Organization and expression of the immediate early genes of human cytomegalovirus

Abstract

The immediate early genes of human cytomegalovirus were characterized according to map location, RNA transcripts, and translation products. Three regions in the large unique component (0.709 to 0.751 map units) were transcribed in the presence of an inhibitor of protein synthesis (anisomycin). A single size class of polyadenylated mRNA, 1.95 kilobases (kb), was transcribed abundantly relative to the other size classes. The predominant 1.95-kb viral RNA was transcribed from right to left on the prototype arrangement of the viral genome and spanned a region of approximately 2.8 kb (0.739 to 0.751 map units). This mRNA codes for a 75,000-dalton protein that represents the predominant immediate early protein detected in infected cells. Immunoprecipitation of viral proteins synthesized in vitro as well as in vivo demonstrated that the predominant immediate early protein is synthesized as a protein of 75,000 daltons, but is presumably modified in vivo, resulting in a broad banding pattern ranging from 75,000 to 68,000 daltons. A different immediate early viral gene (0.732 to 0.739 map units) is transcribed from left to right at relatively low levels. The 3' ends of the above viral RNAs terminate at approximately 230 base pairs apart in the region of approximately 0.739 map units. Five RNA size classes ranging from 2.25 to 1.10 kb were detected, but the 1.75-kb and 1.40-kb RNA size classes were more abundant from this region. At least four minor proteins are coded by these mRNAs, with apparent molecular weights ranging from 56,000 to 16,500. Last, a 1.95-kb mRNA was transcribed from a third region (0.709 to 0.728 map units). This viral mRNA was present at relatively low concentration and coded for a minor protein of 68,000 daltons. Since immediate early gene expression of human cytomegalovirus is dominated by the synthesis of an mRNA from the region of 0.739 to 0.751 map units that codes for the predominant immediate early protein found in the infected cell, we hypothesize that this protein is the major regulatory protein influencing the switch from restricted to extensive transcription.

Expression of Human Cytomegalovirus Immediate Early Antigens Is Responsible for a Novel Chromatin Structure of Infected Human Embryonic Lung Cells

“…The transformant, NIH/3T3-Ea4, expressed three proteins apparently corresponding to those of 78, 70, and 68kD (lane 3), and no such proteins were detected in the control NIH/3T3 cells transformed with pSV2neo (lane 4). These values for virus-specific proteins were within the range so far reported for the major IEA's of HCMV (11,19,24), and these three size-class proteins probably constitute the IEA's detected by ACIF. NIH/3T3-Ea4 cells expressing HCMV IEA's were found to display chromatin fibers composed of conventional nucleosomes and no ellipsoid beads (not shown).…”

“…In the present study, we have confirmed their finding and The cloned HindIII C fragment of HCMV (Towne) DNA (6) was digested with EcoRI, and the EcoRI H fragment carrying the major immediate early (MIE) and immediate early 2 (IE2) genes (15)(16)(17)19) was subsequently cloned by being ligated into the EcoRI site of pSV2neo (13). This plasmid, designated pSV2neoEcoH ( Fig.…”

Physical mapping of human cytomegalovirus genes: Identification of DNA sequences coding for a virion phosphoprotein of 71 kDa and a viral 65-kDa polypeptide

“…The HCMV genome has been cloned in plasmid (Thomsen and Stinski, 1981;Oram et al, 1982;Tai aL, 1982) and cosmid vectors (Fleckenstein et al, 1982), providing a basis for the physical and functional mapping of viral DNA. A small, contiguous region of the viral genome is abundantly transcribed into three or four species of RNA during the immediate early phase of viral replication (DeMarchi, 1981;Wathen and Stinski, 1982;McDonough and Spector, 1983;Jahn et aL, submitted), and the coding region for one immediate early gene product of 72 kDa has been identified (Stinski et aL, 1983). On the other hand, transcripts from most regions of the viral genome are found in infected cells during subsequent phases of viral replication (DeMarchi, 1981;Wathen and Stinski, 1982;McDonough and Spector, 1983), but no correlation has been reported between a defined virion protein and its respective coding sequence.…”

Interactions between cytomegalovirus immediate‐early proteins and the long terminal repeat of human immunodeficiency virus

“…The four IE genes are designated UL36-3814'Gz2, TRSI-IRSI (a US22 family memberz6) and the major IE (MIE) gene (UL122/123) 13,16,17,23 based on the open reading frame classification from the genome sequencing project. '* The IE genes have the capacity to encode multiple proteins as a result of alternative promoter and polyadenylation/cleavage site usage as well as employing differential splicing configurations.…”