40S subunit dissociation and proteasome-dependent RNA degradation in nonfunctional 25S rRNA decay

Abstract

Eukaryotic cells have quality control systems that eliminate nonfunctional rRNAs with deleterious mutations (nonfunctional rRNA decay, NRD). We have previously reported that 25S NRD requires an E3 ubiquitin ligase complex, which is involved in ribosomal ubiquitination. However, the degradation process of nonfunctional ribosomes has remained unknown. Here, using genetic screening, we identified two ubiquitin-binding complexes, the Cdc48-Npl4-Ufd1 complex (Cdc48 complex) and the proteasome, as the factors involved in 25S NRD. We show that the nonfunctional 60S subunit is dissociated from the 40S subunit in a Cdc48 complex-dependent manner, before it is attacked by the proteasome. When we examined the nonfunctional 60S subunits that accumulated under proteasome-depleted conditions, the majority of mutant 25S rRNAs retained their full length at a single-nucleotide resolution. This indicates that the proteasome is an essential factor triggering rRNA degradation. We further showed that ribosomal ubiquitination can be stimulated solely by the suppression of the proteasome, suggesting that ubiquitin-proteasome-dependent RNA degradation occurs in broader situations, including in general rRNA turnover.

Cdc48-associated complex bound to 60S particles is required for the clearance of aberrant translation products

“…Finally, Cdc48 was implicated in the degradation of nonfunctional 60S subunits (25S NRD for nonfunctional rRNA decay) by promoting the dissociation of stalled 80S, thus allowing the recruitment of the proteasome to stalled 60S to perform ribosomal proteins proteolysis (29). These results are consistent with the hypothesis that after dissociation of stalled ribosomes, the 60S particles are not recycled to perform new rounds or translation but targeted for proteasomal degradation, together with the aberrant nascent peptide.…”

The VCP/p97 system at a glance: connecting cellular function to disease pathogenesis

“…Ohno and colleagues recently discovered that this process is mediated by Cdc48 and the proteasome (Fujii et al, 2012). They showed that ribosomes with defective 25S rRNA in the 60S subunit are selectively ubiquitylated by a ubiquitin ligase complex containing Mms1 and Rtt101.…”

“…They showed that ribosomes with defective 25S rRNA in the 60S subunit are selectively ubiquitylated by a ubiquitin ligase complex containing Mms1 and Rtt101. Subsequently, the Cdc48-Ufd1-Npl4 cofactor complex separates the 60S subunit from the ribosome and facilitates the degradation of ubiquitylated ribosomal proteins by the proteasome (Fujii et al, 2012). It is tempting to speculate that Cdc48 does this by disassembling the individual components of the 60S subunit.…”

“…This process is different from starvation-induced degradation of ribosomes in the lysosome (termed 'ribophagy'), which is also governed by Cdc48 (Ossareh-Nazari et al, 2010). Unlike nonfunctional rRNA decay, ribophagy requires the Cdc48 cofactor Ufd3 (also known as Doa1) (Fujii et al, 2012;Ossareh-Nazari et al, 2010), suggesting that the VCP/p97 system can decide the fate of substrates through degradation by either the proteasome or autophagy.…”

Assembly and quality control of protein phosphatase 1 holoenzyme involve Cdc48-Shp1 chaperone

“…These diverse functions of Cdc48/VCP/p97 are mediated by specific adaptors. The best known is the Npl4-Ufd1 complex that is used for Cdc48/VCP/p97 to extract misfolded protein substrates from the ER (Wolf and Stolz, 2012) and to dissociate nonfunctional 60S ribosome subunits for proteasomal degradation (Fujii et al, 2012). Cdc48-Npl4-Ufd1 also remodels chromatin-binding proteins for DNA damage repair (Acs et al, 2011) and promotes turnover of RNA polymerase II upon UV irradiation (Verma et al, 2011).…”