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Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

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At least 685 records · Page 38
All sourcesNASA NTRSDOE OSTI
DOE OSTI2021

Root angle in maize influences nitrogen capture and is regulated by calcineurin B-like protein (CBL)-interacting serine/threonine-protein kinase 15 ( ZmCIPK15 )

Crops with reduced nutrient and water requirements are urgently needed in global agriculture. Root growth angle plays an important role in nutrient and water acquisition. A maize diversity panel of 481 genotypes was screened for variation in root angle employing a high-throughput field phenotyping platform. Genome-wide association mapping identified several single nucleotide polymorphisms (SNPs) associated with root angle, including one located in the root expressed CBL-interacting serine/threonine-protein kinase 15 (ZmCIPK15) gene (LOC100285495). Reverse genetic studies validated the functional importance of ZmCIPK15, causing a approximately 10° change in root angle in specific nodal positions. A steeper root growth angle improved nitrogen capture in silico and in the field. OpenSimRoot simulations predicted at 40 days of growth that this change in angle would improve nitrogen uptake by 11% and plant biomass by 4% in low nitrogen conditions. In field studies under suboptimal N availability, the cipk15 mutant with steeper growth angles had 18% greater shoot biomass and 29% greater shoot nitrogen accumulation compared to the wild type after 70 days of growth. We propose that a steeper root growth angle modulated by ZmCIPK15 will facilitate efforts to develop new crop varieties with optimal root architecture for improved performance under edaphic stress.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2017

MiR-33a suppresses proliferation of NSCLC cells via targeting METTL3 mRNA

Methyltransferase like 3 (METTL3) was incipiently known as a methyltransferase which was responsible for N{sup 6}-methyladenosine (m{sup 6}A) methylation. METTL3 can promote the expression of several crucial oncoproteins and its high expression enhanced proliferation, survival, and invasion of human lung cancer cells. However, how METTL3 was regulated is seldom understood in non-small-cell lung carcinoma (NSCLC). In the present study, miR-33a was suspicious to target to the 3'-untranslated region (3'UTR) of METTL3 mRNA via in silico prediction. Besides, the expressions of METTL3 were higher in NSCLC tissues than those in adjacent tissues, and METTL3 expressions were positively related to the expressions of miR-33a in NSCLC tissues which confirmed by quantitative real-time polymerase chain reaction (qRT-PCR). MiR-33a can directly target to the 3'UTR of METTL3 mRNA which examined by luciferase reporter gene assay. Moreover, we found that miR-33a can reduce the expression of METTL3 at both mRNA and protein levels using reverse transcription-polymerase chain reaction (RT-PCR) and Western blot analysis. Functionally, miR-33a can reduce the proliferation of A549 and NCI-H460 cells. Conversely, inhibition of miR-33a by anti-miR-33a can rescue that using 4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) and 5-ethynyl-2-deoxyuridine (EdU) assay. Similarly, miR-33a can reduce cellular anchorage-independent growth of A549 cells. Additionally, the negative influences of miR-33a on the downstream genes of METTL3 were examined by Western blot analysis. Thus, we concluded that miR-33a can attenuate NSCLC cells proliferation via targeting to the 3'UTR of METTL3 mRNA. Our findings provide new insights into the mechanism of METTL3 regulation by micro RNA, and supports METTL3 as a therapeutic target in NSCLC. - Highlights: • METTL3 expression is negatively correlated with miR-33a. • MiR-33a can directly target to the 3'UTR of METTL3 mRNA. • MiR-33a inhibits NSCLC cells proliferation through decreasing METTL3. • MiR-33a induces the down-regulation of EGFR through targeting METTL3.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2015

Comparing the cardiovascular therapeutic indices of glycopyrronium and tiotropium in an integrated rat pharmacokinetic, pharmacodynamic and safety model

