thanks John Sinclair and Patrick Sissons for their active support of his work at Cambridge and Mat Bentham for his technical assistance

thanks John Sinclair and Patrick Sissons for their active support of his work at Cambridge and Mat Bentham for his technical assistance. REFERENCES 1. Towne infection, but many fewer CR208-infected cells contained the ppUL44 polymerase accessory protein when evaluated at 24 or 48 h after infection. Furthermore, fibroblasts infected with CR208 at a low multiplicity failed to form viral DNA replication compartments, despite having expressed IE2 p86. These low-multiplicity growth and expression defects were corrected in two rescued derivatives of CR208 able to synthesize IE1 p72. One rescued virus (CR249) carried a deletion removing the large intron between exons 1 and 2 of the locus, revealing that this intron was dispensable for growth in cell culture. Human cytomegalovirus (HCMV) is a widespread herpesvirus, which infects 50 to 100% of the human population. HCMV disease is a significant medical problem, although it is mostly restricted EVP-6124 hydrochloride to patients with immature or compromised immune systems (5). HCMV gene expression during productive infection of cultured human fibroblast cells follows an ordered cascade of expression of immediate-early (IE or ) genes, followed by expression of delayed-early (DE or ) genes, followed after viral DNA replication by strong expression of late (L or ) genes. HCMV also modulates the expression of many cellular genes during the virus life cycle (47). IE1 p72 (IE1491aa) is the most abundant product of the strongly transcribed major IE locus of HCMV and is detected in the nuclei of infected cells both in culture and in infected individuals. RNA transcripts EVP-6124 hydrochloride originating from the major IE enhancer-promoter (MIEP) immediately after infection span five major exons and are alternately spliced and polyadenylated to produce messages for either IE1 p72 (exons 1 to 4) or IE2 p86 (IE2579aa) (exons 1 to 3 and 5) (74, 75). The large exon unique to the IE2 p86 message, originally termed exon 7 (75), is here termed exon 5 (47). Translation of IE1 p72 and IE2 p86 initiates in exon 2, and the proteins share 85 identical residues at their amino termini. IE2 p86 is thought to be the major specific transcriptional regulator of the lytic cycle of HCMV. Consistent with a such a role, IE2 p86 is a sequence-specific DNA binding protein, which autoregulates by binding adjacent to the transcription start site of the MIEP (12, 32, 40, 41, 44, 53, 83) and also binds to specific sites in other HCMV promoters (3, 66, 67). IE2 p86 interacts with diverse components of the cellular transcription machinery, including TBP, TFIIB, CREB, CBP, and c-Jun (7, 23, 33, 39, 65, 67), and in transient-cotransfection assays IE2 activates transcription from a wide range of HCMV and cellular promoters (15, 28, 35, 45, 54, 70, 73). Transactivation by IE2 p86 may be mediated by upstream promoter elements (39, 65), but promiscuous activation of heterologous promoters is frequently TATA box mediated (23). In contrast, IE1 p72 acts via discrete promoter elements to stimulate a relatively limited number of viral and cellular promoters. Notably, IE1 p72 transactivates its own promoter, the HCMV MIEP (13, 45, 73), acting via NF-B sites in the 18-bp repeat sequences of the enhancer (13, 62). IE1 p72 also activates the human immunodeficiency virus type 1 long terminal repeat and the cellular DNA polymerase , dihydrofolate reductase, and prointerleukin-1 gene promoters (26, 29, 80, 82). IE1 p72 physically interacts with the cellular transcription factors SP-1, E2F-1, and CTF-1 (26, 43, 46), but binding of IE1 to the basal transcription factors TFIIB and TFIID has not been detected (7, 23). As distinct from its promoter-specific transactivator functions, coexpression of IE1 p72 augments the promiscuous transactivating activities of IE2 p86 on a wide variety of viral and cellular promoters (9, 35, 43, 45, 73). However, the accessory viral transactivators TRS1/IRS1, Mouse monoclonal to COX4I1 UL36-UL38, and UL112-UL113 are in addition required for the maximal EVP-6124 hydrochloride stimulation of a number of HCMV DE promoters (30, 71). Several other distinct activities have been described for IE1 p72, but considered together, these do not form a.