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At least 37 records · Page 2
All sourcesNASA NTRSDOE OSTI
DOE OSTI2024

Activation of CBASS Cap5 endonuclease immune effector by cyclic nucleotides

The bacterial cyclic oligonucleotide-based antiphage signaling system (CBASS) is similar to the cGAS–STING system in humans, containing an enzyme that synthesizes a cyclic nucleotide on viral infection and an effector that senses the second messenger for the antiviral response. Cap5, containing a SAVED domain coupled to an HNH DNA endonuclease domain, is the most abundant CBASS effector, yet the mechanism by which it becomes activated for cell killing remains unknown. We present here high-resolution structures of full-length Cap5 from Pseudomonas syringae (Ps) with second messengers. Here, the key to PsCap5 activation is a dimer-to-tetramer transition, whereby the binding of second messenger to dimer triggers an open-to-closed transformation of the SAVED domains, furnishing a surface for assembly of the tetramer. This movement propagates to the HNH domains, juxtaposing and converting two HNH domains into states for DNA destruction. These results show how Cap5 effects bacterial cell suicide and we provide proof-in-principle data that the CBASS can be extrinsically activated to limit bacterial infections.

36 MATERIALS SCIENCE↗
DOE OSTI2021

Vascular K ATP channel structural dynamics reveal regulatory mechanism by Mg-nucleotides

Vascular tone is dependent on smooth muscle K ATP channels comprising pore-forming Kir6.1 and regulatory SUR2B subunits, in which mutations cause Cantú syndrome. Unique among K ATP isoforms, they lack spontaneous activity and require Mg-nucleotides for activation. Structural mechanisms underlying these properties are unknown. Here, we determined cryogenic electron microscopy structures of vascular K ATP channels bound to inhibitory ATP and glibenclamide, which differ informatively from similarly determined pancreatic K ATP channel isoform (Kir6.2/SUR1). Unlike SUR1, SUR2B subunits adopt distinct rotational “propeller” and “quatrefoil” geometries surrounding their Kir6.1 core. The glutamate/aspartate-rich linker connecting the two halves of the SUR-ABC core is observed in a quatrefoil-like conformation. Molecular dynamics simulations reveal MgADP-dependent dynamic tripartite interactions between this linker, SUR2B, and Kir6.1. The structures captured implicate a progression of intermediate states between MgADP-free inactivated, and MgADP-bound activated conformations wherein the glutamate/aspartate-rich linker participates as mobile autoinhibitory domain, suggesting a conformational pathway toward K ATP channel activation.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Codon bias, nucleotide selection, and genome size predict in situ bacterial growth rate and transcription in rewetted soil

In soils, the first rain after a prolonged dry period represents a major pulse event impacting soil microbial community function, yet we lack a full understanding of the genomic traits associated with the microbial response to rewetting. Genomic traits such as codon usage bias and genome size have been linked to bacterial growth in soils—however, often through measurements in culture. Here, we used metagenome-assembled genomes (MAGs) with 18 O-water stable isotope probing and metatranscriptomics to track genomic traits associated with growth and transcription of soil microorganisms over one week following rewetting of a grassland soil. We found that codon bias in ribosomal protein genes was the strongest predictor of growth rate. We also found higher growth rates in bacteria with smaller genomes, suggesting that reduced genome size enables a faster response to pulses in soil bacteria. Faster transcriptional upregulation of ribosomal protein genes was associated with high codon bias and increased nucleotide skew. We found that several of these relationships existed within phyla, indicating that these associations between genomic traits and activity could be generalized characteristics of soil bacteria. Finally, we used publicly available metagenomes to assess the distribution of codon bias across a pH gradient and found that microbial communities in higher pH soils—which are often more water limited and pulse driven—have higher codon usage bias in their ribosomal protein genes. Together, these results provide evidence that genomic characteristics affect soil microbial activity during rewetting and pose a potential fitness advantage for soil bacteria where water and nutrient availability are episodic.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

In vitro evidence that the vasorelaxant effects of 2‐nitro‐1‐phenyl‐1‐propanol on rat coronary arteries involve cyclic nucleotide pathways

