Mapping of the human RNASEL promoter and expression in cancer and normal cells.
2',5'-Oligoadenylate (2-5A)-dependent RNase L is a ubiquitous endoribonuclease of higher vertebrates that functions in the interferon (IFN) antiviral response by degrading both viral and cellular single-stranded RNA (ssRNA). In addition, the RNase L gene, RNASEL, was mapped to the hereditary prostate cancer 1 (HPC1) gene. Previous analyses of human RNASEL determined its exon/intron structure but lacked a description of the promoter region. We thus mapped the RNASEL transcriptional start site using 5'-rapid amplification of cDNA ends (5'-RACE) and primer extension methods with RNA from human histiocytic lymphoma U937 cells. The promoter sequence was analyzed for potential transcription factor binding sites. Although a canonical IFN-gamma activation site (GAS) element (TTCCAAGAA) was identified (nucleotides -155 to -147), there was only slight induction of RNASEL promoter-reporter activity or of endogenous RNase L expression in response to IFN-alpha or IFN-gamma. Several sites for tissue-specific and general promoters were observed, however, which could explain the widespread expression of RNase L in mammalian cells. Accordingly, RNase L levels were determined and compared in different human and rodent cancer and normal cell types using a radiolabeled 2-5A derivative. In addition, levels of RNase L were established in various normal human tissues and cell types by immunoblotting and immunohistochemistry. Our findings are the first description of the human RNASEL promoter that allows constituitive expression in a range of normal and neoplastic cell types.
Zhou
Aimin
A
Department of Chemistry, Cleveland State University, Cleveland, OH 44115, USA.
Molinaro
Ross J
RJ
Malathi
Krishnamurthy
K
Silverman
Robert H
RH
D000818:Animals; D045744:Cell Line, Tumor; D004722:Endoribonucleases / Q000096:biosynthesis* / Q000235:genetics; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics; D006801:Humans; D007371:Interferon-gamma / Q000378:metabolism; D008297:Male; D051379:Mice; D009369:Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D012367:RNA, Viral / Q000378:metabolism; D020218:Response Elements / Q000235:genetics*; D014777:Virus Diseases / Q000378:metabolism
D012367:RNA, Viral; D007371:Interferon-gamma; D004722:Endoribonucleases; C031956:2-5A-dependent ribonuclease