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Journal of Virology, August 2004, p. 8191-8200, Vol. 78, No. 15
0022-538X/04/$08.00+0     DOI: 10.1128/JVI.78.15.8191-8200.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Analysis of Splice Variants of the Immediate-Early 1 Region of Human Cytomegalovirus

Sita Awasthi,{dagger} Jennifer A. Isler,{dagger} and James C. Alwine*

Department of Cancer Biology, Abramson Family Cancer Research Institute, School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104-6142

Received 5 February 2004/ Accepted 2 April 2004

The major immediate-early (MIE) gene of human cytomegalovirus (HCMV) produces multiple mRNAs through differential splicing and polyadenylation. Reverse transcriptase PCR was used to characterize transcripts from exons 1, 2, 3, and 4 (immediate-early 1 [IE1]). The expected IE72 and IE19 mRNAs were detected, as well as two heretofore-uncharacterized transcripts designated IE17.5 and IE9. The IE72, IE19, and IE17.5 transcripts utilized the same 5'-splice site in exon 3. IE9 utilized a cryptic 5'-splice site within exon 3. The IE19, IE17.5, and IE9 transcripts all used different 3'-splice sites within exon 4. These spliced species occur in infected human foreskin fibroblast (HFF) cells, with accumulation kinetics similar to those of IE72 mRNA. IE19 and IE9 RNAs were much more abundant than IE17.5 RNA. Transfection of CV-1 cells with cDNAs resulted in IE19 and IE17.5 proteins detectable by antibodies to either N-terminal or C-terminal epitopes. No IE9 protein product has been detected. We have not been able to detect IE19, IE17.5, or IE9 proteins during infection of HFF, HEL, or U373MG cells. Failure to detect IE19 protein contrasts with a previous report (M. Shirakata, M. Terauchi, M. Ablikin, K. Imadome, K. Hirai, T. Aso, and Y. Yamanashi, J. Virol. 76:3158-3167, 2002) of IE19 protein expression in HCMV-infected HEL cells. Our analysis suggests that an N-terminal breakdown product of IE72 may be mistaken for IE19. Expression of IE19 or IE17.5 from its respective cDNA results in repression of viral gene expression in infected cells. We speculate that expression of these proteins during infection may be restricted to specific conditions or cell types.

* Corresponding author. Mailing address: 314 Biomedical Research Building II/III, 421 Curie Blvd., University of Pennsylvania, Philadelphia, PA 19104. Phone: (215) 898-3256. Fax: (215) 573-3888. E-mail: alwine{at}mail.med.upenn.edu.

{dagger} S.A. and J.A.I. contributed equally.

Journal of Virology, August 2004, p. 8191-8200, Vol. 78, No. 15
0022-538X/04/$08.00+0     DOI: 10.1128/JVI.78.15.8191-8200.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



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