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DOE OSTI2025

A network-enabled pipeline for gene discovery and validation in non-model plant species

Identifying key regulators of important genes in non-model crop species is challenging due to limited multi-omics resources. To address this, we introduce the network-enabled gene discovery pipeline NEEDLE, a user-friendly tool that systematically generates coexpression gene network modules, measures gene connectivity, and establishes network hierarchy to pinpoint key transcriptional regulators from dynamic transcriptome datasets. After validating its accuracy with two independent datasets, we applied NEEDLE to identify transcription factors (TFs) regulating the expression of cellulose synthase-like F6 ( CSLF6 ), a crucial cell wall biosynthetic gene, in Brachypodium and sorghum. Our analyses uncover regulators of CSLF6 and also shed light on the evolutionary conservation or divergence of gene regulatory elements among grass species. These results highlight NEEDLE’s capability to provide biologically relevant TF predictions and demonstrate its value for non-model plant species with dynamic transcriptome datasets.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2022

Case study on the novel permitting and authorization of PacWave South, a US grid-connected wave energy test facility: Development, challenges, and insights

Marine energy (i.e., energy from waves, tides, currents, and rivers) in the United States is a nascent industry. In particular, permitting processes—an uncertainty for industry advancement that can be costly and time consuming to navigate—have rarely been tested and used for marine energy. The novelty of the marine energy industry and utilization of open ocean permitting processes that were not originally developed for marine energy have led to extensive efforts to gain consensus amongst state and federal regulatory agencies to authorize marine energy projects. In 2021, Oregon State University successfully completed permitting of a wave energy test facility, called PacWave South, off the coast of Oregon, which is designed to advance wave energy research and development. This article documents the multi-year process that Oregon State University used to receive federal and state authorization for a pre-permitted commercial-scale grid connected facility by detailing the development of the test facility, management of uncertainty and challenges, and key decisions. The PacWave South case study provides insights for the larger marine energy community as the industry advances towards commercialization.

16 TIDAL AND WAVE POWER↗

DOE OSTI2024

Bridging the Energy Future: The Role and Potential of Hydrogen Co-Firing with Natural Gas

In the transition toward sustainable energy systems, the co-firing of hydrogen (H2) with natural gas emerges as a critical technology for cleaner power generation. This study examines its benefits, including emissions reduction, enhanced efficiency, and compatibility with existing infrastructure. Concurrently, it addresses the inherent challenges, such as combustion dynamics, necessary infrastructure modifications, and economic considerations. Incorporating global case studies, the review explores recent advancements in hydrogen production and co-firing, evaluating their practicality and operational efficiency. It also provides a critical examination of the economic implications and societal acceptance, which are crucial for the adoption of hydrogen technologies. Moreover, the review investigates the synergy between hydrogen and renewable energy technologies, assessing potential impacts on energy policy and regulation. Ultimately, this comprehensive analysis underscores the essential role of hydrogen co-firing in the energy sector and advocates for a comprehensive strategy that synergizes technological progress, economic viability, regulatory support, and public engagement to promote a resilient and sustainable energy future.

hydrogen co-firing↗

DOE OSTI2026

Enhancers that direct gene expression to central nervous system vascular endothelial cells in vivo

CNS vascular endothelial cells (ECs) exhibit a distinctive gene expression program that is foundational for the blood-brain barrier (BBB). Previous research identified candidate cis-regulatory elements (CREs) that were hypothesized to control this program. In this work, transgenic mice and recombinant adeno-associated virus (rAAV) vectors have been used to interrogate these candidate CREs in vivo. These experiments show that an 850 bp genomic DNA segment ∼60 kb 5′ of Slc2a1 possesses enhancer activity that is (1) specific for BBB+ CNS ECs and (2) both necessary and sufficient for BBB+ EC gene expression. A screen of >8,000 genomic DNA segments from CNS EC-specific CRE candidates reveals several hundred with enhancer activity. Transcription factors ERG and LEF1 are shown to occupy sites in brain ECs that are highly enriched in candidate and experimentally validated CREs, lending strong support to a model in which canonical Wnt signaling activates the BBB program via LEF1.

