Expression and splicing patterns of human papillomavirus type‐16 mrnas in pre‐cancerous lesions and carcinomas of the cervix, in human keratinocytes immortalized by HPV 16, and in cell lines established from cervical cancers

Abstract

We have analysed the splicing patterns of human papillomavirus (HPV) type-16 mRNAs in a human epithelial cell line immortalized by HPV 16 (HPKII), in cell lines established from cervical carcinomas (SiHa and CaSki) and in pre-invasive and invasive carcinomas of the cervix. The presence of mRNA species previously described, which could encode the E6, E6I, E6II, E6III, E7, E2, E2C, E4, E5 and L1 proteins, was determined, using the RNA polymerase chain reaction (PCR) technique with primers that flank unique splice sites. The state of the viral DNA in the tumor biopsies was established by Southern blot analysis. The various HPV 16 transcripts could be detected in cell lines and in tumor biopsies. The size of the RNA PCR products were in agreement with the previously mapped splice sites. The full range of transcripts was revealed in the HPKII cell line and in a number of pre-invasive carcinomas. Messenger RNAs which could encode the E6III, E4 and E5 proteins were most prevalent in all types of tumor. The overall results of DNA and RNA analyses in cell lines and tumor specimens indicate that (1) expression of either of the early or late transcripts studied is not specifically related to (a) tumor stage or (b) the physical state of the viral genome; and (2) alterations in the splicing patterns of HPV 16 transcripts may not be involved in tumor progression.

Diagnosing Cervical Cancer and High-Grade Precursors by HPV16 Transcription Patterns

“…Three LSIL and two NIL/M samples were positive for both housekeeping transcripts but negative for any HPV16 transcript and were excluded in the following two paragraphs. In accordance with published data (27), the 1302^5639 spliced transcript was not detected.…”

Stomach Cancer in Transgenic Mice Expressing Human Papillomavirus Type 16 Early Region Genes From a Keratin Promoter

“…The predominant bands seen in these tissues and also in the glandular stomach and vagina/cervix, though not in the duodenum or uterus, correspond to the sizes predicted for m R N A spliced 226~409. This m R N A includes a truncated E6*I open reading frame (Smotkin et al, 1989) and the intact E7 coding sequence; this was previously found to be the predominant splicing pattern of the E6/E7 region in CIN and invasive carcinomas (Smotkin et al, 1989;Sherman et al, 1992). Equimolar amounts of the 200 and 471 nt protected bands would give a > twofold greater intensity of the upper band; we interpret the approximately equal intensities, together with the results shown in Fig.…”

Induction of cell death in human papillomavirus 18-positive cervical cancer cells by E6 siRNA

“…It is known that mRNA coding E6 and E7 is transcribed from the same promoter in the form of a bicistronic transcript, with a number of alternative splicing variants designated as E6*I-IV encoding E6*I proteins, which are truncated forms of E6. [39][40][41] Importantly, E7 is equally and efficiently translated from E6-E7 and E6*I-E7 transcripts. 42 One siRNA, Ex-18E6, was selected within the common region of E6-E7 and E6*I-E7 mRNA so that it recognized and destructed both E6-E7 and E6*I-E7 mRNA, thereby knocking down both E6 and E7 (E6E7-specific siRNA).…”