Long acting inhaled muscarinic receptor antagonists, such as tiotropium, are widely used as bronchodilator therapy for chronic obstructive pulmonary disease (COPD). Although this class of compounds is generally considered to be safe and well tolerated in COPD patients the cardiovascular safety of tiotropium has recently been questioned. We describe a rat in vivo model that allows the concurrent assessment of muscarinic antagonist potency, bronchodilator efficacy and a potential for side effects, and we use this model to compare tiotropium with NVA237 (glycopyrronium bromide), a recently approved inhaled muscarinic antagonist for COPD. Anaesthetized Brown Norway rats were dosed intratracheally at 1 or 6 h prior to receiving increasing doses of intravenous methacholine. Changes in airway resistance and cardiovascular function were recorded and therapeutic indices were calculated against the ED{sub 50} values for the inhibition of methacholine-induced bronchoconstriction. At both time points studied, greater therapeutic indices for hypotension and bradycardia were observed with glycopyrronium (19.5 and 28.5 fold at 1 h; > 200 fold at 6 h) than with tiotropium (1.5 and 4.2 fold at 1 h; 4.6 and 5.5 fold at 6 h). Pharmacokinetic, protein plasma binding and rat muscarinic receptor binding properties for both compounds were determined and used to generate an integrated model of systemic M{sub 2} muscarinic receptor occupancy, which predicted significantly higher M{sub 2} receptor blockade at ED{sub 50} doses with tiotropium than with glycopyrronium. In our preclinical model there was an improved safety profile for glycopyrronium when compared with tiotropium. - Highlights: • We use an in vivo rat model to study CV safety of inhaled muscarinic antagonists. • We integrate protein and receptor binding and PK of tiotropium and glycopyrrolate. • At ED{sub 50} doses for bronchoprotection we model systemic M{sub 2} receptor occupancy. • Glycopyrrolate demonstrates lower M{sub 2} occupancy at bronchoprotective doses. • Glycopyrrolate demonstrates an improved CV safety profile, versus tiotropium.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2025

Phosphoproteomics Modifications in Women with Rheumatoid Arthritis─Application of Web-Based Software to Enhance Data Visualization

Individuals with rheumatoid arthritis (RA) are at increased risk of functional disability, cardiovascular disease, and obesity, all of which are influenced by dysregulated skeletal muscle. Here, this pilot study aims to identify phosphoproteomics changes in RA skeletal muscle and visualize modifications through development of a web-based app designed to promote user-friendly data interpretation and visualization. NanoLC–MS/MS analysis was performed on vastus lateralis biopsies from three women with RA and matched healthy controls. Differential analysis was performed using the Limma R package. Kinase substrate enrichment analysis (KSEA) predicted changes in kinase activity. RA muscle displayed 35 upregulated and 60 downregulated phosphosites, including the cytoskeletal proteins TTN (Ser33201, Ser33013, Ser20925), NEB (Ser2219, Thr254, Ser33013, Ser20925), FLNA (Ser1459), and LASP1 (Ser146). Compared to healthy controls, KSEA predicted decreased activity of several kinases in RA muscle, including PRKACA and CDKs. All such changes were visualized by use of our web-based app. Overall, phosphoproteome analysis reveals signaling alterations in RA skeletal muscle linked to cytoskeletal proteins, representing candidate disease biomarkers; these modifications can be explored through use of our web-based software.

phosphoproteomics↗
NASA NTRS1989

Novel protein crystal growth technology: Proof of concept

A technology for crystal growth, which overcomes certain shortcomings of other techniques, is developed and its applicability to proteins is examined. There were several unknowns to be determined: the design of the apparatus for suspension of crystals of varying (growing) diameter, control of the temperature and supersaturation, the methods for seeding and/or controlling nucleation, the effect on protein solutions of the temperature oscillations arising from the circulation, and the effect of the fluid shear on the suspended crystals. Extensive effort was put forth to grow lysozyme crystals. Under conditions favorable to the growth of tetragonal lysozyme, spontaneous nucleation could be produced but the number of nuclei could not be controlled. Seed transfer techniques were developed and implemented. When conditions for the orthorhombic form were tried, a single crystal 1.5 x 0.5 x 0.2 mm was grown (after in situ nucleation) and successfully extracted. A mathematical model was developed to predict the flow velocity as a function of the geometry and the operating temperatures. The model can also be used to scaleup the apparatus for growing larger crystals of other materials such as water soluble non-linear optical materials. This crystal suspension technology also shows promise for high quality solution growth of optical materials such as TGS and KDP.