CUT&RUN↗

DOE OSTI2024

Integrative analysis of the 3D genome and epigenome in mouse embryonic tissues

While a rich set of putative cis-regulatory sequences involved in mouse fetal development have been annotated recently on the basis of chromatin accessibility and histone modification patterns, delineating their role in developmentally regulated gene expression continues to be challenging. To fill this gap, here we mapped chromatin contacts between gene promoters and distal sequences across the genome in seven mouse fetal tissues and across six developmental stages of the forebrain. We identified 248,620 long-range chromatin interactions centered at 14,138 protein-coding genes and characterized their tissue-to-tissue variations and developmental dynamics. Integrative analysis of the interactome with previous epigenome and transcriptome datasets from the same tissues revealed a strong correlation between the chromatin contacts and chromatin state at distal enhancers, as well as gene expression patterns at predicted target genes. We predicted target genes of 15,098 candidate enhancers and used them to annotate target genes of homologous candidate enhancers in the human genome that harbor risk variants of human diseases. We present evidence that schizophrenia and other adult disease risk variants are frequently found in fetal enhancers, providing support for the hypothesis of fetal origins of adult diseases.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2026

Regulation compliant AI for fusion: explainable image-based feedback control of divertor detachment in DIII-D tokamak

While artificial intelligence (AI) has been promising for fusion control, its inherent black-box nature will make compliant implementation in regulatory environments a challenge. This study implements and validates a real-time AI-enabled linear and interpretable control system for successful divertor detachment control with the DIII-D lower divertor camera. Using D 2 gas, we demonstrate successful feedback divertor detachment control with a mean absolute difference of 2% from the target for both detachment and reattachment. This automatic training and linear processing framework can be extended to any image-based diagnostic for future fusion reactors.

computer vision↗

DOE OSTI2024

Sorghum SbGhd7 is a major regulator of floral transition and directly represses genes crucial for flowering activation

Molecular genetic understanding of flowering time regulation is crucial for sorghum development. GRAIN NUMBER, PLANT HEIGHT AND HEADING DATE 7 (SbGhd7) is one of the six classical loci conferring photoperiod sensitivity of sorghum flowering. However, its functions remain poorly studied. The molecular functions of SbGhd7 were characterized. The gene regulatory network controlled by SbGhd7 was constructed and validated. The biological roles of SbGhd7 and its major targets were studied. SbGhd7overexpression (OE) completely prevented sorghum flowering. Additionally, we show thatSbGhd7is a major negative regulator of flowering, binding to the promoter motif TGAATG(A/T)(A/T/C) and repressing transcription of the major florigen FLOWERING LOCUS T 10(SbFT10) and floral activators EARLY HEADING DATE(SbEhd1),FLAVIN-BINDING,KELCH REPEAT,F-BOX1(SbFKF1) and EARLY FLOWERING 3(SbELF3). Reinforcing the direct effect ofSbGhd7,SbEhd1OE activated the promoters of three functional florigens (SbFT1,SbFT8andSbFT10), dramatically accelerating flowering. Here our studies demonstrate that SbGhd7 is a major repressor of sorghum flowering by directly and indirectly targeting genes for flowering activation. The mechanism appears ancient. Our study extends the current model of floral transition regulation in sorghum and provides a framework for a comprehensive understanding of sorghum photoperiod response.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2021

