THE DEAH-BOX PROTEIN PRP22 IS AN ATPASE THAT MEDIATES ATP-DEPENDENT MESSENGER-RNA RELEASE FROM THE SPLICEOSOME AND UNWINDS RNA DUPLEXES

Of the proteins required for pre-mRNA splicing, at least four, the DEA H-box proteins, are closely related due to the presence of a central ' RNA helicase-like' region, and extended homology through a large porti on of the protein. A major unresolved question is the function of thes e proteins. Indirect evidence suggests that several of these proteins are catalysts for important structural rearrangements in the spliceoso me. However, the mechanism for the proposed alterations is presently u nknown. We present evidence that PRP22, a DEAH-box protein required fo r mRNA release from the spliceosome, unwinds RNA duplexes in a concent ration-and ATP-dependent manner. This demonstrates that PRP22 can modi fy RNA structure directly. We also show that the PRP22-dependent relea se of mRNA from the spliceosome is an ATP-dependent process and that r ecombinant PRP22 is an ATPase, Nonhydrolyzable ATP analogs did not sub stitute for ATP in the RNA-unwinding reaction, suggesting that ATP hyd rolysis is required for this reaction. Specific mutation of a putative ATP phosphate-binding motif in the recombinant protein eliminated the ATPase and RNA-unwinding capacity. Significantly, these data suggest that the DEAH-box proteins act directly on RNA substrates within the s pliceosome.