RNA degradation paths in a 12-subunit nuclear exosome complex

Abstract

The eukaryotic exosome is a conserved RNA-degrading complex that functions in RNA surveillance, turnover and processing. How the same machinery can either completely degrade or precisely trim RNA substrates has long remained unexplained. Here we report the crystal structures of a yeast nuclear exosome containing the 9-subunit core, the 3'-5' RNases Rrp44 and Rrp6, and the obligate Rrp6-binding partner Rrp47 in complex with different RNAs. The combined structural and biochemical data of this 12-subunit complex reveal how a single-stranded RNA can reach the Rrp44 or Rrp6 active sites directly or can bind Rrp6 and be threaded via the central channel towards the distal RNase Rrp44. When a bulky RNA is stalled at the entrance of the channel, Rrp6-Rrp47 swings open. The results suggest how the same molecular machine can coordinate processive degradation and partial trimming in an RNA-dependent manner by a concerted swinging mechanism of the two RNase subunits.

CryoEM structure of yeast cytoplasmic exosome complex

“…In the RNA-bound complex, this tail is released from the binding pocket and refolds to become the extension of Rrp45's C-terminal α-helix, which no longer interacts with Rrp44 ( Figure 4B). A similar conformational change in the C-terminal tail of Rrp45 was observed through X-ray crystallography [15]. The dramatic conformational change of Rrp44's C-terminal portion also markedly decreased the interaction area between the RNase II-like domain and the rest of the Exo10 complex from ~2 573 Å 2 in the RNA-free state ( Figure 5A) to ~913 Å 2 in the RNA-bound state ( Figure 5B).…”

“…As we and others have previously discovered, RNA molecules with a long enough single-strand (ss) 3 overhang induce a major rearrangement of the Rrp44 RNase II-like domain relative to the core complex so that the ssRNA substrates coming out of the core directly thread into the exonuclease active site for degradation [13][14][15]17]. However, how this major conformational change in Rrp44 occurs upon RNA substrate binding was unknown.…”

“…Rrp44 is a multi-domain protein with a processive hydrolytic exonuclease activity at its C-terminal region, which is homologous to bacterial RNase II, and a relatively weak endonuclease activity at its N-terminal region [10][11][12]. Previous structural studies have shown that Rrp44 specifically interacts with Rrp41, Rrp43 and Rrp45 of the core complex, aligning with the central channel's exit of the core complex [13][14][15]. The Rrp44-bound exosome complex is considered to be an important active core complex in eukaryotes and is termed Exo10 [16].…”

“…In the best studied Saccharomyces cerevisiae system, Rrp6, an RNase D-type enzyme with distributive exonuclease activity, interacts physically and functionally with the exosome in the nucleus [18]. Rrp6 has been revealed to interact with the Exo10 complex at the top region, next to the entry site of the core [14,15,19]. In the cytoplasm, Ski7, a putative GTPase, bridges the exosome with the ribosome complex and the Ski2/3/8 complex, a cofactor complex, thus promoting recognition and degradation of aberrant mRNAs by the exosome [20].…”

The S. pombe mRNA decapping complex recruits cofactors and an Edc1-like activator through a single dynamic surface

“…After deadenylation, the mRNA is unstable and can be degraded in one of two independent pathways. In the 3 ′ -5 ′ degradation pathway, the mRNA body is processively degraded by the exosome complex (Mitchell et al 1997;Makino et al 2015), after which the scavenger decapping enzyme exploits a well-regulated mechanism to hydrolyze the remaining 5 ′ cap structure Neu et al 2015). In the 5 ′ -3 ′ degradation pathway, the deadenylated 3 ′ end of the mRNA recruits the Lsm1-7:Pat1 complex (Tharun et al 2000;Sharif and Conti 2013).…”