Abstract
We have identified and characterized the transcript for the human cytomegalovirus (HCMV) UL102 gene. The UL102 gene product is proposed to encode the primase-associated factor. The primase-associated factor is one of the three components of the helicase-primase complex, along with UL105 (helicase) and UL70 (primase). In order to characterize the UL102 transcription unit we used single-stranded antisense RNA probes to identify an abundant 2.7-kb transcript originating from the UL102 region. This transcript can be initially detected at 24 h postinfection and in the presence of phosphonoformic acid but not in the presence of cycloheximide. A 2.7-kb cDNA clone containing this transcript was isolated from a 72-h HCMV (strain Towne) cDNA library. Sequence analysis of this clone revealed a continuous unspliced transcript between the region of UL101X and UL102; the only in-frame translational stop codon is 2,619 bp downstream from the first ATG in the message. Genome sequencing of the UL102 region from strains AD169 and Towne revealed that the UL101X stop codon TAA was actually TAC and that the cDNA and genomic sequences were in agreement. The cDNA clone starts 5 nucleotides (nt) upstream of the UL101X ATG, continues through the putative ATG of UL102, and ends 97 nt downstream of the putative termination codon of the UL102 open reading frame. Primer extension analysis indicated a transcriptional start site 23 nt upstream of the UL101X open reading frame.
Full Text
The Full Text of this article is available as a PDF (360.0 KB).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Calder J. M., Stow N. D. Herpes simplex virus helicase-primase: the UL8 protein is not required for DNA-dependent ATPase and DNA helicase activities. Nucleic Acids Res. 1990 Jun 25;18(12):3573–3578. doi: 10.1093/nar/18.12.3573. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Chee M. S., Bankier A. T., Beck S., Bohni R., Brown C. M., Cerny R., Horsnell T., Hutchison C. A., 3rd, Kouzarides T., Martignetti J. A. Analysis of the protein-coding content of the sequence of human cytomegalovirus strain AD169. Curr Top Microbiol Immunol. 1990;154:125–169. doi: 10.1007/978-3-642-74980-3_6. [DOI] [PubMed] [Google Scholar]
- Colberg-Poley A. M., Santomenna L. D., Harlow P. P., Benfield P. A., Tenney D. J. Human cytomegalovirus US3 and UL36-38 immediate-early proteins regulate gene expression. J Virol. 1992 Jan;66(1):95–105. doi: 10.1128/jvi.66.1.95-105.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Crute J. J., Lehman I. R. Herpes simplex virus-1 helicase-primase. Physical and catalytic properties. J Biol Chem. 1991 Mar 5;266(7):4484–4488. [PubMed] [Google Scholar]
- Crute J. J., Tsurumi T., Zhu L. A., Weller S. K., Olivo P. D., Challberg M. D., Mocarski E. S., Lehman I. R. Herpes simplex virus 1 helicase-primase: a complex of three herpes-encoded gene products. Proc Natl Acad Sci U S A. 1989 Apr;86(7):2186–2189. doi: 10.1073/pnas.86.7.2186. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Dodson M. S., Crute J. J., Bruckner R. C., Lehman I. R. Overexpression and assembly of the herpes simplex virus type 1 helicase-primase in insect cells. J Biol Chem. 1989 Dec 15;264(35):20835–20838. [PubMed] [Google Scholar]
- Dodson M. S., Lehman I. R. Association of DNA helicase and primase activities with a subassembly of the herpes simplex virus 1 helicase-primase composed of the UL5 and UL52 gene products. Proc Natl Acad Sci U S A. 1991 Feb 15;88(4):1105–1109. doi: 10.1073/pnas.88.4.1105. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Ertl P. F., Powell K. L. Physical and functional interaction of human cytomegalovirus DNA polymerase and its accessory protein (ICP36) expressed in insect cells. J Virol. 1992 Jul;66(7):4126–4133. doi: 10.1128/jvi.66.7.4126-4133.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Ertl P. F., Thomas M. S., Powell K. L. High level expression of DNA polymerases from herpesviruses. J Gen Virol. 1991 Jul;72(Pt 7):1729–1734. doi: 10.1099/0022-1317-72-7-1729. [DOI] [PubMed] [Google Scholar]
- Fixman E. D., Hayward G. S., Hayward S. D. trans-acting requirements for replication of Epstein-Barr virus ori-Lyt. J Virol. 1992 Aug;66(8):5030–5039. doi: 10.1128/jvi.66.8.5030-5039.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
- He Y. S., Xu L., Huang E. S. Characterization of human cytomegalovirus UL84 early gene and identification of its putative protein product. J Virol. 1992 Feb;66(2):1098–1108. doi: 10.1128/jvi.66.2.1098-1108.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Heilbronn R., Jahn G., Bürkle A., Freese U. K., Fleckenstein B., zur Hausen H. Genomic localization, sequence analysis, and transcription of the putative human cytomegalovirus DNA polymerase gene. J Virol. 1987 Jan;61(1):119–124. doi: 10.1128/jvi.61.1.119-124.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Jenkins D. E., Martens C. L., Mocarski E. S. Human cytomegalovirus late protein encoded by ie2: a trans-activator as well as a repressor of gene expression. J Gen Virol. 1994 Sep;75(Pt 9):2337–2348. doi: 10.1099/0022-1317-75-9-2337. [DOI] [PubMed] [Google Scholar]