Towards a dynamic photosynthesis model to guide yield improvement in C4 crops

Summary The most productive C4 food and biofuel crops, such as Saccharum officinarum (sugarcane), Sorghum bicolor (sorghum) and Zea mays (maize), all use NADP‐ME‐type C4 photosynthesis. Despite high productivities, these crops fall well short of the theoretical maximum solar conversion efficiency of 6%. Understanding the basis of these inefficiencies is key for bioengineering and breeding strategies to increase the sustainable productivity of these major C4 crops. Photosynthesis is studied predominantly at steady state in saturating light. In field stands of these crops light is continually changing, and often with rapid fluctuations. Although light may change in a second, the adjustment of photosynthesis may take many minutes, leading to inefficiencies. We measured the rates of CO 2 uptake and stomatal conductance of maize, sorghum and sugarcane under fluctuating light regimes. The gas exchange results were combined with a new dynamic photosynthesis model to infer the limiting factors under non‐steady‐state conditions. The dynamic photosynthesis model was developed from an existing C4 metabolic model for maize and extended to include: (i) post‐translational regulation of key photosynthetic enzymes and their temperature responses; (ii) dynamic stomatal conductance; and (iii) leaf energy balance. Testing the model outputs against measured rates of leaf CO 2 uptake and stomatal conductance in the three C4 crops indicated that Rubisco activase, the pyruvate phosphate dikinase regulatory protein and stomatal conductance are the major limitations to the efficiency of NADP‐ME‐type C4 photosynthesis during dark‐to‐high light transitions. We propose that the level of influence of these limiting factors make them targets for bioengineering the improved photosynthetic efficiency of these key crops.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2025

A novel regulator of the fungal phosphate starvation response revealed by transcriptional profiling and DNA affinity purification sequencing

Cells must accurately sense and respond to nutrients to compete for resources and establish growth. Phosphate is a critical nutrient source necessary for signaling, energy metabolism, and synthesis of nucleic acids, phospholipids, and cellular metabolites. During phosphate limitation, fungi import phosphate from the environment and liberate phosphate from phosphate-containing molecules in the cell. In the model filamentous fungus Neurospora crassa, the phosphate starvation response is regulated by the conserved transcription factor NUC-1. The activity of NUC-1 is repressed by a complex of the cyclin-dependent kinase MDK-1 and the cyclin PREG when phosphate is plentiful. When phosphate is limiting, NUC-1 repression by MDK-1/PREG is relieved by the cyclin-dependent kinase inhibitor NUC-2. We investigated the global response of N. crassa to phosphate starvation. During phosphate starvation, NUC-1 directly activated the expression of genes encoding phosphatases, nucleases, and a phosphate transporter and directly repressed genes associated with the ribosome. Additionally, NUC-1 indirectly activated the expression of an uncharacterized transcription factor, which we named nuc-3. NUC-3 directly repressed the expression of genes involved in phosphate acquisition and liberation after an extended period of phosphate starvation. Additionally, NUC-3 directly repressed the expression of the cyclin-dependent kinase inhibitor nuc-2. Thus, through the combination of NUC-3 direct repression of genes in the phosphate starvation response and nuc-2, an activator of the phosphate starvation response, NUC-3 serves to act as a brake on the phosphate starvation response after an extended period of phosphate starvation. This braking mechanism could reduce transcription, a phosphate-intensive process, under conditions of extended phosphate limitation.IMPORTANCEFungi have evolved regulatory networks to respond to available nutrients. Phosphate is often a limiting nutrient for fungi that is critical for many cellular functions, including nucleic acid and phospholipid biosynthesis, cell signaling, and energy metabolism. The fungal response to phosphate limitation is important in interactions with plants and animals. We investigated the global transcriptional response to phosphate starvation and the role of a major transcriptional regulator, NUC-1, in the model filamentous fungus Neurospora crassa. Our data show that NUC-1 is a bifunctional transcription factor that directly activates phosphate acquisition genes, while directly repressing genes associated with phosphate-intensive processes. NUC-1 indirectly regulates an uncharacterized transcription factor, which we named nuc-3. NUC-3 directly represses phosphate acquisition genes and nuc-2, an activator of the phosphate starvation response, during extended periods of phosphate starvation. Thus, NUC-3 acts as a brake on the phosphate starvation response to reduce phosphate-intensive activities, like transcriptional activation, when phosphate starvation persists.

