Nucleotide sequences of cDNAs for human papillomavirus type 18 transcripts in HeLa cells

Papillomaviruses are small double-stranded DNA viruses that replicate episomally in the nuclei of infected cells. The full-length E1 protein of papillomaviruses is required for the replication of viral DNA. The viral mRNA from which the human papillomavirus type 18 E1 protein is expressed is not known. We demonstrate that in eukaryotic cells, the E1 protein is expressed from polycistronic mRNA containing E6, E7, and E1 open reading frames (ORFs). The translation of adjacent E7 and E1 ORFs is not associated; it is performed by separate populations of ribosomes. The translation of the downstream E1 gene is preceded by ribosome scanning. Scanning happens at least at the 5′ end of the polycistronic mRNA and also approximately 100 bp in front of the E1 gene. Long areas in middle of the mRNA are bypassed by ribosomes, possibly by ribosomal “shunting.” Inactivation of short minicistrons in the upstream area of the E1 gene did not change the expression level of the E1 gene.

Section: Discussionmentioning

confidence: 96%

Human Papillomavirus Type 18 E1 Protein Is Translated from Polycistronic mRNA by a Discontinuous Scanning Mechanism

Remm

Ustav

1999

“…The trans-activation function was localized to the 30 C-terminal amino acid residues, E7 dm67-98, which contains only one of the two Cys-X-X-Cys motifs. This amino acid sequence is relatively well conserved among the human papillomaviruses (6,16,23,40,43,57). Analysis of E7 plasmid mutants indicates that some mutations in the E7 domain 2 can lead to defects in trans activation (11,53).…”

Section: Discussionmentioning

confidence: 99%

Chemical synthesis of human papillomavirus type 16 E7 oncoprotein: autonomous protein domains for induction of cellular DNA synthesis and for trans activation

Rawls

Pusztai

Green

1990

The human papillomavirus type 16 E7 protein belongs to a family of nuclear oncoproteins that share amino acid sequences and functional homology. To localize biochemical activities associated with E7, we chemically synthesized the full-length 98-amino-acid polypeptide and several deletion mutant peptides. We show that the E7 polypeptide is biologically active and possesses at least two functional domains; the first induces cellular DNA synthesis in quiescent rodent cells, and the second trans activates the adenovirus E1A-inducible early E2 promoter and binds zinc. Further, each domain is autonomous and can function on separate peptides. DNA synthesis induction activity maps within the N-terminal portion of the molecule, which contains sequences related to adenovirus E1A conserved domains 1 and 2 required for cell transformation and binding of the retinoblastoma gene product. trans-Activation and Zn-binding activities map within the C-terminal portion of the molecule, a region which contains Cys-X-X-Cys motifs. trans Activation does not require protein synthesis, implying a mechanism that involves interaction with a preexisting cellular factor(s). E7 trans activates the adenovirus E2 promoter but not other E1A-inducible viral promoters, suggesting the possibility that E7 trans activation involves interaction, directly or indirectly, with cellular transcription factor E2F.

“…In general, in HPV-16and HPV-18-containing carcinomas and cell lines derived from carcinomas, transcripts encoding a full-length E6 protein have been demonstrated (17,33,37,39). However, quantities of such transcripts have been found to be relatively low (37,39).…”

Section: Cda Pcr Smentioning

confidence: 99%

“…Low amounts of HPV-33-specific unspliced E6 mRNAs could have been missed because the method certainly favors the amplification of the shorter E6* cDNAs. Moreover, in cervical carcinomas and carcinoma-derived cell lines containing HPV-16 or HPV-18, the viral genome is often integrated into the host genome, usually interrupting E1/E2 sequences and leaving the E6 and E7 ORFs undisrupted and transcriptionally active by means of the generation of viral-cellular fusion transcripts (17,29,32,37). The carcinoma that we investigated was found to contain both integrated and episomal HPV-33 DNA, which is not unique since a proportion of HPV-16 containing cervical carcinomas have also been found to contain both viral forms (6,12).…”

Section: Cda Pcr Smentioning

confidence: 99%

“…Such spliced E6/E7 transcripts, two of which have been identified for HPV-16 (E6*I and E6*II), were not present in HPV-6and HPV-11-containing condylomas (4,38). Moreover, HPV-16 and HPV-18 can generate E6/E7-encoding viralcellular fusion transcripts (17,29,32,37), a phenomenon which is uncommon among the low-risk HPVs. Knowledge about the mechanism by which E6/E7 mRNAs of additional high-risk HPVs such as HPV-33 are generated is necessary to determine which viral features are most critical in determining the in vivo oncogenic potential.…”

mentioning

confidence: 93%

See 1 more Smart Citation

Human papillomavirus type 33 in a tonsillar carcinoma generates its putative E7 mRNA via two E6* transcript species which are terminated at different early region poly(A) sites

Snijders

Brule

Schrijnemakers

et al. 1992

Human papillomavirus type 33 (HPV-33)-specific early region transcripts in a tonsillar carcinoma were analyzed by using the RNA polymerase chain reaction method. A total of five cDNA species including species with potential to encode E6*I, E6*II, and E6*III, could be identified. As determined by 3' cDNA end mapping, one E6*I cDNA species was found to utilize a novel early region poly(A) site and was polyadenylated at or near the putative initiation codon of the El open reading frame (ORF). Compared with the HPV-16 and HPV-18 E6* mRNAs, the HPV-33 E6*I and E6*II species utilize different splice acceptor sites, the latter being localized writhin the E7 ORF. Furthermore, HPV-33 E6* mRNAs were found to contain a short overlapping ORF resulting in alternative coding potentials if translation were to start at an internal AUG codon within the E6 region. These results indicate that like HPV-16 and HPV-18, HPV-33 generates E6* mRNAs which may serve as efficient mRNAs for E7. However, HPV-33 has the ability to generate its putative E7 mRNAs by the utilization of two early region poly(A) sites, which offers the possibility of expressing E7 in different ways.