Codon Optimality Is a Major Determinant of mRNA Stability

“Alignment and Half-Life Calculation Reads were aligned to the S. cerevisiae reference genome using bowtie (Langmead et al., 2009), with the unaligned reads then aligned to the sequences of the controls in the same way.”

“Several ribosomal profiling studies have failed to detect codon-specific differences in the translation of optimal and non-optimal codons (Ingolia et al., 2009; Qian et al., 2012; Charneski and Hurst, 2013).”

“The overall result is not surprising, as codon bias has been well studied (Gustafsson et al., 2004); however, the pattern of codon usage demonstrates that certain classes of mRNAs predominately use either optimal or non-optimal codons (Figures 3A and 3B; overrepresented codons in yellow,…”

“Critically, optimal codons are postulated to be decoded faster and more accurately by the ribosome than non-optimal codons (Akashi, 1994; Drummond and Wilke, 2008), which are hypothesized to slow translation elongation (Novoa and Ribas de Pouplana, 2012; Tuller et al., 2010).”

“Critically, optimal codons are postulated to be decoded faster and more accurately by the ribosome than non-optimal codons (Akashi, 1994; Drummond and Wilke, 2008), which are hypothesized to slow translation elongation (Novoa and Ribas de Pouplana, 2012; Tuller et al., 2010).”

“The rare codons used in our previous studies belong to a general class of codons defined as non-optimal (Pechmann and Frydman, 2013; dos Reis et al., 2004).”

“These transcripts were determined to be stable both previously and in our genome-wide analysis (median half-life = 43.4 min; Wang et al., 2002).”

“Ribosomal run-off experiments were performed similarly to normal polyribosome analysis, except cells were resuspended in media lacking glucose for 10 min before harvesting (Coller and Parker, 2005).”

“For instance, large-scale RNA profiling experiments have demonstrated that tRNA concentrations vary widely between proliferating and differentiating cells (Gingold et al., 2014).”

“The term codon optimality has been introduced in an attempt to define the differential recognition of codons by the translational apparatus (Zhou et al., 2009; Pechmann and Frydman, 2013).”

“Despite being targeted by a common decay pathway, turnover rates for individual yeast mRNAs differ dramatically, with half-lives ranging from <1 min to 60 min or greater (Coller and Parker, 2004).”

“This effect was not dependent on RNA surveillance pathways such as No-Go, Nonsense-Mediated, or Non-Stop Decay (Shoemaker and Green, 2012).”

“We repeated our analysis of codon usage versus mRNA half-life using mRNA decay rates obtained by Miller et al. (2011).”

“RNA-Seq rpb1-1 mutant cells (Nonet et al., 1987) were grown to mid-log phase at 24 C and shifted to a non-permissive temperature of 37 C.”

“Although it is firmly established that deadenylation is the rate-limiting step in mRNA turnover, we and others have observed that specific mRNAs persist in cells as ‘‘stable’’ deadenylated species (Hu et al., 2009; Muhlrad et al., 1995).”

“Northern RNA Analysis and Sucrose Density Gradients Northern RNA analysis of GAL-driven reporters and sucrose density gradients for polyribosome analysis were performed as previously described (Hu et al., 2009).”

“Several ribosomal profiling studies have failed to detect codon-specific differences in the translation of optimal and non-optimal codons (Ingolia et al., 2009; Qian et al., 2012; Charneski and Hurst, 2013).”

“Several ribosomal profiling studies have failed to detect codon-specific differences in the translation of optimal and non-optimal codons (Ingolia et al., 2009; Qian et al., 2012; Charneski and Hurst, 2013).”

“Numerous examples of each have been described (Goldstrohm et al., 2007; Olivas and Parker, 2000).”

“However, these features regulate mRNA stability predominantly in a transcript-specific manner through binding of regulatory factors and cannot account for the wide variation in half-lives observed across the entire transcriptome (Geisberg et al., 2014).”

“…introduced the synthetic ORFs into a reporter bearing the 50 and 30 UTRs ofMFA2, a well-studied mRNA that is rapidly degraded in the cell (half-life = 3.0 min), a phenomenon shown to be mediated, in part, by elements encoded within its 30 UTR (LaGrandeur and Parker, 1999; Muhlrad and Parker, 1992).”

“The link between rare codons and enhanced mRNA turnover rates of reporter mRNAs is consistent with earlier observations for several endogenous transcripts in yeast (Caponigro et al., 1993; Hoekema et al., 1987).”

“The term codon optimality has been introduced in an attempt to define the differential recognition of codons by the translational apparatus (Zhou et al., 2009; Pechmann and Frydman, 2013).”

“Numerous examples of each have been described (Goldstrohm et al., 2007; Olivas and Parker, 2000).”

“We have previously shown that inclusion of a cluster of rare arginine codons within the open reading frame (ORF) of a reporter mRNA dramatically enhanced its turnover (Hu et al., 2009; Sweet et al., 2012).”

“…introduced the synthetic ORFs into a reporter bearing the 50 and 30 UTRs ofMFA2, a well-studied mRNA that is rapidly degraded in the cell (half-life = 3.0 min), a phenomenon shown to be mediated, in part, by elements encoded within its 30 UTR (LaGrandeur and Parker, 1999; Muhlrad and Parker, 1992).”

“(C) Representative A260 trace of sucrose density gradient analysis under run-off (D) Distribution of the optimal and non-optimal HIS3 reporters and the RPS20 and toring factor is the DEAD-box RNA helicase DHH1, an integral component of the mRNA decaymachinery (Presnyak and Coller, 2013) that has been shown to act as a…”

“Degradation of most mRNAs in Saccharomyces cerevisiae is initiated by removal of the 30 poly(A) tail (deadenylation), followed by cleavage of the 50 7mGpppN cap (decapping) and exonucleolytic degradation of the mRNA body in a 50-30 direction (Coller and Parker, 2004; Ghosh and Jacobson, 2010).”

“The link between rare codons and enhanced mRNA turnover rates of reporter mRNAs is consistent with earlier observations for several endogenous transcripts in yeast (Caponigro et al., 1993; Hoekema et al., 1987).”

“Therefore, codon optimality is hypothesized to play an important role in modulation of translation elongation rates and the kinetics of protein synthesis (Krisko et al., 2014; Novoa and Ribas de Pouplana, 2012; Pechmann and Frydman, 2013; dos Reis et al., 2004).”

“RNA features that influence transcript stability have long been sought, and some sequence and/or structural elements located within 50 and 30 UTRs have been implicated in contributing to the decay of a subset of mRNAs (Lee and Lykke-Andersen, 2013; Muhlrad and Parker, 1992; Geisberg et al., 2014).”

“The modification of rare codons in protein-coding sequences with synonymous frequently occurring codons, so-called codon optimization, can result in order-of-magnitude changes in expression levels.(5,6) Modification of the 50 mRNA cap can also enhance mRNA translation by inhibiting RNA decapping and improving resistan…”

“Codon Optimality: The fraction of the optimal codon for each gene was calculated using a custom script based on codon stabilization coefficients presented in a previous report (Presnyak et al., 2016).”