- Kemble G. W., McCormick A. L., Pereira L., Mocarski E. S. A cytomegalovirus protein with properties of herpes simplex virus ICP8: partial purification of the polypeptide and map position of the gene. J Virol. 1987 Oct;61(10):3143–3151. doi: 10.1128/jvi.61.10.3143-3151.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kerry J. A., Priddy M. A., Stenberg R. M. Identification of sequence elements in the human cytomegalovirus DNA polymerase gene promoter required for activation by viral gene products. J Virol. 1994 Jul;68(7):4167–4176. doi: 10.1128/jvi.68.7.4167-4176.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Klinedinst D. K., Challberg M. D. Helicase-primase complex of herpes simplex virus type 1: a mutation in the UL52 subunit abolishes primase activity. J Virol. 1994 Jun;68(6):3693–3701. doi: 10.1128/jvi.68.6.3693-3701.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kouzarides T., Bankier A. T., Satchwell S. C., Weston K., Tomlinson P., Barrell B. G. Sequence and transcription analysis of the human cytomegalovirus DNA polymerase gene. J Virol. 1987 Jan;61(1):125–133. doi: 10.1128/jvi.61.1.125-133.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Malone C. L., Vesole D. H., Stinski M. F. Transactivation of a human cytomegalovirus early promoter by gene products from the immediate-early gene IE2 and augmentation by IE1: mutational analysis of the viral proteins. J Virol. 1990 Apr;64(4):1498–1506. doi: 10.1128/jvi.64.4.1498-1506.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Martignetti J. A., Barrell B. G. Sequence of the HindIII T fragment of human cytomegalovirus, which encodes a DNA helicase. J Gen Virol. 1991 May;72(Pt 5):1113–1121. doi: 10.1099/0022-1317-72-5-1113. [DOI] [PubMed] [Google Scholar]
- McGeoch D. J., Dalrymple M. A., Davison A. J., Dolan A., Frame M. C., McNab D., Perry L. J., Scott J. E., Taylor P. The complete DNA sequence of the long unique region in the genome of herpes simplex virus type 1. J Gen Virol. 1988 Jul;69(Pt 7):1531–1574. doi: 10.1099/0022-1317-69-7-1531. [DOI] [PubMed] [Google Scholar]
- McGeoch D. J., Dalrymple M. A., Dolan A., McNab D., Perry L. J., Taylor P., Challberg M. D. Structures of herpes simplex virus type 1 genes required for replication of virus DNA. J Virol. 1988 Feb;62(2):444–453. doi: 10.1128/jvi.62.2.444-453.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Olivo P. D., Nelson N. J., Challberg M. D. Herpes simplex virus type 1 gene products required for DNA replication: identification and overexpression. J Virol. 1989 Jan;63(1):196–204. doi: 10.1128/jvi.63.1.196-204.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Pari G. S., Anders D. G. Eleven loci encoding trans-acting factors are required for transient complementation of human cytomegalovirus oriLyt-dependent DNA replication. J Virol. 1993 Dec;67(12):6979–6988. doi: 10.1128/jvi.67.12.6979-6988.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Pari G. S., Kacica M. A., Anders D. G. Open reading frames UL44, IRS1/TRS1, and UL36-38 are required for transient complementation of human cytomegalovirus oriLyt-dependent DNA synthesis. J Virol. 1993 May;67(5):2575–2582. doi: 10.1128/jvi.67.5.2575-2582.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Parry M. E., Stow N. D., Marsden H. S. Purification and properties of the herpes simplex virus type 1 UL8 protein. J Gen Virol. 1993 Apr;74(Pt 4):607–612. doi: 10.1099/0022-1317-74-4-607. [DOI] [PubMed] [Google Scholar]
- Peppel K., Baglioni C. A simple and fast method to extract RNA from tissue culture cells. Biotechniques. 1990 Dec;9(6):711–713. [PubMed] [Google Scholar]
- Sherman G., Gottlieb J., Challberg M. D. The UL8 subunit of the herpes simplex virus helicase-primase complex is required for efficient primer utilization. J Virol. 1992 Aug;66(8):4884–4892. doi: 10.1128/jvi.66.8.4884-4892.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Takebe Y., Seiki M., Fujisawa J., Hoy P., Yokota K., Arai K., Yoshida M., Arai N. SR alpha promoter: an efficient and versatile mammalian cDNA expression system composed of the simian virus 40 early promoter and the R-U5 segment of human T-cell leukemia virus type 1 long terminal repeat. Mol Cell Biol. 1988 Jan;8(1):466–472. doi: 10.1128/mcb.8.1.466. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Tenney D. J., Hurlburt W. W., Micheletti P. A., Bifano M., Hamatake R. K. The UL8 component of the herpes simplex virus helicase-primase complex stimulates primer synthesis by a subassembly of the UL5 and UL52 components. J Biol Chem. 1994 Feb 18;269(7):5030–5035. [PubMed] [Google Scholar]
- Wright D. A., Spector D. H. Posttranscriptional regulation of a class of human cytomegalovirus phosphoproteins encoded by an early transcription unit. J Virol. 1989 Jul;63(7):3117–3127. doi: 10.1128/jvi.63.7.3117-3127.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Wu C. A., Nelson N. J., McGeoch D. J., Challberg M. D. Identification of herpes simplex virus type 1 genes required for origin-dependent DNA synthesis. J Virol. 1988 Feb;62(2):435–443. doi: 10.1128/jvi.62.2.435-443.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]