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Tissue ischemia results in intracellular pH (pHIN) acidification, and while accumulation of metabolites such as lactate is a known driver of acidic pHIN, less is known about how acidic pHIN regulates metabolism. Furthermore, acidic extracellular (pHEX) during early reperfusion confers cardioprotection, but how this impacts metabolism is unclear. Herein we employed LCMS based targeted metabolomics to analyze perfused mouse hearts exposed to: (i) control perfusion, (ii) hypoxia, (iii) ischemia, (iv) enforced acidic pHIN, (v) control reperfusion, and (vi) acidic pHEX (6.8) reperfusion. Surprisingly little overlap was seen between metabolic changes induced by hypoxia, ischemia, and acidic pHIN. Acidic pHIN elevated metabolites in the top half of glycolysis, and enhanced glutathione redox state. Acidic pHEX reperfusion induced substantial metabolic changes in addition to those seen in control reperfusion. This included elevated metabolites in the top half of glycolysis, prevention of purine nucleotide loss, and an enhancement in glutathione redox state. These data led to parallel hypotheses regarding potential roles for methylglyoxal inhibiting the mitochondrial permeability transition pore, and for acidic inhibition of ecto-5’-nucleotidase, as potential mediators of cardioprotection by acidic pHEX reperfusion. However, neither hypothesis was supported by subsequent experiments. In contrast, analysis of cardiac effluents revealed complex effects of pHEX on metabolite transport, suggesting that mildly acidic pHEX may protect in part by enhancing succinate release during reperfusion. Overall, each intervention had distinct and overlapping metabolic effects, suggesting acidic pH is an independent metabolic regulator regardless which side of the cell membrane it is imposed.HIGHLIGHTSHypoxia, ischemia and acidic pHIN each induce unique cardiac metabolic profiles.Acidic pHEX at reperfusion prevents purine loss and enhances succinate release.

Nonstructural protein 5 (Nsp5) is the main protease of SARS-CoV-2 that cleaves viral polyproteins into individual polypeptides necessary for viral replication. Here, we show that Nsp5 binds and cleaves human tRNA methyltransferase 1 (TRMT1), a host enzyme required for a common post-transcriptional modification in tRNAs. Human cells infected with SARS-CoV-2 exhibit a decrease in TRMT1 protein levels and TRMT1-catalyzed tRNA modifications, consistent with TRMT1 cleavage and inactivation by Nsp5. Nsp5 cleaves TRMT1 at a specific position that matches the consensus sequence of SARS-CoV-2 polyprotein cleavage sites, and a single mutation within the sequence inhibits Nsp5-dependent proteolysis of TRMT1. The TRMT1 cleavage fragments exhibit altered RNA binding activity and are unable to rescue tRNA modification in TRMT1-deficient human cells. Compared to wildtype human cells, TRMT1-deficient human cells infected with SARS-CoV-2 exhibit reduced levels of intracellular viral RNA. These findings suggest that Nsp5-dependent cleavage of TRMT1 and perturbation of tRNA modification patterns contribute to the cellular pathogenesis and biology of SARS-CoV-2 infection.

Poly-[(

Ribosomal frameshifting, a process whereby a translating ribosome is diverted from one reading frame to another on a contiguous mRNA, is an important regulatory mechanism in biology and an opportunity for therapeutic intervention in several human diseases. In HIV, ribosomal frameshifting controls the ratio of Gag and Gag-Pol, two polyproteins critical to the HIV life cycle. We have previously reported compounds able to selectively bind an RNA stemloop within the Gag-Pol mRNA; these compounds alter the production of Gag-Pol in a manner consistent with increased frameshifting. Importantly, they also display antiretroviral activity in human T-cells. Here, we describe new compounds with significantly reduced molecular weight, but with substantially maintained affinity and anti-HIV activity. These results suggest that development of more “ligand efficient” enhancers of ribosomal frameshifting is an achievable goal.

Poly-[(R)-3-hydroxyalkanoate] biopolymers, or PHAs, are biocompatible and biodegradable polyesters that can be produced by diverse microbial strains. PHA polymers have found widespread uses in applications ranging from sustainable replacements of nonbiodegradable bulk-commodity plastics to biomaterials. However, further expansion into other markets and industries has generally been limited by the inability to chemically modify these polymers. Recently, our lab engineered E. coli LSBJ, a microbial strain able to produce PHA copolymers with controlled unit compositions from simple and accessible fatty acid feedstocks. We envisioned meaningfully broadening the application spectrum of these materials via production of chemically tractable PHA biopolymers containing “click”-ready chemical functionalities. With a myriad of applications in mind, in this study we demonstrate the synthesis and biopolymerization of a panel of ω-azido fatty acids and take the first exploratory steps toward demonstrating their conjug...

This study assessed the nutrient acquisition strategies of two scleractinian corals, Montastraea cavernosa and Agaricia spp., collected from shallow (depths less than 20 m) and mesophotic (depths of 30–150 m) habitats. The composition of biomarker sterols, bulk stable carbon and nitrogen isotope values, and compound specific stable isotope analysis (CSIA) of the sterols were analyzed to assess changes in feeding strategies of the corals. Both species acquired nutrients by heterotrophic feeding and translocation from symbionts. Colonies of M. cavernosa acquired photosynthetic nutrients in shallow and mesophotic habitats, and the relative sterol (and phytosterol) composition did not change with depth. CSIA evidence suggests that photosynthesis slowed with increasing depth. Colonies of Agaricia spp. used heterotrophic feeding throughout their depth range and acquired some photosynthetic nutrients in shallow habitats and few in mesophotic habitats. Both corals, Agaricia spp. and M. cavernosa, may be able to take advantage of deep reef refugia to maintain populations in a changing ocean by using distinct nutrient acquisition strategies.