Interaction between the RNA-dependent ATPase and poly(A) polymerase subunits of the TRAMP complex is mediated by short peptides and important for snoRNA processing.

Jillian S Losh,Sean J Johnson,John Loomis,Jason A Rosenzweig,Alejandra Klauer King,Ambro Van Hoof,Jeremy Bakelar,Lacy Taylor

doi:10.1093/nar/gkv005

Abstract

The RNA exosome is one of the main 3′ to 5′ exoribonucleases in eukaryotic cells. Although it is responsible for degradation or processing of a wide variety of substrate RNAs, it is very specific and distinguishes between substrate and non-substrate RNAs as well as between substrates that need to be 3′ processed and those that need to be completely degraded. This specificity does not appear to be determined by the exosome itself but rather by about a dozen other proteins. Four of these exosome cofactors have enzymatic activity, namely, the nuclear RNA-dependent ATPase Mtr4, its cytoplasmic paralog Ski2 and the nuclear non-canonical poly(A) polymerases, Trf4 and Trf5. Mtr4 and either Trf4 or Trf5 assemble into a TRAMP complex. However, how these enzymes assemble into a TRAMP complex and the functional consequences of TRAMP complex assembly remain unknown. Here, we identify an important interaction site between Mtr4 and Trf5, and show that disrupting the Mtr4/Trf interaction disrupts specific TRAMP and exosome functions, including snoRNA processing.

Highlights

Almost all cellular RNAs undergo extensive processing before becoming fully mature and functional RNAs
The Trf4/5 Air1/2 Mtr4 polyadenylation (TRAMP) complex was initially identified in a yeast twohybrid screen for Mtr4 interacting proteins [6]
In the yeast two-hybrid analysis, the Mtr4-archless interaction with Trf5 98–117 allowed for more robust growth, as we have previously reported for the longer Trf5 53–199 fragment. These results identify an interaction of a small peptide in the unstructured N-terminus of Trf4/5 with the helicase core of Mtr4 that may be important for TRAMP complex assembly

Summary

Introduction

Almost all cellular RNAs undergo extensive processing before becoming fully mature and functional RNAs. The pathways involved in these post-transcriptional modifications, tightly regulated, are not error-free. Cells have evolved a variety of surveillance mechanisms to ensure the fidelity of this RNA maturation process and gene expression. The RNA exosome complex is a major player in both RNA processing and surveillance mechanisms. The RNA exosome has 3 to 5 exoribonuclease activity that is required for removal of 3 extensions from precursor RNAs (e.g. 5.8S rRNA; 1,2) and to degrade misprocessed RNAs (e.g. hypomethylated tRNAi; 3). Many exosome substrates have been identified, but how the exosome distinguishes between substrates that require processing, substrates that require degradation and nonsubstrates is poorly understood

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Results

Conclusion