DNA affinity purification sequencing↗

DOE OSTI2023

REC protein family expansion by the emergence of a new signaling pathway

This report presents multi-genome evidence that REC protein family expansion occurs when the emergence of new pathways gives rise to functional discordance. Specificity between residues in REC domain containing response regulators with paired histidine kinases is under negative purifying selection, constrained by the presence of other bacterial two-component systems signaling cascades that share sequence and structural identity. Presuming that the two-component systems can evolve by neutral amino acid changes (neutral drift) when purifying evolutionary constraints are relaxed, how might the REC protein family expand by amino acid changes when these constraints remain intact? Using an unsupervised machine learning approach to observe the sequence landscape of REC domains across long phylogenetic distances, we find that within-gene recombination, a subcategory of gene conversion, switched the effector domain and, consequently, the regulatory context of a duplicated response regulator from transcriptional regulation by σ54 to that by σ70. We determined that the recombined response regulator diverged from its parent by episodic diversifying selection and neutral drift. Functional experiments of the parent of recombined response regulators in a model Pseudomonas putida KT2440 model system revealed that the parent and recombined response regulators sense and respond to different carboxylic acids. Finally, a residue-switching experiment using structural predictions and functional characterization suggests that the new residues in the recombined regulator could form a new interaction interface and mediate condition-specific phosphotransfer. Overall, our study finds that genetic perturbations can create conditions of functional discordance, whereby the REC protein family can evolve by episodic diversifying selection.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2020

ATAC-Seq Reveals an Isl1 Enhancer That Regulates Sinoatrial Node Development and Function

Cardiac pacemaker cells (PCs) in the sinoatrial node (SAN) have a distinct gene expression program that allows them to fire automatically and initiate the heartbeat. Although critical SAN transcription factors, including Isl1 (Islet-1), Tbx3 (T-box transcription factor 3), and Shox2 (short-stature homeobox protein 2), have been identified, the cis-regulatory architecture that governs PC-specific gene expression is not understood, and discrete enhancers required for gene regulation in the SAN have not been identified. Here, to define the epigenetic profile of PCs using comparative ATAC-seq (assay for transposase-accessible chromatin with sequencing) and to identify novel enhancers involved in SAN gene regulation, development, and function.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2021

Coding and noncoding variants in EBF3 are involved in HADDS and simplex autism

Abstract Background Previous research in autism and other neurodevelopmental disorders (NDDs) has indicated an important contribution of protein-coding (coding) de novo variants (DNVs) within specific genes. The role of de novo noncoding variation has been observable as a general increase in genetic burden but has yet to be resolved to individual functional elements. In this study, we assessed whole-genome sequencing data in 2671 families with autism (discovery cohort of 516 families, replication cohort of 2155 families). We focused on DNVs in enhancers with characterized in vivo activity in the brain and identified an excess of DNVs in an enhancer named hs737. Results We adapted the fitDNM statistical model to work in noncoding regions and tested enhancers for excess of DNVs in families with autism. We found only one enhancer (hs737) with nominal significance in the discovery (p = 0.0172), replication (p = 2.5 × 10 −3 ), and combined dataset (p = 1.1 × 10 −4 ). Each individual with a DNV in hs737 had shared phenotypes including being male, intact cognitive function, and hypotonia or motor delay. Our in vitro assessment of the DNVs showed they all reduce enhancer activity in a neuronal cell line. By epigenomic analyses, we found that hs737 is brain-specific and targets the transcription factor gene EBF3 in human fetal brain. EBF3 is genome-wide significant for coding DNVs in NDDs (missense p = 8.12 × 10 −35 , loss-of-function p = 2.26 × 10 −13 ) and is widely expressed in the body. Through characterization of promoters bound by EBF3 in neuronal cells, we saw enrichment for binding to NDD genes (p = 7.43 × 10 −6 , OR = 1.87) involved in gene regulation. Individuals with coding DNVs have greater phenotypic severity (hypotonia, ataxia, and delayed development syndrome [HADDS]) in comparison to individuals with noncoding DNVs that have autism and hypotonia. Conclusions In this study, we identify DNVs in the hs737 enhancer in individuals with autism. Through multiple approaches, we find hs737 targets the gene EBF3 that is genome-wide significant in NDDs. By assessment of noncoding variation and the genes they affect, we are beginning to understand their impact on gene regulatory networks in NDDs.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2022

