COMPLEX ALTERNATIVE RNA PROCESSING GENERATES AN UNEXPECTED DIVERSITY OF POLY(A) POLYMERASE ISOFORMS

Multiple forms of poly(A) polymerase (PAPs I, II, and III) cDNA have p reviously been isolated from bovine, human, and/or frog cDNA libraries , PAPs I and II are long forms of the enzyme that contain four functio nal domains: an apparent ribonucleoprotein-type RNA-binding domain, a catalytic region that may be related to the polymerase module, two nuc lear localization signals (NLSs 1 and 2), and a C-terminal Ser/Thr-ric h region, PAP III would encode a truncated protein that lacks the NLSs and the S/T-rich region, To investigate further the structure and exp ression of these forms, we isolated the mouse PAP gene and an intronle ss pseudogene from a mouse liver genomic library, The structure of the gene indicates that different forms of PAP are produced by alternativ e splicing (PAPs I and II) or by competition between polyadenylation a nd splicing (PAP III), The pseudogene appears to reflect yet another f orm of long PAP, which we call PAP TV, Mouse PAP III and two additiona l truncated forms, PAPs V and VI, which would be produced by use of po ly(A) sites in adjacent introns, were also isolated from a mouse brain cDNA library, RNase protection and reverse transcription-PCR analyses showed that PAP II, V, and VI are expressed in all tissues tested but that PAP I and/or IV and III are tissue specific, However, immunoblot analysis detected only the long forms, raising the possibility that t he short-form RNAs are not translated. Purified recombinant baculoviru s-expressed PAPs were tested in several in vitro assays, and the short forms were found to be inactive. We discuss the possible significance of this complex expression pattern.