Nyce, Thomas A.↗
DOE OSTI2024

Genetic underpinnings of arthropod community distributions in Populus trichocarpa

Community genetics seeks to understand the mechanisms by which natural genetic variation in heritable host phenotypes can encompass assemblages of organisms such as bacteria, fungi, and many animals including arthropods. Prior studies that focused on plant genotypes have been unable to identify genes controlling community composition, a necessary step to predict ecosystem structure and function as underlying genes shift within plant populations. Here, we surveyed arthropods within an association population of Populus trichocarpa in three common gardens to discover plant genes that contributed to arthropod community composition. We analyzed our surveys with traditional single-trait genome-wide association analysis (GWAS), multitrait GWAS, and functional networks built from a diverse set of plant phenotypes. Plant genotype was influential in structuring arthropod community composition among several garden sites. Candidate genes important for higher level organization of arthropod communities had broadly applicable functions, such as terpenoid biosynthesis and production of dsRNA binding proteins and protein kinases, which may be capable of targeting multiple arthropod species. We have demonstrated the ability to detect, in an uncontrolled environment, individual genes that are associated with the community assemblage of arthropods on a host plant, further enhancing our understanding of genetic mechanisms that impact ecosystem structure.

Community genetics↗
DOE OSTI2020

Genome-Wide Analysis of RNA Decay in the Cyanobacterium Synechococcus sp. Strain PCC 7002

ABSTRACT RNA degradation is an important process that influences the ultimate concentration of individual proteins inside cells. While the main enzymes that facilitate this process have been identified, global maps of RNA turnover are available for only a few species. Even in these cases, there are few sequence elements that are known to enhance or destabilize a native transcript; even fewer confer the same effect when added to a heterologous transcript. To address this knowledge gap, we assayed genome-wide RNA degradation in the cyanobacterium Synechococcus sp. strain PCC 7002 by collecting total RNA samples after stopping nascent transcription with rifampin. We quantified the abundance of each position in the transcriptome as a function of time using RNA-sequencing data and later analyzed the global mRNA decay map using machine learning principles. Half-lives, calculated on a per-ORF (open reading frame) basis, were extremely short, with a median half-life of only 0.97 min. Despite extremely rapid turnover of most mRNA, transcripts encoding proteins involved in photosynthesis were both highly expressed and highly stable. Upon inspection of these stable transcripts, we identified an enriched motif in the 3′ untranslated region (UTR) that had similarity to Rho-independent terminators. We built statistical models for half-life prediction and used them to systematically identify sequence motifs in both 5′ and 3′ UTRs that correlate with stabilized transcripts. We found that transcripts linked to a terminator containing a poly(U) tract had a longer half-life than both those without a poly(U) tract and those without a terminator. IMPORTANCE RNA degradation is an important process that affects the final concentration of individual mRNAs, affecting protein expression and cellular physiology. Studies of how RNA is degraded increase our knowledge of this fundamental process as well as enable the creation of genetic tools to manipulate RNA stability. By studying global transcript turnover, we searched for sequence elements that correlated with transcript (in)stability and used these sequences to guide tool design. This study probes global RNA turnover in a cyanobacterium, Synechococcus sp. strain PCC 7002, that both has a unique array of RNases that facilitate RNA degradation and is an industrially relevant strain that could be used to convert CO 2 and sunlight into useful products.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2013

Structural and Histone Binding Ability Characterizations of Human PWWP Domains

The PWWP domain was first identified as a structural motif of 100-130 amino acids in the WHSC1 protein and predicted to be a protein-protein interaction domain. It belongs to the Tudor domain 'Royal Family', which consists of Tudor, chromodomain, MBT and PWWP domains. While Tudor, chromodomain and MBT domains have long been known to bind methylated histones, PWWP was shown to exhibit histone binding ability only until recently. The PWWP domain has been shown to be a DNA binding domain, but sequence analysis and previous structural studies show that the PWWP domain exhibits significant similarity to other 'Royal Family' members, implying that the PWWP domain has the potential to bind histones. In order to further explore the function of the PWWP domain, we used the protein family approach to determine the crystal structures of the PWWP domains from seven different human proteins. Our fluorescence polarization binding studies show that PWWP domains have weak histone binding ability, which is also confirmed by our NMR titration experiments. Furthermore, we determined the crystal structures of the BRPF1 PWWP domain in complex with H3K36me3, and HDGF2 PWWP domain in complex with H3K79me3 and H4K20me3. PWWP proteins constitute a new family of methyl lysine histone binders. The PWWP domain consists of three motifs: a canonical {beta}-barrel core, an insertion motif between the second and third {beta}-strands and a C-terminal {alpha}-helix bundle. Both the canonical {beta}-barrel core and the insertion motif are directly involved in histone binding. The PWWP domain has been previously shown to be a DNA binding domain. Therefore, the PWWP domain exhibits dual functions: binding both DNA and methyllysine histones.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Robust deep learning–based protein sequence design using ProteinMPNN