Abstract The synthetic nitro‐alcohol 2‐nitro‐1‐phenyl‐1‐propanol (NPP) has endothelium‐independent relaxing properties in isolated preparations of rat aorta and mesenteric artery. In this study, we investigated whether the vasodilator effects occur in coronary vessels and explored whether hyperpolarization is involved in the underlying mechanism of NPP‐induced smooth muscle relaxation. The relaxing responses were studied in isolated preparations of the left anterior descending coronary (ADC) and the septal coronary (SC) arteries, which had been previously maintained under sustained contraction induced by the thromboxane A 2 analogue U‐46619. Administered cumulatively, NPP elicited concentration‐dependent vasorelaxation with similar potency in both vessels. The relaxant effect remained unaffected by the nitric oxide synthase inhibitor L‐NAME, the protein kinase C inhibitor bisindolylmaleimide IV and the Rho‐associated protein kinase inhibitor Y‐27632. However, it was significantly diminished by the adenylyl cyclase inhibitor MDL‐12,330A, the guanylyl cyclase inhibitor ODQ, as well as the K + channel inhibitors tetraethylammonium and CsCl. In ADC preparations impaled with intracellular micropipettes, NPP hyperpolarized the vascular preparation. When the isolated preparation was precontracted by 5‐hydroxytryptamine or 80 mM KCl, NPP‐induced relaxation with lower pharmacological potency compared to the vessels contracted by U‐46619. In conclusion, NPP exhibits vasorelaxant effects on rat coronary arteries, likely involving pathways that include cyclic nucleotide production and membrane hyperpolarization.

Vasconcelos‐Silva, Alfredo Augusto↗
DOE OSTI2022

A noncoding single-nucleotide polymorphism at 8q24 drives IDH1 -mutant glioma formation

Establishing causal links between inherited polymorphisms and cancer risk is challenging. Here, we focus on the single-nucleotide polymorphism rs55705857, which confers a sixfold greater risk of isocitrate dehydrogenase (IDH)–mutant low-grade glioma (LGG). Here, we reveal that rs55705857 itself is the causal variant and is associated with molecular pathways that drive LGG. Mechanistically, we show that rs55705857 resides within a brain-specific enhancer, where the risk allele disrupts OCT2/4 binding, allowing increased interaction with the Myc promoter and increased Myc expression. Mutating the orthologous mouse rs55705857 locus accelerated tumor development in an Idh1 R132H -driven LGG mouse model from 472 to 172 days and increased penetrance from 30% to 75%. Our work reveals mechanisms of the heritable predisposition to lethal glioma in ~40% of LGG patients.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Using intrahost single nucleotide variant data to predict SARS-CoV-2 detection cycle threshold values

Over the last four years, each successive wave of the COVID-19 pandemic has been caused by variants with mutations that improve the transmissibility of the virus. Despite this, we still lack tools for predicting clinically important features of the virus. In this study, we show that it is possible to predict the PCR cycle threshold (Ct) values from clinical detection assays using sequence data. Ct values often correspond with patient viral load and the epidemiological trajectory of the pandemic. Using a collection of 36,335 high quality genomes, we built models from SARS-CoV-2 intrahost single nucleotide variant (iSNV) data, computing XGBoost models from the frequencies of A, T, G, C, insertions, and deletions at each position relative to the Wuhan-Hu-1 reference genome. Our best model had an R 2 of 0.604 [0.593–0.616, 95% confidence interval] and a Root Mean Square Error (RMSE) of 5.247 [5.156–5.337], demonstrating modest predictive power. Overall, we show that the results are stable relative to an external holdout set of genomes selected from SRA and are robust to patient status and the detection instruments that were used. This study highlights the importance of developing modeling strategies that can be applied to publicly available genome sequence data for use in disease prevention and control.

COVID19↗
DOE OSTI2024

An empirical DNA-based identification of morphologically similar snappers (Lutjanus campechanus, Lutjanus purpureus) using a versatile bioinformatics workflow for the discovery and analysis of informative single-nucleotide polymorphisms

The commercially important speciesLutjanus campechanus(Northern/Gulf red snapper) andLutjanus purpureus(Southern/Caribbean red snapper) are the protagonists of a decade’s long taxonomic debate over their species delimitation, due in part to partial habitat overlap, extensive morphological similarity, and the lack of resolution when applying canonically reliable DNA barcoding approaches. In this study, we leveraged publicly available RAD-Seq data forL. campechanusandL. purpureusto identify species-informative single‐nucleotide polymorphisms (SNPs) at the genome scale that were successful in distinguishing the Northern and Southern red snappers, while also detecting individuals exhibiting introgression. This 4-step empirical approach demonstrates the value of applying novel bioinformatics pipelines to existing genome-scale data to maximize the distillation of informative subsets. Our results facilitate economically relevant species identification in addition to confirming or challenging species identifications for specimens with data in public databases. These findings and their applications will benefit future sustainability strategies and broader research questions surrounding these overfished and evolutionarily entangled snapper species.

