At least three separate regions of the Epstein-Barr virus (EBV) genome encode RNA in a cell line that is growth transformed and nonpermissively infected with EBV. Six polyadenylylated cytoplasmic RNAs have been identified from these three regions. An abundant RNA 3.0-3.1 kilobases (kb) long is encoded by DNA of the internal reiteration, IR, and DNA that maps at 25.7-30 megadaltons. A second, abundant, 2.9-kb RNA is primarily encoded by DNA at 110-03 megadaltons but probably has a 3' end to the left of 110 megadaltons. A third, abundant, 3.7-kb RNA is largely encoded by DNA at 63-66 megadaltons and has a 5' end to the left of 63 megadaltons. A less-abundant 1.5-kb RNA is also encoded by IR. The least-abundant polyadenylylated RNAs identified are 2.3 and 2.0 kb. These RNAs have 3' ends mapping of 5-7 megadaltons and 5' ends mapping to the right of 7 megadaltons. The data suggest that there may be two additional polyadenylylated cytoplasmic RNAs, a 3-kb RNA mapping at 26.2-30 megadaltons and a minor RNA mapping at 102-110 megadaltons. An abundant 0.16-kb nonpolyadenylylated RNA is also present in the cytoplasm of IB-4 cells. This RNA precipitates from the cytoplasm in the presence of high concentrations of magnesium, indicating that it is complexed with protein or polyribosomes.

The most abundant Epstein-Barr virus mRNA in a latently infected cell line, IB4, established by in vitro growth transformation with virus, was a 2.8-kilobase RNA encoded by largely unique DNA near the right end of the genome. The RNA was transcribed from right to left, and two introns were spliced out. This region of the genome was sequenced, and the exons of the RNA were identified by S1 analysis of DNA-RNA hybrids and primer extension. The first start codon in the RNA was 40 nucleotides from its 5' end. Beginning with the start codon, there was a 1,158-nucleotide open reading frame which crossed both introns. The important characteristics of the translated protein were as follows. (i) The amino terminus was highly charged and not suggestive of a leader sequence. (ii) There were six markedly hydrophobic alpha-helical domains, each having 21 amino acids and connected by 5 to 7 amino acid segments predicted to be reverse turns. (iii) The carboxy-terminal 200 amino acids were markedly acidic, containing 6 glutamic and 37 aspartic acids. The hydrophobic region is predicted to form six membrane-spanning regions, leaving the short charged amino terminus and long acidic carboxy terminus on the inside of the membrane. This protein could be responsible for the new antigen detected in the plasma membrane of Epstein-Barr virus-transformed cells, lymphocyte-determined membrane antigen. There were two other open reading frames in the RNA.

The size of the Epstein-Barr virus (EBV) nuclear antigen (EBNA) in cells infected with different EBV isolates varies directly with the size of the EBV triplet repeat array, IR3. The isolate with the largest IR3 fragment has approximately 170 more codons than the isolates with the smallest IR3 fragment; it encodes an EBNA which is approximately 17,000 daltons larger than the smallest EBNA. The EBV IR3 encodes part of a 2-kilobase exon of a latently infected cell messenger RNA which must be translated into a repetitive amino acid domain of EBNA.

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We have studied the interaction of two of the Ul small nuclear ribonucleoprotein (snRNP)-specific proteins, Ul-70K and U1-A, with Ul small nuclear RNA (snRNA). The U1-70K protein is a Ul-specific RNA-binding protein. Deletion and mutation analyses of a ,-galactosidase/Ul-70K partial fusion protein indicated that the central portion of the protein, including the RNP sequence domain, is both necessary and sufficient for specific Ul snRNA binding in vitro. The highly conserved eight-amino-acid RNP consensus sequence was found to be essential for binding. Deletion and mutation analyses of Ul snRNA showed that both the Ul-70K fusion protein and the native HeLa U1-70K protein bound directly to loop I of Ul snRNA. Binding was sequence specific, requiring 8 of the 10 bases in the loop. The U1-A snRNP protein also interacted specifically with Ul snRNA, principally with stem-loop II.The splicing of eucaryotic mRNA precursors (pre-mRNAs) involves a group of small nuclear ribonucleoprotein particles (U snRNPs) that are subassemblies of a larger ribonucleoprotein complex, the spliceosome. At least four different U snRNPs are required for splicing. Ul snRNP mediates recognition of 5' splice sites, and U2, U4/6, and U5 snRNPs mediate recognition of the 3' ends of intervening sequences (reviewed in references 10 and 27). An understanding of the mechanisms by which the various U snRNPs interact with pre-mRNAs, with each other, and with other components of the spliceosome will ultimately require an analysis of the individual snRNP components and their functions.Human Ul snRNP consists of the 164-nucleotide Ul small nuclear RNA (snRNA), a group of at least seven core (Sm) proteins common to all U snRNPs, and three specific proteins: U1-70K, U1-A, and U1-C (5). In the absence of an snRNA, the Sm core proteins associate to form a 6S particle (9). It seems likely that the core snRNP proteins serve a scaffold function, organizing snRNP architecture, and that the specific proteins mediate the functions of Ul snRNP in pre-mRNA splicing.The recent isolation of cDNAs encoding all three of the human Ul-specific snRNP proteins (24-26, 29, 37) has provided considerable structural information on these polypeptides. The U1-70K and U1-A proteins both contain sequence motifs that occur in several known nucleic acidbinding proteins, including the RNP sequence domain (1, 28; reviewed in reference 7) which is apparently involved in RNA binding (6,19). The RNP sequence domain contains within it a short, even more highly conserved segment, the RNP consensus sequence (rY'GlyPheGlyPhelleXPh¾e; reviewed in reference 7). In addition, the U1-70K protein contains two arginine-rich tracts reminiscent of those in protamines, sperm histones, and chicken galline, proteins that bind DNA * Corresponding author. t Paper 3044 from the