Dataset for report: Non-Technical Barriers to Geothermal Development in California and Nevada

In California and Nevada, geothermal projects are subject to non-technical barriers, which may create development delays leading to higher project costs and risks and decreased competitiveness with other electricity generation technologies. These non-technical barriers may include federal and state permits, authorizations, environmental reviews, and other regulatory requirements that are applicable throughout different phases of geothermal project development. The 2022 National Renewable Energy Laboratory (NREL) report, "Non-Technical Barriers to Geothermal Development in California and Nevada" presents findings of a study on non-technical barriers that may influence geothermal project development. The set of data resources relied upon for the report includes: 1) interviews conducted with relevant geothermal stakeholders including regulators and project developers, 2) federal and state environmental review documents developed for specific projects, 3) a techno-economic analysis conducted using the NREL Annual Technology Baseline (ATB) framework, and 4) an analysis of the impacts of site-specific land access and permitting considerations on project development readiness conducted using the Geothermal Resource Portfolio Optimization and Reporting Technique (GeoRePORT) Socioeconomic Assessment Tool (SEAT).

15 GEOTHERMAL ENERGY↗

DOE OSTI2020

Summary Overview of Content Guidance For Early Site Permit Applications

The Early Site Permit (ESP) process offered under Title 10 Code of Federal Regulations (CFR) Part 52, Subpart A, was promulgated by the Nuclear Regulatory Commission (NRC) in 1989 to address industry concerns with the former licensing process under 10 CFR Part 50. Previously, the licensing process required large expenditures of time and money by applicants well before key site specific environmental, safety, and emergency planning issues could be resolved. As envisioned, the ESP process is meant to resolve these issues well in advance of when a decision is made to build a nuclear power facility and before substantial capital is invested in the construction of a new nuclear facility. The ESP application is required to provide design information pertaining to structures, systems, and components along with site-specific parameters (such as meteorology, demographics, and hydrology) and address site safety, environmental protection, and emergency planning. Although an ESP applicant does not need to specify a particular nuclear plant design, as in construction permit applications, it does need to provide sufficient surrogate design information (developed to bound the nuclear plant design(s) that are being considered by the applicant) so that the NRC can make a determination on the acceptability of the site and the environmental impacts, and determine whether designs bounded by the surrogate design information provided by the applicant can be qualified for the proposed site.

11 NUCLEAR FUEL CYCLE AND FUEL MATERIALS↗

DOE OSTI2022

Mining G.O.L.D. (Geothermal Opportunities Leveraged Through Data): Exploring Synergies Between the Geothermal and Mining Industries

This report analyzes potential collaborations between the geothermal and locatable mineral industries (focused on the portion of the Basin and Range Province within Nevada, United States of America). The objectives of this study included analyzing: 1. The type and quality of data collected by the locatable mineral industry to determine feasibility for geothermal resource exploration; 2. The regulatory pathways and potential barriers that could prevent development of geothermal resources discovered via a mining claim (and vice versa) in the United States; 3. The historical development of geothermal resources discovered via mineral exploration data in the United States and illustrations of co-located mining and geothermal power projects; 4. The value propositions for both the locatable mineral and the geothermal industries to collaborate. This article concludes that many of the data collected by the mining industry as part of locatable mineral exploration (e.g., copper, gold, lithium) would also be useful for identifying and developing previously unknown geothermal resources (and in some notable cases, already have led to geothermal resource development). In addition, for minimal costs, the mining industry could catalogue these data and potentially monetize the data itself or use the data in the future to develop a geothermal project. Leveraging these locatable mineral data to develop geothermal resources and/or co-located minerals and geothermal resources would represent significant cost savings when compared to developing geothermal resources under a business-as-usual scenario as well as compared to current generating technologies (e.g., diesel-powered generators) employed at remote mining operations. Ultimately, leveraging mining industry data, knowledge, and expertise serves to effectively expand the geothermal exploration workforce, increase the rate of geothermal resource discovery, and potentially reduce geothermal electricity's levelized cost of energy (LCOE) by 23%-29%.

