Abstract
Recent studies have suggested that a sub-complex of RNA polymerase II composed of Rpb4 and Rpb7 couples the nuclear and cytoplasmic stages of gene expression by associating with newly made mRNAs in the nucleus, and contributing to their translation and degradation in the cytoplasm. Here we show by yeast two hybrid and co-immunoprecipitation experiments, followed by ribosome fractionation and fluorescent microscopy, that a subunit of the Ccr4-Not complex, Not5, is essential in the nucleus for the cytoplasmic functions of Rpb4. Not5 interacts with Rpb4; it is required for the presence of Rpb4 in polysomes, for interaction of Rpb4 with the translation initiation factor eIF3 and for association of Rpb4 with mRNAs. We find that Rpb7 presence in the cytoplasm and polysomes is much less significant than that of Rpb4, and that it does not depend upon Not5. Hence Not5-dependence unlinks the cytoplasmic functions of Rpb4 and Rpb7. We additionally determine with RNA immunoprecipitation and native gel analysis that Not5 is needed in the cytoplasm for the co-translational assembly of RNA polymerase II. This stems from the importance of Not5 for the association of the R2TP Hsp90 co-chaperone with polysomes translating RPB1 mRNA to protect newly synthesized Rpb1 from aggregation. Hence taken together our results show that Not5 interconnects translation and transcription.
Highlights
The life of an mRNA molecule in eukaryotic cells is considered to be the sum of distinct events separated in time and space
Not5 interacts with the Rpb4 subunit of polymerase that is known to readily dissociate from the rest of the polymerase, and it is essential for Rpb4 to associate with mRNAs at the completion of transcription to contribute to translation and mRNA degradation in the cytoplasm
R2TP is composed of the Tah1 tetratricopeptide repeat (TPR) protein, Pih1 and the two AAA+ ATPases, Rvb1 and Rvb2
Summary
The life of an mRNA molecule in eukaryotic cells is considered to be the sum of distinct events separated in time and space This separation seems to constitute the characteristic difference distinguishing eukaryotes from prokaryotes, where translation is co-transcriptional and occurs in a single cellular compartment. The heptapeptide repeatcontaining C-terminal domain (CTD) of the largest subunit of eukaryotic RNA polymerase II (RNA Pol II) was found to direct post-transcriptional RNA processing events. It serves as a landing platform for components of the machines involved in mRNA capping, splicing, and mRNA export [1,2,3]. An RNA Pol II subunit, Rpb, has been suggested to play roles in the nucleus during the transcription process, and subsequently in the cytoplasm, contributing to both the RNA degradation and translation processes [4,5]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