Although deep learning has revolutionized protein structure prediction, almost all experimentally characterized de novo protein designs have been generated using physically based approaches such as Rosetta. Here, we describe a deep learning–based protein sequence design method, ProteinMPNN, that has outstanding performance in both in silico and experimental tests. On native protein backbones, ProteinMPNN has a sequence recovery of 52.4% compared with 32.9% for Rosetta. The amino acid sequence at different positions can be coupled between single or multiple chains, enabling application to a wide range of current protein design challenges. We demonstrate the broad utility and high accuracy of ProteinMPNN using x-ray crystallography, cryo–electron microscopy, and functional studies by rescuing previously failed designs, which were made using Rosetta or AlphaFold, of protein monomers, cyclic homo-oligomers, tetrahedral nanoparticles, and target-binding proteins.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

De novo transcriptome in roots of switchgrass ( Panicum virgatum L. ) reveals gene expression dynamic and act network under alkaline salt stress

Background: Soil salinization is a major limiting factor for crop cultivation. Switchgrass is a perennial rhizomatous bunchgrass that is considered an ideal plant for marginal lands, including sites with saline soil. Here we investigated the physiological responses and transcriptome changes in the roots of Alamo (alkaline-tolerant genotype) and AM314/MS-155 (alkaline-sensitive genotype) under alkaline salt stress. Results: Alkaline salt stress significantly affected the membrane, osmotic adjustment and antioxidant systems in switchgrass roots, and the ASTTI values between Alamo and AM-314/MS-155 were divergent at different time points. A total of 108,319 unigenes were obtained after reassembly, including 73,636 unigenes in AM-314/MS-155 and 65,492 unigenes in Alamo. A total of 10,219 DEGs were identified, and the number of upregulated genes in Alamo was much greater than that in AM-314/MS-155 in both the early and late stages of alkaline salt stress. The DEGs in AM-314/MS-155 were mainly concentrated in the early stage, while Alamo showed greater advantages in the late stage. These DEGs were mainly enriched in plant-pathogen interactions, ubiquitin-mediated proteolysis and glycolysis/gluconeogenesis pathways. We characterized 1480 TF genes into 64 TF families, and the most abundant TF family was the C2H2 family, followed by the bZIP and bHLH families. A total of 1718 PKs were predicted, including CaMK, CDPK, MAPK and RLK. WGCNA revealed that the DEGs in the blue, brown, dark magenta and light steel blue 1 modules were associated with the physiological changes in roots of switchgrass under alkaline salt stress. The consistency between the qRT-PCR and RNA-Seq results confirmed the reliability of the RNA-seq sequencing data. A molecular regulatory network of the switchgrass response to alkaline salt stress was preliminarily constructed on the basis of transcriptional regulation and functional genes. Conclusions: Alkaline salt tolerance of switchgrass may be achieved by the regulation of ion homeostasis, transport proteins, detoxification, heat shock proteins, dehydration and sugar metabolism. These findings provide a comprehensive analysis of gene expression dynamic and act network induced by alkaline salt stress in two switchgrass genotypes and contribute to the understanding of the alkaline salt tolerance mechanism of switchgrass and the improvement of switchgrass germplasm.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2017

MiR-361 targets Yes-associated protein (YAP) mRNA to suppress cell proliferation in lung cancer

