Abstract
Non-coding RNAs (ncRNAs) play critical roles in gene regulation. In eukaryotic cells, ncRNAs are processed and/or degraded by the nuclear exosome, a ribonuclease complex containing catalytic subunits Dis3 and Rrp6. The TRAMP (Trf4/5-Air1/2-Mtr4 polyadenylation) complex is a critical exosome cofactor in budding yeast that stimulates the exosome to process/degrade ncRNAs and human TRAMP components have recently been identified. Importantly, mutations in exosome and exosome cofactor genes cause neurodegenerative disease. How the TRAMP complex interacts with other exosome cofactors to orchestrate regulation of the exosome is an open question. To identify novel interactions of the TRAMP exosome cofactor, we performed a high copy suppressor screen of a thermosensitive air1/2 TRAMP mutant. Here, we report that the Nab3 RNA-binding protein of the Nrd1-Nab3-Sen1 (NNS) complex is a potent suppressor of TRAMP mutants. Unlike Nab3, Nrd1 and Sen1 do not suppress TRAMP mutants and Nrd1 binding is not required for Nab3-mediated suppression of TRAMP suggesting an independent role for Nab3. Critically, Nab3 decreases ncRNA levels in TRAMP mutants, Nab3-mediated suppression of air1/2 cells requires the nuclear exosome component, Rrp6, and Nab3 directly binds Rrp6. We extend this analysis to identify a human RNA binding protein, RALY, which shares identity with Nab3 and can suppress TRAMP mutants. These results suggest that Nab3 facilitates TRAMP function by recruiting Rrp6 to ncRNAs for processing/degradation independent of Nrd1. The data raise the intriguing possibility that Nab3 and Nrd1 can function independently to recruit Rrp6 to ncRNA targets, providing combinatorial flexibility in RNA processing.
Highlights
Non-coding RNAs are fast emerging as key regulators of gene expression in eukaryotic cells [1,2]
Eukaryotic genomes from yeast to man express numerous non-coding RNAs that regulate the expression of messenger RNAs encoding the proteins vital for cell and body function
To better understand TRAMP function, we performed a genetic screen to identify genes that improve the growth of TRAMP mutant yeast cells that grow poorly at high temperature
Summary
Non-coding RNAs (ncRNAs) are fast emerging as key regulators of gene expression in eukaryotic cells [1,2]. Understanding how ncRNAs are produced, processed, and destroyed is critical to elucidating gene regulation and has become an area of major research focus in recent years. Transcriptional termination and 3’-end processing of ncRNAs are functionally linked but how they are coupled is poorly characterized [5]. In Saccharomyces cerevisiae, unlike most mRNAs, ncRNAs are not terminated and 3’-end processed by the conventional cleavage and polyadenylation machinery, but by a trio of evolutionarily conserved complexes: the RNA exosome, the TRAMP complex, and the NNS complex [5]. The interactions and mechanisms employed by these complexes to terminate and process/degrade ncRNAs are not well understood
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