Characterization of two murine (2′‐5′)(A) n ‐dependent endonucleases of different molecular mass

RNase L is considered as the major if not unique target of (2′‐5′)(A) n and therefore as an important intracellular mediator of interferon action. It behaves as an 185‐kDa species in various cell extracts when analyzed by gel filtration. SDS/PAGE analysis of the polypeptides covalently labelled with a (2′‐5′)(A) 4 ‐3′‐[ 32 P]p Cp probe reveals a single 80‐kDa species, thus attesting a multimeric form of the 185‐kDa protein. At variance with such data, mouse spleen extracts reveal an additional 40‐kDa polypeptide with (2′‐5′)(A) n ‐dependent ribonucleolytic activity. This seemingly new form of RNase L migrates as a 40‐kDa polypeptide when analyzed under native or denaturing conditions. It bears some structural similarity with the larger‐molecular‐mass RNase L as revealed by partial proteolysis. It is probably not generated through proteolytic degradation of the 185‐kDa RNase L during extract preparation, although its physiological significance is unknown. Indeed various protease inhibitors do not significantly alter the ratio of 40‐kDa and 185‐kDa (2′‐5′)(A) n ‐dependent ribonucleases; moreover, the (2′‐5′)(A) n ‐ binding capacity of the 40‐kDa polypeptide is more stable than that of the 185‐kDa one.