Pyruvate Kinase M (PKM) binds ribosomes in a poly-ADP ribosylation dependent manner to induce translational stalling.

In light of the numerous studies identifying post-transcriptional regulators on the surface of the endoplasmic reticulum (ER), we asked whether there are factors that regulate compartment specific mRNA translation in human cells. Using a proteomic survey of spatially regulated polysome interacting proteins, we identified the glycolytic enzyme Pyruvate Kinase M (PKM) as a cytosolic (i.e. ER-excluded) polysome interactor and investigated how it influences mRNA translation. We discovered that the PKM-polysome interaction is directly regulated by ADP levels-providing a link between carbohydrate metabolism and mRNA translation. By performing enhanced crosslinking immunoprecipitation-sequencing (eCLIP-seq), we found that PKM crosslinks to mRNA sequences that are immediately downstream of regions that encode lysine- and glutamate-enriched tracts. Using ribosome footprint protection sequencing, we found that PKM binding to ribosomes causes translational stalling near lysine and glutamate encoding sequences. Lastly, we observed that PKM recruitment to polysomes is dependent on poly-ADP ribosylation activity (PARylation)-and may depend on co-translational PARylation of lysine and glutamate residues of nascent polypeptide chains. Overall, our study uncovers a novel role for PKM in post-transcriptional gene regulation, linking cellular metabolism and mRNA translation.

SARS-CoV-2 targets ribosomal RNA biogenesis.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) hinders host gene expression, curbing defenses and licensing viral protein synthesis and virulence. During SARS-CoV-2 infection, the virulence factor non-structural protein 1 (Nsp1) targets the mRNA entry channel of mature cytoplasmic ribosomes, limiting translation. We show that Nsp1 also restrains translation by targeting nucleolar ribosome biogenesis. SARS-CoV-2 infection disrupts 18S and 28S ribosomal RNA (rRNA) processing. Expression of Nsp1 recapitulates the processing defects. Nsp1 abrogates rRNA production without altering the expression of critical processing factors or nucleolar organization. Instead, Nsp1 localizes to the nucleolus, interacting with precursor-rRNA and hindering its maturation separately from the viral protein's role in restricting mature ribosomes. Thus, SARS-CoV-2 Nsp1 limits translation by targeting ribosome biogenesis and mature ribosomes. These findings revise our understanding of how SARS-CoV-2 Nsp1 controls human protein synthesis, suggesting that efforts to counter Nsp1's effect on translation should consider the protein's impact from ribosome manufacturing to mature ribosomes.

Non-Darwinian Molecular Biology.

With the discovery of the double helical structure of DNA, a shift occurred in how biologists investigated questions surrounding cellular processes, such as protein synthesis. Instead of viewing biological activity through the lens of chemical reactions, this new field used biological information to gain a new profound view of how biological systems work. Molecular biologists asked new types of questions that would have been inconceivable to the older generation of researchers, such as how cellular machineries convert inherited biological information into functional molecules like proteins. This new focus on biological information also gave molecular biologists a way to link their findings to concepts developed by genetics and the modern synthesis. However, by the late 1960s this all changed. Elevated rates of mutation, unsustainable genetic loads, and high levels of variation in populations, challenged Darwinian evolution, a central tenant of the modern synthesis, where adaptation was the main driver of evolutionary change. Building on these findings, Motoo Kimura advanced the neutral theory of molecular evolution, which advocates that selection in multicellular eukaryotes is weak and that most genomic changes are neutral and due to random drift. This was further elaborated by Jack King and Thomas Jukes, in their paper "Non-Darwinian Evolution", where they pointed out that the observed changes seen in proteins and the types of polymorphisms observed in populations only become understandable when we take into account biochemistry and Kimura's new theory. Fifty years later, most molecular biologists remain unaware of these fundamental advances. Their adaptionist viewpoint fails to explain data collected from new powerful technologies which can detect exceedingly rare biochemical events. For example, high throughput sequencing routinely detects RNA transcripts being produced from almost the entire genome yet are present less than one copy per thousand cells and appear to lack any function. Molecular biologists must now reincorporate ideas from classical biochemistry and absorb modern concepts from molecular evolution, to craft a new lens through which they can evaluate the functionality of transcriptional units, and make sense of our messy, intricate, and complicated genome.

mRNA localization as a rheostat to regulate subcellular gene expression.

It is currently believed that certain messenger RNAs (mRNAs) are localized to distinct subcellular regions to efficiently target their encoded proteins. However, this simplistic model does not explain why in certain scenarios mRNA localization is dispensable for proper protein distribution. In other cases, mRNA localization is accompanied by translational silencing and degradation by the localization machinery. Here we propose that in certain scenarios mRNAs are localized so that they can either be stabilized and translated, or silenced and degraded, in response to the needs of the subcellular locale. In these cases, the localized mRNA, and its cadre of associated factors, act as a rheostat that regulates protein production and/or mRNA stability in response to the needs of its immediate subcellular environment. WIREs RNA 2017, 8:e1416. doi: 10.1002/wrna.1416 For further resources related to this article, please visit the WIREs website.