Summary Emerging studies have linked the ribosome to more selective control of gene regulation. However, an outstanding question is whether ribosome heterogeneity at the level of core ribosomal proteins (RPs) exists and enables ribosomes to preferentially translate specific mRNAs genome-wide. Here, we measured the absolute abundance of RPs in translating ribosomes and profiled transcripts that are enriched or depleted from select subsets of ribosomes within embryonic stem cells. We find that heterogeneity in RP composition endows ribosomes with differential selectivity for translating subpools of transcripts including those controlling metabolism, cell cycle, and development. As an example, mRNAs enriched in binding to RPL10A/uL1-containing ribosomes are shown to require RPL10A/uL1 for their efficient translation. Within several of these transcripts, this level of regulation is mediated, at least in part, by internal ribosome entry sites. Together, these results reveal a critical functional link between ribosome heterogeneity and the post-transcriptional circuitry of gene expression.

Emerging evidence suggests a regulatory function of the ribosome in directing how the genome is translated in time and space. However, how this regulation is encoded in mRNA sequence remains largely unknown. Here we uncover unique RNA regulons embedded in Homeobox (Hox) 5′UTRs that confer ribosome-mediated control of gene expression. These structured RNA elements, resembling viral Internal Ribosome Entry Sites (IRESes), are found in subsets of Hox mRNAs. They facilitate ribosome recruitment and require Ribosomal Protein L38 for their activity. Despite numerous layers of Hox gene regulation, these IRES elements are essential for converting Hox transcripts into proteins to pattern the mammalian body plan. This specialized mode of IRES-dependent translation is enabled by a regulatory element, the Translational Inhibitory Element (TIE), which blocks cap-dependent translation of these transcripts. Together, these data uncover a new paradigm for ribosome-mediated control of gene expression and organismal development.

Quality control mechanisms operate in various steps of ribosomal biogenesis to ensure the production of functional ribosome particles. It was reported previously that mature ribosome particles containing nonfunctional mutant rRNAs are also recognized and selectively removed by a cellular quality control system (nonfunctional rRNA decay [NRD]). Here, we show that the NRD of 25S rRNA requires a ubiquitin E3 ligase component Rtt101p and its associated protein Mms1p, identified previously as factors involved in DNA repair. We revealed that a group of proteins associated with nonfunctional ribosome particles are ubiquitinated in a Rtt101-Mms1-dependent manner. 25S NRD was disrupted when ubiquitination was inhibited by the overexpression of modified ubiquitin molecules, demonstrating a direct role for ubiquitin in this pathway. These results uncovered an unexpected connection between DNA repair and the quality control of rRNAs. Our findings support a model in which responses to DNA and rRNA damages are triggered by a common ubiquitin ligase complex during genotoxic stress harmful to both molecules.[Keywords: Ubiquitin; ribosome; genotoxic stress; quality control; rRNA] Supplemental material is available at http://www.genesdev.org. Gene mutations often result in the production of nonfunctional RNA molecules. In addition, RNA itself is continuously damaged by endogenous and exogenous stress, including ionizing radiation, exposure to certain chemical compounds, and the intracellular generation of reactive oxygen species (Bregeon and Sarasin 2005). Rare but measurable errors in transcription also produce mutant RNAs that do not properly fulfill their roles and aims. In order to avoid a breakdown of cellular order, it is important for cells to detect and selectively dismantle such irregular RNA molecules continuously. It is well documented that various types of aberrant RNAs are selectively removed in eukaryotic cells (Doma and Parker 2007). Three pathways requiring distinct factors degrade different classes of aberrant mRNAs, including mRNAs with a nonsense mutation in their ORFs (nonsensemediated mRNA decay) (Isken and Maquat 2007), mRNAs with no termination codon (nonstop mRNA decay) van Hoof et al. 2002), and mRNAs with a highly stable structure that prevents ribosomal progression (no-go mRNA decay) (Doma and Parker 2006). Recently, it has been reported that tRNAs with hypomodifications are also selectively degraded in vivo, indicating that stable RNAs are monitored by cellular quality control systems (Kadaba et al. 2004;Chernyakov et al. 2008). However, it is not clear how the quality control of ribosomal RNAs (rRNAs), another species of stable RNAs, is achieved, although rRNAs are highly abundant and essential for life.The eukaryotic ribosome is a massive ribonucleoprotein (RNP) complex that consists of four rRNAs and about 80 ribosomal proteins (Venema and Tollervey 1999). The precursor 35S rRNA transcribed by RNA polymerase (Pol) I is processed into three parts; 18S, 5.8S, and 25S rRNA. 5S rRNA is synthesized indep...

Therapeutic mRNAs and vaccines are being developed for a broad range of human diseases, including COVID-19. However, their optimization is hindered by mRNA instability and inefficient protein expression. Here, we describe design principles that overcome these barriers. We develop an RNA sequencing-based platform called PERSIST-seq to systematically delineate in-cell mRNA stability, ribosome load, as well as in-solution stability of a library of diverse mRNAs. We find that, surprisingly, in-cell stability is a greater driver of protein output than high ribosome load. We further introduce a method called In-line-seq, applied to thousands of diverse RNAs, that reveals sequence and structure-based rules for mitigating hydrolytic degradation. Our findings show that highly structured “superfolder” mRNAs can be designed to improve both stability and expression with further enhancement through pseudouridine nucleoside modification. Together, our study demonstrates simultaneous improvement of mRNA stability and protein expression and provides a computational-experimental platform for the enhancement of mRNA medicines.

Most solid-state materials are composed of p-block anions, only in recent years the introduction of hydride anions (1s2) in oxides (e.g., SrVO2H, BaTi(O,H)3) has allowed the discovery of various interesting properties. Here we exploit the large polarizability of hydride anions (H–) together with chalcogenide (Ch2–) anions to construct a family of antiperovskites with soft anionic sublattices. The M3HCh antiperovskites (M = Li, Na) adopt the ideal cubic structure except orthorhombic Na3HS, despite the large variation in sizes of M and Ch. This unconventional robustness of cubic phase mainly originates from the large size-flexibility of the H– anion. Theoretical and experimental studies reveal low migration barriers for Li+/Na+ transport and high ionic conductivity, possibly promoted by a soft phonon mode associated with the rotational motion of HM6 octahedra in their cubic forms. Aliovalent substitution to create vacancies has further enhanced ionic conductivities of this series of antiperovskites, resulting in Na2.9H(Se0.9I0.1) achieving a high conductivity of ~1 × 10–4 S/cm (100 °C).