Molecular basis of human poly(A) polymerase recruitment by mPSF.

3' end processing of most eukaryotic precursor-mRNAs (pre-mRNAs) is a crucial cotranscriptional process that generally involves the cleavage and polyadenylation of the precursor transcripts. Within the human 3' end processing machinery, the four-subunit mammalian polyadenylation specificity factor (mPSF) recognizes the polyadenylation signal (PAS) in the pre-mRNA and recruits the poly(A) polymerase α (PAPOA) to it. To shed light on the molecular mechanisms of PAPOA recruitment to mPSF, we used a combination of cryogenic-electron microscopy (cryo-EM) single-particle analysis, computational structure prediction, and in vitro biochemistry to reveal an intricate interaction network.

Concerted structural rearrangements enable RNA channeling into the cytoplasmic Ski238-Ski7-exosome assembly.

The Ski2-Ski3-Ski8 (Ski238) helicase complex directs cytoplasmic mRNAs toward the nucleolytic exosome complex for degradation. In yeast, the interaction between Ski238 and exosome requires the adaptor protein Ski7. We determined different cryo-EM structures of the Ski238 complex depicting the transition from a rigid autoinhibited closed conformation to a flexible active open conformation in which the Ski2 helicase module has detached from the rest of Ski238.

Purification and Reconstitution of the S. cerevisiae TRAMP and Ski Complexes for Biochemical and Structural Studies.

The RNA exosome is a macromolecular machine that degrades a large variety of RNAs from their 3'-end. It comprises the major 3'-to-5' exonuclease in the cell, completely degrades erroneous and overly abundant RNAs, and is also involved in the precise processing of RNAs. To degrade transcripts both specifically and efficiently the exosome functions together with compartment-specific cofactors.