2′-adenylated derivatives of Ap3A activate RNase L

The exact physiological function of Ap(3)A (A5'ppp5 " A, 5'5 " diadenosine triphosphate) remains unclear. Previously we have demonstrated that the human p46 2-5A synthetase (OAS1) efficiently utilises Ap(3)A as an acceptor substrate for oligoadenylate synthesis. Here we show that Ap(3)A(2'p5'A)(n) oligonucleotides can activate the 2-5A-dependent RNase (RNase L), when the number of 2',5'-linked adenyl residues is two or more. Under the experimental conditions applied the half-maximal activation (AC(50)) of RNase L for 2'-adenylated Ap3A derivatives was determined to be in nanomolar range while the AC(50) for 2-5A(3) was 0.4 nM, The Ap(3)A(2'p5'A)(n) oligonucleotides are thus less effective in activating RNase L than 2-5A, We also investigated the occurrence of 2'-adenylated Ap(3)A in interferon and poly(I).poly(C)-treated HeLa cells. In purified trichloroacetic acid-soluble extracts about 40% of RNase L-activating material is resistant to phosphatase treatment, whereas the removal of 5'-terminal phosphates greatly reduces the activating properties of 2-5A, We assume that this activity at least partly may be associated with the presence of 2'-adenylated Ap(n)A derivatives with blocked 5'-terminal phosphates. (C) 1999 Federation of European Biochemical Societies.