Yes-associated protein (YAP) contributes to the development of multiple tumors, but the post-transcription modulation of YAP remains unexplored. Here, we present a new regulatory microRNA of YAP, miR-361, which directly targets YAP to inhibit cell proliferation in lung cancer. We used bioinformatics to predict that miR-361 could target 3′-untranslated region (3′UTR) of YAP mRNA. Luciferase reporter gene assays demonstrated that miR-361 could decrease the luciferase activities of 3′UTR vector of YAP. Furthermore, YAP expression was obviously abated by miR-361 using RT-PCR and immunoblotting in lung cancer A549 cells. In terms of function, MTT and colony formation analysis showed that ectopic miR-361 expression significantly suppressed cell proliferation in lung cancer. Notably, overexpressed YAP accelerated miR-361-bated proliferation of lung cancer cells. MiR-361 inhibitor promoted cell proliferation in lung cancer, but YAP RNA interference reversed miR-361 inhibitor-driven cell proliferation. Interestingly, miR-361 was capable of affecting the cell cycle in lung cancer progression. Finally, the negative correlation of miR-361 with YAP was found in clinical human lung cancer tissues. In conclusion, miR-361 targets 3′UTR of YAP mRNA to depress the proliferation of lung cancer cells. - Highlights: • MiR-361 is able to target 3′UTR of YAP mRNA. • MiR-361 suppresses the proliferation of lung cancer by inhibiting YAP. • YAP is negatively correlated with miR-361 in lung cancer tissues. • YAP is up-regulated in clinical lung cancer tissues.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

PAG1 promotes the inherent radioresistance of laryngeal cancer cells via activation of STAT3

Highlights: • PAG1 is overexpressed in laryngeal cancer. • PAG1 promotes the inherent radioresistance of laryngeal cancer cells. • PAG1 targets and regulates the expression of p-STAT3. • PAG1-integrin β1 complex is involved in the activation of STAT3. Phosphoprotein associated with glycosphingolipid-enriched microdomains 1(PAG1) is a ubiquitous protein that is essential for the development and progression of various malignancies. A previous study in our laboratory confirmed that PAG1 plays an important role in modulating the inherent radioresistance of laryngeal cancer cells, but the underlying mechanisms are still poorly defined. In this study, we found that PAG1 was significantly increased in laryngeal cancer tissues compared to adjacent non-tumor tissues (P < 0.05). The expression of PAG1 was positively correlated with lymph node metastasis (P < 0.05) and TNM stage (P < 0.05). High expression of PAG1 also predicted a poor prognosis in patients with laryngeal cancer. Moreover, gain-of-function and loss-of-function studies showed that PAG1 overexpression was able to promote growth, increase migration and invasion, and enhance inherent radioresistance of laryngeal cancer cells. Mechanistic investigations revealed that the activation of STAT3 was required for PAG1-mediated inherent radioresistance of laryngeal cancer. Inhibition of STAT3 activity with a chemical inhibitor sensitized radioresistant cells to radiation. Importantly, PAG1-integrin β1 complex was involved in the regulation of STAT3 activation. In addition, downregulation of PAG1 could suppress tumor growth and reverse inherent radioresistance in the nude mouse xenograft model. Taken together, these results suggested that PAG1 conferred inherent radioresistance by activating STAT3, which provided a novel therapeutic strategy for laryngeal cancer.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2012

Bioinformatics resource manager v2.3: an integrated software environment for systems biology with microRNA and cross-species analysis tools

Background: MicroRNAs (miRNAs) are noncoding RNAs that direct post-transcriptional regulation of protein coding genes. Recent studies have shown miRNAs are important for controlling many biological processes, including nervous system development, and are highly conserved across species. Given their importance, computational tools are necessary for analysis, interpretation and integration of high-throughput (HTP) miRNA data in an increasing number of model species. The Bioinformatics Resource Manager (BRM) v2.3 is a software environment for data management, mining, integration and functional annotation of HTP biological data. In this study, we report recent updates to BRM for miRNA data analysis and cross-species comparisons across datasets. Results: BRM v2.3 has the capability to query predicted miRNA targets from multiple databases, retrieve potential regulatory miRNAs for known genes, integrate experimentally derived miRNA and mRNA datasets, perform ortholog mapping across species, and retrieve annotation and cross-reference identifiers for an expanded number of species. Here we use BRM to show that developmental exposure of zebrafish to 30 uM nicotine from 6–48 hours post fertilization (hpf) results in behavioral hyperactivity in larval zebrafish and alteration of putative miRNA gene targets in whole embryos at developmental stages that encompass early neurogenesis. We show typical workflows for using BRM to integrate experimental zebrafish miRNA and mRNA microarray datasets with example retrievals for zebrafish, including pathway annotation and mapping to human ortholog. Functional analysis of differentially regulated (p<0.05) gene targets in BRM indicates that nicotine exposure disrupts genes involved in neurogenesis, possibly through misregulation of nicotine-sensitive miRNAs. Conclusions: BRM provides the ability to mine complex data for identification of candidate miRNAs or pathways that drive phenotypic outcome and, therefore, is a useful hypothesis generation tool for systems biology. The miRNA workflow in BRM allows for efficient processing of multiple miRNA and mRNA datasets in a single software environment with the added capability to interact with public data sources and visual analytic tools for HTP data analysis at a systems level. BRM is developed using Java™ and other open-source technologies for free distribution (http://www.sysbio.org/dataresources/brm.stm).

