In situ analysis of the transcriptional activity of integrated viral DNA using tyramide-FISH
Van Tine, B.A.; Knops, J.; Broker, T.R.; Chow, L.T.; Moen, P.T.
Developments in Biologicals 106: 381-385
2001
ISSN/ISBN: 1424-6074 PMID: 11761253 Document Number: 536191
Infection by the oncogenic human papillomavirus (HPV) types 16 and type 18 can progress to cancers. Two well studied cervical carcinoma cell lines, SiHa and CaSki, contain two to four copies, or several hundred copies of integrated HPV-16, respectively. To define the chromosomal loci from which HPV mRNAs are transcribed in these cells, we have simultaneously visualized chromosomal DNA territories, HPV DNA or nascent HPV RNA sequences by using a highly sensitive in situ hybridization (T-FISH) technique employing deposition of fluorescent tyramides. We found that, in SiHa cells, nascent HPV RNAs co-localized with both integrated HPV copies on chromosome 13. Surprisingly, in CaSki cells, nascent HPV RNA only co-localized with one minor HPV DNA-positive locus on chromosome 14. The DNA signal intensity of this locus was consistent with a single to a few HPV intergrants. The tyramide methodologies described here provide an in-depth molecular cytological analyses applicable to research and diagnosis.