15 GEOTHERMAL ENERGY↗

DOE OSTI2022

Offshore Wind Energy Basics: Navigating Offshore Wind Energy Decision-Making Processes [Slides]

This webinar will provide a high-level summary of decision-making processes for siting and permitting, with a focus on the points at which local stakeholders can meaningfully engage in these processes. It will differentiate itself from other offshore wind webinars by presenting information relevant to a national audience (i.e., it will not be state specific) that helps stakeholders to "connect the dots" across agencies and processes. It will provide neutral, fact-based information from NREL staff, as well as from relevant staff at the federal, state, and local levels. This webinar will provide a helpful foundation for webinar #3 in this series, which will highlight opportunities for community engagement in the offshore wind development process more broadly, including but not limited to the regulatory processes covered in this webinar.

17 WIND ENERGY↗

DOE OSTI2022

Marine Energy Environmental Toolkit for Permitting and Licensing

The permitting and licensing process for marine energy in the U.S. could take up to 7 years for even small demonstrations or pilot studies despite existing resources like Tethys, MarineCadstre, and the FERC E-Library. The goal of the Marine Energy Environmental Toolkit Project was to increase regulators, developers, and marine energy innovators’ understanding of potential environmental effects of marine energy conversion devices deployments and provide information to make efficient and effective determinations during permitting and licensing; develop a one-stop shop for existing environmental, spatial, regulatory, and scientific data that allows users to access disparate sources of data through a series of tags and spatial queries. The Toolkit (marineenergy.app) builds upon prior U.S. Department of Energy (DOE) Water Power Technologies Office (WPTO) investments by pulling from WPTO-funded and outside resources to facilitate the permitting and licensing process, inform technology developers of required permits, and disseminate the latest knowledge of the environmental effects of these devices to regulators, developers, and marine energy innovators and stakeholders.

16 TIDAL AND WAVE POWER↗

DOE OSTI2023

Non-Technical Barriers to Geothermal Development in California and Nevada [Slides]

This presentation presents the findings of our study, including an analysis of federal, state, and local geothermal regulatory and permitting processes, case studies analyzing site-specific attributes that may impact project development at four selected geothermal project sites, an analysis of cost and timeline implications for geothermal project development, and the results of a qualitative study focused on inter-agency coordination and collaboration efforts between federal, state, and/or local agencies for geothermal projects located in California and Nevada. Our analysis found that development timelines may be impacted by multiple federal and state environmental review processes and duplicative permitting requirements as well as coordination efforts between numerous federal, state, and local agencies involved in issuing authorizations and permits necessary for project development. In addition, projects in California and Nevada may face site-specific environmental challenges due to the presence of sensitive resources (e.g., biological species and species habitat, cultural resources) that may result in project construction delays. Our study results also indicate that protracted geothermal development timelines caused by delays in acquiring necessary permits and environmental reviews may result in loss of generated electricity revenue and additional financing costs, which may increase economic uncertainty associated with project development. Through our qualitative analysis, we found that utilization of best practices, including tiering to existing environmental review documents and developing memoranda of understanding, which clearly delineate agency roles and responsibilities may reduce overall project timelines, costs, and uncertainties associated with geothermal project development in California and Nevada.

15 GEOTHERMAL ENERGY↗