Distinct RNP complexes of shuttling hnRNP proteins with pre-mRNA and mRNA:Candidate intermediates in formation and export of mRNA

Nascent pre-mRNAs associate with hnRNP proteins in hnRNP complexes, the nat ural substrates for mRNA processing. Several lines of evidence indicate tha t hnRNP complexes undergo substantial remodeling during mRNA formation and export. Here we report the isolation of three distinct types of pre-mRNP an d mRNP complexes from HeLa cells associated with hnRNP A1, a shuttling hnRN P protein. Based on their RNA and protein compositions, these complexes are likely to represent distinct stages in the nucleocytoplasmic shuttling pat hway of hnRNP A1 with its bound RNAs. In the cytoplasm, A1 is associated wi th its nuclear import receptor (transportin), the cytoplasmic poly(A)-bindi ng protein, and mRNA. In the nucleus, A1 is found in two distinct types of complexes that are differently associated with nuclear structures. One clas s contains pre-mRNA and mRNA and is identical to previously described hnRNP complexes. The other class behaves as freely diffusible nuclear mRNPs (nmR NPs) at late nuclear stages of maturation and possibly associated with nucl ear mRNA export. These nmRNPs differ from hnRNPs in that while they contain shuttling hnRNP proteins, the mRNA export factor REF, and mRNA, they do no t contain nonshuttling hnRNP proteins or pre-mRNA. Importantly, nmRNPs also contain proteins not found in hnRNP complexes. These include the alternati vely spliced isoforms D01 and D02 of the hnRNP D proteins, the EO isoform o f the hnRNP E proteins, and LRP130, a previously reported protein with unkn own function that appears to have a novel type of RNA-binding domain. The c haracteristics of these complexes indicate that they result from RNP remode ling associated with mRNA maturation and delineate specific changes in RNP protein composition during formation and transport of mRNA in vivo.