MOLECULAR-CLONING AND CHARACTERIZATION OF 2 RELATED AND INTERFERON-INDUCED 56-KDA AND 30-KDA PROTEINS HIGHLY SIMILAR TO 2'-5'-OLIGOADENYLATE SYNTHETASE

The 2'-5' oligoadenylate synthetase (OAS) represents a family of inter feron-induced proteins which when activated by double-stranded (ds) RN A polymerizes ATP into 2'-5' linked oligomers with the general formula pppA(2'p5'A)(n), n greater than or equal to 1. Three forms of human O ASs have been described corresponding to proteins of 40/46, 69/71 and 100 kDa (p40/p46, p69/p71 and p100). Polyclonal antibodies raised agai nst p100 cross reacted with the other forms p40/p46, p69/p71 but also with an interferon-induced 56-kDa protein (p56). By screening a cDNA e xpression library, these polyclonal antibodies selected a cDNA encodin g p56. Further studies by the RACE procedure using primers correspondi ng to this cDNA, a p56-related protein of 30 kDa (p30) was isolated. B oth p56 and p30 mRNA are expressed in interferon-treated cells as tran scripts of 2 kb and 1.8 kb, respectively. The 1.8-kb mRNA is homologou s to the 2-kb mRNA but with a 243-nucleotide deletion at position 1011 , which results in a frameshift. Consequently, the p56 and p30 have th eir first 219 amino acid residues identical but differ at their C-term ini. In vitro transcription-translation of p56 and p30 cDNAs generated proteins of 56 and 30 kDa, respectively. The deduced amino acid seque nce of p56 kDa shares strong similarity with the previously cloned OAS s, and contains the subdomains conserved in p40/p46 and p69/p71 forms. Transient expression in HeLa cells indicated that p30 has a cytoplasm ic localization, whereas p56 has cytoplasmic and nucleolar localizatio ns. The p56 isolated from transfected cells was shown to bind dsRNA an d DNA, but it was devoid of 2'-5'OAS activity typical of the three kno wn farms of this enzyme. Thus, p56 and p30 are two related and interfe ron-induced proteins outside the family of 2'-5'OAS, which might have as yet unidentified catalytic activities or functions.