Control of messenger RNA (mRNA) decay rate is intimately connected to translation elongation, but the spatial coordination of these events is poorly understood. The Ccr4-Not complex initiates mRNA decay through deadenylation and activation of decapping. We used a combination of cryo–electron microscopy, ribosome profiling, and mRNA stability assays to examine the recruitment of Ccr4-Not to the ribosome via specific interaction of the Not5 subunit with the ribosomal E-site in Saccharomyces cerevisiae. This interaction occurred when the ribosome lacked accommodated A-site transfer RNA, indicative of low codon optimality. Loss of the interaction resulted in the inability of the mRNA degradation machinery to sense codon optimality. Our findings elucidate a physical link between the Ccr4-Not complex and the ribosome and provide mechanistic insight into the coupling of decoding efficiency with mRNA stability.
Ribosome stalling triggers the ribosome-associated quality control (RQC) pathway, which targets collided ribosomes and leads to subunit dissociation, followed by proteasomal degradation of the nascent peptide. In yeast, RQC is triggered by Hel2-dependent ubiquitination of uS10, followed by subunit dissociation mediated by the RQC-trigger (RQT) complex. In mammals, ZNF598-dependent ubiquitination of collided ribosomes is required for RQC, and activating signal cointegrator 3 (ASCC3), a component of the ASCC complex, facilitates RQC. However, the roles of other components and associated factors of the ASCC complex remain unknown. Here, we demonstrate that the human RQCtrigger (hRQT) complex, an ortholog of the yeast RQT complex, plays crucial roles in RQC. The hRQT complex is composed of ASCC3, ASCC2, and TRIP4, which are orthologs of the RNA helicase Slh1(Rqt2), ubiquitin-binding protein Cue3(Rqt3), and zinc-finger type protein yKR023W(Rqt4), respectively. The ATPase activity of ASCC3 and the ubiquitin-binding activity of ASCC2 are crucial for triggering RQC. Given the proposed function of the RQT complex in yeast, we propose that the hRQT complex recognizes the ubiquitinated stalled ribosome and induces subunit dissociation to facilitate RQC. Cells have evolved various quality control mechanisms to guarantee accurate gene expression 1-4. Ribosome stalling induces quality control mechanisms for mRNA, referred to as No-go decay (NGD) 5-7 , as well as for protein, referred to as ribosome-associated quality control (RQC) 8-11. RQC is conserved throughout species and consists primarily of four steps: (i) recognition of abnormal ribosome stalling; (ii) ubiquitination of specific residue(s) on the stalled ribosome; (iii) dissociation of ribosome into 40S and 60S subunits; and (iv) degradation of the nascent polypeptide on the 60S subunit 2,4. In the first step of RQC, the stalling of a ribosome at a specific sequence results in the formation of a di-ribosome (disome), which consists of the leading stalled ribosome and the following collided ribosome 5,6,10. Cryo-EM structural analysis has shown that the leading stalled ribosome is in the POST-state, with an empty A-site, whereas the colliding ribosome is in a rotated state with hybrid tRNAs 6,8,10. In the second step, the RING-type E3 ubiquitin ligase Hel2 recognizes the ribosome collision and ubiquitinates ribosomal protein uS10 (in yeast, at residue(s) K6/8) 8. In the third step, ubiquitinated ribosomes are dissociated into 40S subunits and 60S ribosome-nascent chain complexes (60S-RNCs), leading to subsequent RQC reactions. We recently proposed a model in which these ubiquitinated ribosomes are targeted by the RQT complex 8. The RQT complex is composed of three proteins: RNA helicase Slh1(Rqt2), ubiquitin-binding protein Cue3(Rqt3), and zinc-finger domain-containing protein yKR023W(Rqt4). The ubiquitin-binding activity of Cue3 and the ATPase activity of Slh1 are crucial for triggering RQC 8. After subunit dissociation, Rqc2 binds to tRNA at the subunit interface of 6...
Haploinsufficiency of SETD5 is implicated in syndromic autism spectrum disorder (ASD), but the molecular mechanism underlying the pathological role of this protein has remained unclear. We have now shown that Setd5 +/mice manifest ASDrelated behavioral phenotypes and that the expression of ribosomal protein genes and rDNA is disturbed in the brain of these mice. SETD5 recruited the HDAC3 complex to the rDNA promoter, resulting in removal of the histone mark H4K16ac and its reader protein TIP5, a repressor of rDNA expression. Depletion of SETD5 attenuated rDNA expression, translational activity, and neural cell proliferation, whereas ablation of TIP5 in SETD5-deficient cells rescued these effects. Translation of cyclin D1 mRNA was specifically down-regulated in SETD5-insufficient cells. Our results thus suggest that SETD5 positively regulates rDNA expression via an HDAC3-mediated epigenetic mechanism and that such regulation is essential for translation of cyclin D1 mRNA and neural cell proliferation.
Read‐through or mutations of a stop codon resulting in translation of the 3′‐UTR produce potentially toxic C‐terminally extended proteins. However, quality control mechanisms for such proteins are poorly understood in mammalian cells. Here, a comprehensive analysis of the 3′‐UTRs of genes associated with hereditary diseases identified novel arrest‐inducing sequences in the 3′‐UTRs of 23 genes that can repress the levels of their protein products. In silico analysis revealed that the hydrophobicity of the polypeptides encoded in the 3′‐UTRs is correlated with arrest efficiency. These results provide new insight into quality control mechanisms mediated by 3′‐UTRs to prevent the production of C‐terminally extended cytotoxic proteins.
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