pre-mRNA 3’-end processing

 

pre-mRNA 3’ -end maturation is part of a general scheme of pre-mRNA processing events comprising 5'-capping and intron splicing. All these maturation events are essential and tightly coupled and controlled for proper gene expression. mRNAs poly(A) tails are produced by cleavage and polyadenylation of the pre-mRNA molecule. Occurring co-transcriptionally, pre-mRNA 3’-end processing is critical for termination of transcription and mRNA export. As opposed to the striking divergence of the cis-acting sequence elements that direct cleavage and polyadenylation, the protein components of the pre-mRNA 3’-end processing complexes are quite well conserved from yeast to mammals. In metazoans, cleavage of the precursor requires the trimeric complex cleavage stimulation factor (CstF) and the cleavage and polyadenylation specificity factor CPSF. Both of them are crucial to identify during a preliminary step the precise sequence elements on the precursor where cleavage, and hence polyadenylation thereafter, would occur. Additional factors are then recruited, CFIm, CFIIm, and the poly(A)-polymerase PAP, to stabilize the initial interaction and trigger the processing. A network of physical interactions between subunits of the 3’-end processing machinery and the transcription apparatus has been partially drawn that could explain to some extent how processing, transcription termination and export can be regulated. Many interactions between the pre-mRNA 3'-end processing factors have been reported for the human, drosophila and yeast systems. 

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CstF is a multimeric complex essential for the reaction to occur. In human and drosophila, CstF is composed of CstF-50, CstF-64 and CstF-77. The more likely yeast counterparts are respectively, Pfs2, Rna15 and Rna14 . CstF-50 exhibits characteristic WD-repeats which are involved in the assembly of multi-protein factors. CstF-64 bears an RRM-type RNA-binding domain required for the recognition of U/GU-rich elements located downstream of the poly(A) site. It plays a key role in the choice of the cleavage site and hence, in the efficiency of the reaction. CstF-77 is critical for the assembly of the complex, bridging CstF64 and CstF-50. It is the prototypical Half-a-TPR-containing (HAT) protein. Moreover, CstF-77 is located at the crossroads in the network of interactions with other 3’-end formation factors such as CPSF and CFIIm. It connects CstF to CPSF160 and hPcf11. Mutations in Rna15 and Rna14 not only impair formation of the mRNA 3’-end but also prevent RNA polymerase II to terminate transcription properly. Export of the imperfect transcripts is affected and, as a consequence, they are subsequently degraded. A growing number of structural studies have shed light on how the catalytic reactions and the regulation may occur in this complex biological machinery. The structure of protein interacting domains and protein– RNA complexes have been also reported. Many basic questions are still open such as the exact subunit composition of some specific complexes and their overall architecture and role in the reaction.

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