Messenger RNA biogenesis and export are processes of fundamental importance in eukaryotic gene expression, alterations in which have been implicated in numerous human diseases. This research field is now in the stage of exponential growth. The view that has been emerging in the past several years is that the synthesis and processing of mRNA precursors and the assembly of the export-competent mRNP complexes is carried out by an integrated """"""""mRNA factory"""""""", which comprises RNA polymerase II and numerous processing and export factors, many of which interact with the RNA polymerase II and/or with each other. Implicit in this model is a complex network of the quality control checkpoints, all of which must be passed in order for the mRNA to be successfully exported and to enter the cytoplasmic transactions. Poly(A) binding protein (PABP) is a prototypical RRM (RNA Recognition Motif)-containing RNA binding protein that is conserved in all eukaryotes. Our key hypothesis is that PABP, the key mediator of the role of poly(A) tails in the cell and a component of polyadenylation machinery, has a conserved nuclear function in facilitating mRNA biogenesis and export. While the cytoplasmic functions of PABP in mRNA stability and translation are well understood, its role in these nuclear processes, and particularly in the inner workings of the polyadenylation-dependent quality control checkpoint of mRNA biogenesis and export, as well as its relationships to other trans acting factors involved in it, are poorly understood. These issues will be addressed, using Saccharomyces cerevisiae as model system. The following Specific Aims will be pursued: 1) To genetically separate the function of S. cerevisiae Pablp in mRNA biogenesis from its cytoplasmic functions, 2) To test the hypothesis that Pab1 p acts in the early events of mRNA biogenesis, at least some of which occur near the site of transcription, and 3) To identify additional factors cooperating with Pablp in mRNA biogenesis, and elucidate the consequences of Pablp recruitment to mRNA uncoupled from the normal 3' end mRNA processing. ? ?
