Herpes simplex virus type 1 HindIII fragment L encodes spliced and complementary mRNA species

Abstract

We have used DNA bound to cellulose to isolate and translate in vitro herpes simplex virus type 1 (HSV-1) mRNA's encoded by HindIII fragment L (mapping between 0.592 and 0.647), an 8,450-base-pair (8.45-kb) portion of the long unique region of the viral genome. Readily detectable, late mRNA's 2.7 and 1.9 kb in size encoding 69,000-and 58,000-dalton polypeptides, respectively, were isolated. A very minor late mRNA family composed of two colinear forms, one 2.6 kb and one 2.8 kb, was isolated and found to encode only an 85,000-dalton polypeptide. A major early mRNA, 1.8 kb in size encoding a 64,000-dalton polypeptide, was also isolated. High-resolution mapping of these mRNA's by using Si nuclease and exonuclease VII digestion of hybrids between them and 5' and 3' end-labeled DNA fragments from the region indicated that the major early mRNA contained no detectable splices, and about half of its 3' end was complementary to the 3' region of the very minor 2.6-to 2.8-kb mRNA's encoded on the opposite strand. These mRNA's also contained no detectable splices. The major late 2.7-kb mRNA was found to be a family made up of members with no detectable splices and members with variable-length (100 to 300 bases) segments spliced out very near (ca. 50 to 100 bases) the 5' end. Like all herpesviruses, herpes simplex virus type 1 (HSV-1) is characterized by an unusual arrangement of its genome. The viral DNA has a molecular weight of 95 x 106 to 100 x 106 daltons (d; reviewed in reference 44), which corresponds to a length of 150,000 base pairs (150 kb; 50). The linear HSV-1 genome is segmented into a long unique region (UL, ca. 105 kb, 70%)

Kinetics of expression of the gene encoding the 65-kilodalton DNA-binding protein of herpes simplex virus type 1

“…This probe detected only a single major mRNA species of approximately 1.8 kilobases in infected cells at all times examined ( Fig. 5), in agreement with the previously reported size of the transcript of this gene (7). The 65KDBP mRNA was first detected at 4 h p.i.…”

“…The studies described in this report clearly demonstrate that the gene encoding 65KDBP is a member of the early or P kinetic class of HSV-1 genes as assessed by a number of parameters. Frink et al (7) initially classified the transcript from this gene as a ,B transcript on the basis of the presence of the transcript when DNA synthesis was inhibited with adenosine arabinoside. The pattern of protein accumulation that we found was consistent with the gene belonging either to the P or ,3y class.…”

“…The localization studies demonstrated that 65KDBP was expressed as early as ICP8, which has been classified as a ,B polypeptide (13). Analysis of transcripts which were synthesized in the presence of adenosine arabinoside also indicated that the 65KDBP transcripts accumulated as expected for a , gene (7). However, to better determine the kinetic class to which 65KDBP belongs, we examined a number of parameters indicative of gene expression, including protein and mRNA accumulation, and compared these in the same infected cells with the patterns of expression of well-characterized model , and y genes.…”

“…In an early report, Frink et al (7) demonstrated that a transcript which mapped to a region on the HSV-1 genome now known to encode the 65KDBP was expressed in the presence of the viral DNA synthesis inhibitor adenosine arabinoside. It was, therefore, classified as an early or ,( transcript.…”

Characterization of the gene encoding herpes simplex virus type 2 glycoprotein C and comparison with the type 1 counterpart

“…Within the sequence which codes for gC-2, both of the other reading frames on that strand are closed, one with 14 termination codons, and the other with 4. Mapping of HSV-1 RNAs from this region has suggested that spliced RNAs may be present (11); however, additional RNAs or translation products have not been clearly demonstrated in this region of HSV-2. The mRNA which specifies the 18K protein uses a different reading frame from that of gC-2.…”