polypeptide, was mapped in the TR L /IR L region with its 3′ end directed toward the U L region. Three 1.8-kilobase mRNA species were mapped. One, mapping in the IR S region with its 3′ end in the U S , encoded a 68,000-dalton polypeptide. One mapped in the TR S region and had its 3′ end in the U S region; the third one encoded a 64,000-dalton polypeptide and mapped in the U L region near the IR L region. One minor species 5.2 kilobases in size was clearly detectable mapping in the U L region. Furthermore, there were indications that one or more immediate-early mRNA species approximately 3 kilobases in size hybridized to regions near the TR L and in or near the TR S /IR S regions. Nuclear immediate-early RNA mapped only in those regions where polyribosomal immediate-early mRNA mapped, although minor differences were seen. Finally, we demonstrated that at least three major immediate-early mRNA's—4.2 kilobases, 2.8 kilobases, and the 1.8-kilobase one mapping in the IR S /U S region—continued to appear on polyribosomes as functional mRNA late after infection.