INTERFERON ACTION - COVALENT LINKAGE OF (2'-5')PPPAPAPA (P-32) PCP TO (2'-5')(A)N-DEPENDENT RIBONUCLEASES IN CELL-EXTRACTS BY ULTRAVIOLET-IRRADIATION
- 1 January 1982
- journal article
- research article
- Vol. 257 (15) , 8584-8587
Abstract
One of the mediators of interferon action is an endonuclease system. This consists of (2''-5'')(A)n synthetase, which , if activated by double-stranded RNA, converts ATP into (2''-5'')(A)n and RNase L, a latent endoribonuclease, which binds (2''-5'')(A)n and is thereby activated. A derivative of (2''-5'')(A)n (i.e., (2''-5'')pppApApA[32P]pCp) can be convalently cross-linked by UV irradiation to a protein in cytoplasmic extracts from mouse Ehrlich ascites tumor cells. This protein has a MW of .apprx. 77,000 as determined by gel electrophoresis in sodium dodecyl sulfate. It appears to be identical with RNase L according to the following criteria: co-chromatography on DEAE-cellulose and Sephacryl S300. The gel filtration in Sephacryl S300 reveals that the apparent MW of the protein is between 70,000 and 90,000, indicating that it is a monomer. The cross-linking is oligonucleotide specific. It is inhibited by 10 nM (2''-5'')pppApApA or 1 .mu.M (2''-5'')ApApA, i.e., compounds known to block, even at low concentration, the binding of (2''-5'')pppApApApCp to RNase L. (3''-5'')ApApA inhibits only at a 0.1-1 mM concentration, and 1 mM ATP, 2''-AMP or 5'', 3''-pCp have no effect. (2''-5'')-pppApApApCp was also cross-linked to a protein with a MW of .apprx. 78,000 (as determined by gel electrophoresis in sodium dodecyl sulfate) in cytoplasmic extracts from human (HeLa) cells and to protein(s) with MW of 75,000-77,000 (as determined by gel electrophoresis in sodium dodecyl sulfate) in nuclear extracts from Ehrlich ascites tumor cells.This publication has 14 references indexed in Scilit:
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