Environmental Sciences & Ecology↗
DOE OSTI2022

Population Structure of White Sturgeon (Acipenser transmontanus) in the Columbia River Inferred from Single-Nucleotide Polymorphisms

White sturgeon (Acipenser transmontanus) are the largest freshwater fish in North America, with reproducing populations in the Sacramento-San Joaquin, Fraser, and Columbia River Basins. Of these, the Columbia River is the largest, but it is also highly fragmented by hydroelectric dams, and many segments are characterized by declining abundance and persistent recruitment failure. Efforts to conserve and supplement these fish requires an understanding of their spatial genetic structure. Here, we assembled a large set of samples from throughout the Columbia River Basin, along with representative collections from adjacent basins, and genotyped them using a panel of 325 single-nucleotide markers. Results from individual- and group-based analyses of these data indicate that white sturgeon in the uppermost Columbia River Basin, in the Kootenai and upper Snake Rivers, are the most distinct, while the remaining populations downstream in the basin can be described as a genetic gradient consistent with an isolation-by-distance effect. Notably, the population in the lowest reaches of the Columbia River is more distinct from the middle or upper reaches than from outside basins, and suggests historically a higher or more recent gene exchange through coastal routes than with populations in the interior Columbia Basin. Nonetheless, proximal reaches were generally only marginally or non-significantly divergent, suggesting that transplanting larvae or juveniles from nearby sources poses relatively little risk of outbreeding depression. Indeed, we inferred examples of dispersal between reaches via close-kin mark-recapture and genetic mark-recapture that indicate movement between nearby reaches is not unusual. Samples from the Kootenai and upper Snake Rivers exhibited notably lower genetic diversity than the remaining samples as a result of population bottlenecks, genetic drift, and/or historical divergence. Conservation actions, such as supplementation, are underway to maintain population viability and will require balanced efforts to increase demographic abundance while maintaining genetic diversity.

Willis, Stuart C.↗
DOE OSTI2020

Viral inhibitory nucleotide sequences and vaccines

The invention relates to inhibitory nucleotide signal sequences or “INS” sequences in the genomes of lentiviruses. In particular the invention relates to the AGG motif present in all viral genomes. The AGG motif may have an inhibitory effect on a virus, for example by reducing the levels of, or maintaining low steady-state levels of, viral RNAs in host cells, and inducing and/or maintaining in viral latency. In one aspect, the invention provides vaccines that contain, or are produced from, viral nucleic acids in which the AGG sequences have been mutated. In another aspect, the invention provides methods and compositions for affecting the function of the AGG motif, and methods for identifying other INS sequences in viral genomes.

Rabadan, Raul↗
DOE OSTI2023

The mitochondrially targeted peptide elamipretide (SS-31) improves ADP sensitivity in aged mitochondria by increasing uptake through the adenine nucleotide translocator (ANT)

Aging muscle experiences functional decline in part mediated by impaired mitochondrial ADP sensitivity. Elamipretide (ELAM) rapidly improves physiological and mitochondrial function in aging and binds directly to the mitochondrial ADP transporter ANT. We hypothesized that ELAM improves ADP sensitivity in aging leading to rescued physiological function. We measured the response to ADP stimulation in young and old muscle mitochondria with ELAM treatment, in vivo heart and muscle function, and compared protein abundance, phosphorylation, and S-glutathionylation of ADP/ATP pathway proteins. ELAM treatment increased ADP sensitivity in old muscle mitochondria by increasing uptake of ADP through the ANT and rescued muscle force and heart systolic function. Protein abundance in the ADP/ATP transport and synthesis pathway was unchanged, but ELAM treatment decreased protein s-glutathionylation incuding of ANT. Mitochondrial ADP sensitivity is rapidly modifiable. This research supports the hypothesis that ELAM improves ANT function in aging and links mitochondrial ADP sensitivity to physiological function.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2021

Pharmacological Inhibition of HSP90 Radiosensitizes Head and Neck Squamous Cell Carcinoma Xenograft by Inhibition of DNA Damage Repair, Nucleotide Metabolism, and Radiation-Induced Tumor Vasculogenesis

Recent preclinical studies suggest combining the HSP90 inhibitor AT13387 (Onalespib) with radiation (IR) against colon cancer and head and neck squamous cell carcinoma (HNSCC). These studies emphasized that AT13387 downregulates HSP90 client proteins involved in oncogenic signaling and DNA repair mechanisms as major drivers of enhanced radiosensitivity. Given the large array of client proteins HSP90 directs, we hypothesized that other key proteins or signaling pathways may be inhibited by AT13387 and contribute to enhanced radiosensitivity. Metabolomic analysis of HSP90 inhibition by AT13387 was conducted to identify metabolic biomarkers of radiosensitization and whether modulations of key proteins were involved in IR-induced tumor vasculogenesis, a process involved in tumor recurrence.

62 RADIOLOGY AND NUCLEAR MEDICINE↗