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2014

The RNA-binding protein PCBP2 facilitates gastric carcinoma growth by targeting miR-34a

Highlights: • PCBP2 is overexpressed in human gastric cancer. • PCBP2 high expression predicts poor survival. • PCBP2 regulates gastric cancer growth in vitro and in vivo. • PCBP2 regulates gastric cancer apoptosis by targeting miR-34a. - Abstract: Gastric carcinoma is the fourth most common cancer worldwide, with a high rate of death and low 5-year survival rate. However, the mechanism underling gastric cancer is still not fully understood. Here in the present study, we identify the RNA-binding protein PCBP2 as an oncogenic protein in human gastric carcinoma. Our results show that PCBP2 is up-regulated in human gastric cancer tissues compared to adjacent normal tissues, and that high level of PCBP2 predicts poor overall and disease-free survival. Knockdown of PCBP2 in gastric cancer cells inhibits cell proliferation and colony formation in vitro, whereas opposing results are obtained when PCBP2 is overexpressed. Our in vivo subcutaneous xenograft results also show that PCBP2 can critically regulate gastric cancer cell growth. In addition, we find that PCBP2-depletion induces apoptosis in gastric cancer cells via up-regulating expression of pro-apoptotic proteins and down-regulating anti-apoptotic proteins. Mechanically, we identify that miR-34a as a target of PCBP2, and that miR-34a is critically essential for the function of PCBP2. In summary, PCBP2 promotes gastric carcinoma development by regulating the level of miR-34a.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2023

CasDinG is a 5′-3′ dsDNA and RNA/DNA helicase with three accessory domains essential for type IV CRISPR immunity

Abstract CRISPR-associated DinG protein (CasDinG) is essential to type IV-A CRISPR function. Here, we demonstrate that CasDinG from Pseudomonas aeruginosa strain 83 is an ATP-dependent 5′-3′ DNA translocase that unwinds double-stranded (ds)DNA and RNA/DNA hybrids. The crystal structure of CasDinG reveals a superfamily 2 helicase core of two RecA-like domains with three accessory domains (N-terminal, arch, and vestigial FeS). To examine the in vivo function of these domains, we identified the preferred PAM sequence for the type IV-A system (5′-GNAWN-3′ on the 5′-side of the target) with a plasmid library and performed plasmid clearance assays with domain deletion mutants. Plasmid clearance assays demonstrated that all three domains are essential for type IV-A immunity. Protein expression and biochemical assays suggested the vFeS domain is needed for protein stability and the arch for helicase activity. However, deletion of the N-terminal domain did not impair ATPase, ssDNA binding, or helicase activities, indicating a role distinct from canonical helicase activities that structure prediction tools suggest involves interaction with dsDNA. This work demonstrates CasDinG helicase activity is essential for type IV-A CRISPR immunity as well as the yet undetermined activity of the CasDinG N-terminal domain.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Knocking out the carboxyltransferase interactor 1 (CTI1) in Chlamydomonas boosted oil content by fivefold without affecting cell growth

Summary The first step in chloroplast de novo fatty acid synthesis is catalysed by acetyl‐CoA carboxylase (ACCase). As the rate‐limiting step for this pathway, ACCase is subject to both positive and negative regulation. In this study, we identify a Chlamydomonas homologue of the plant carboxyltransferase interactor 1 (CrCTI1) and show that this protein interacts with the Chlamydomonas α‐carboxyltransferase (Crα‐CT) subunit of the ACCase by yeast two‐hybrid protein–protein interaction assay. Three independent CRISPR‐Cas9 mediated knockout mutants for CrCTI1 each produced an ‘enhanced oil’ phenotype, accumulating 25% more total fatty acids and storing up to fivefold more triacylglycerols (TAGs) in lipid droplets. The TAG phenotype of the crcti1 mutants was not influenced by light but was affected by trophic growth conditions. By growing cells under heterotrophic conditions, we observed a crucial function of CrCTI1 in balancing lipid accumulation and cell growth. Mutating a previously mapped in vivo phosphorylation site (CrCTI1 Ser108 to either Ala or to Asp), did not affect the interaction with Crα‐CT. However, mutating all six predicted phosphorylation sites within Crα‐CT to create a phosphomimetic mutant reduced this pairwise interaction significantly. Comparative proteomic analyses of the crcti1 mutants and WT suggested a role for CrCTI1 in regulating carbon flux by coordinating carbon metabolism, antioxidant and fatty acid β‐oxidation pathways, to enable cells to adapt to carbon availability. Taken together, this study identifies CrCTI1 as a negative regulator of fatty acid synthesis in algae and provides a new molecular brick for the genetic engineering of microalgae for biotechnology purposes.

Li, Zhongze [Aix‐Marseille Université, CEA, CNRS, ↗
NASA NTRS2016

In-Flight Personalized Medication Management

Current medication selection for treatment of astronauts during spaceflight missions is primarily dictated by the task of efficiently treating the widest possible range of physiological conditions and illnesses with a limited set of medications. Dosage and recommendations on the combination of drugs are based on the assumption of genetically equal drug sensitivity and unchanged metabolism. To our knowledge, there was no pre-flight drug sensitivity testing on a genetic level for any of the previous manned NASA space missions. Although many of the common, binary drug-drug interactions are, most likely, already considered in the ISS Medical kit composition, multi-drug and multi-drug-gene factors are not incorporated in the medication selection or prescription. Furthermore, due to the physiological changes occurring in microgravity environments, astronauts might be susceptible to potential increased drug toxicity as a result of decreased clearance of numerous drugs. In particular, perturbation of CYP450 enzymes which contribute to the hepatic metabolism of the majority of drugs may have significant effects on therapeutic efficacy and increase treatment-related toxicity5. The genes encoding the CYP450 enzymes are highly variable in humans. Inheritable variations of CYP450 hepatic metabolizer enzymes and transport proteins play a crucial role in the inter-individual variability of drug efficiency and risks of adverse drug reactions5. Additionally, there are some reports that document changes in the levels of production of drug-metabolizing enzymes in microgravity. These data can be extrapolated to provide reasonable assumptions of decreased levels of expression for most CYP450 enzymes in human body during prolonged space travel. If the prescribed medication regiment is not fully effective or causes undesirable side effects, the ability of the astronauts to function and maintain peak performance levels during space flight could be seriously compromised. Therefore, technologies capable of predicting and managing medication side effects, interactions, and toxicity of drugs during spaceflight are needed. We propose to develop and customize for NASAs applications available on the market Personalized Prescribing System (PPS) that would provide a comprehensive, non-invasive solution for safer, targeted medication management for every crew member resulting in safer and more effective treatment and, consequently, better performance. PPS will function as both decision support and record-keeping tool for flight surgeons and astronauts in applying the recommended medications for situations arising in flight. The information on individual drug sensitivity will translate into personalized risk assessment for adverse drug reactions and treatment failures for each drug from the medication kit as well as predefined outcome of any combination of them. Dosage recommendations will also be made individually. The mobile app will facilitate ease of use by crew and medical professionals during training and flight missions.

Personalized Medication↗
DOE OSTI2023

Predicting Mössbauer Parameters of Nonheme Diiron Complexes with Density Functional Theory

Mössbauer spectroscopy provides significant insights into the electronic structure and environment of the metal centers. In this work, we investigate the electronic structures of a set of nonheme diiron complexes by evaluating two key parameters pertaining to Mössbauer spectroscopy, namely, the isomer shift (δ) and quadrupole splitting (|ΔE Q |), using different levels of density functional theory (DFT). The diiron systems investigated here span diverse oxidation states, bridging motifs, and spin coupling patterns, which present a challenging case for theoretical predictions. We demonstrate that the combination of B97-D3/def2-TZVP is an efficient approach in modeling both the δ and |ΔEQ| values with high accuracy for the representative nonheme diiron complexes. We also show that δ is accurately predicted irrespective of the choice of approximate density functional while the |ΔE Q | is sensitive to the level of theory employed. Further investigation shows that the present methodology assessed using synthetic nonheme diiron complexes could be extended to nonheme diiron enzyme active sites, featuring both ferromagnetic and antiferromagnetic coupling between the iron centers.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