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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 217 records · Page 12
All sourcesNASA NTRSDOE OSTI
DOE OSTI2021

Mercury methylation by metabolically versatile and cosmopolitan marine bacteria

Microbes transform aqueous mercury (Hg) into methylmercury (MeHg), a potent neurotoxin that accumulates in terrestrial and marine food webs, with potential impacts on human health. This process requires the gene pair hgcAB, which encodes for proteins that actuate Hg methylation, and has been well described for anoxic environments. However, recent studies report potential MeHg formation in suboxic seawater, although the microorganisms involved remain poorly understood. In this study, we conducted large-scale multi-omic analyses to search for putative microbial Hg methylators along defined redox gradients in Saanich Inlet, British Columbia, a model natural ecosystem with previously measured Hg and MeHg concentration profiles. Analysis of gene expression profiles along the redoxcline identified several putative Hg methylating microbial groups, including Calditrichaeota, SAR324 and Marinimicrobia, with the last the most active based on hgc transcription levels. Marinimicrobia hgc genes were identified from multiple publicly available marine metagenomes, consistent with a potential key role in marine Hg methylation. Computational homology modelling predicts that Marinimicrobia HgcAB proteins contain the highly conserved amino acid sites and folding structures required for functional Hg methylation. Furthermore, a number of terminal oxidases from aerobic respiratory chains were associated with several putative novel Hg methylators. Our findings thus reveal potential novel marine Hg-methylating microorganisms with a greater oxygen tolerance and broader habitat range than previously recognized.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Comparative genome analyses suggest a hemibiotrophic lifestyle and virulence differences for the beech bark disease fungal pathogens Neonectria faginata and Neonectria coccinea

Abstract Neonectria faginata and Neonectria coccinea are the causal agents of the insect-fungus disease complex known as beech bark disease (BBD), known to cause mortality in beech forest stands in North America and Europe. These fungal species have been the focus of extensive ecological and disease management studies, yet less progress has been made toward generating genomic resources for both micro- and macro-evolutionary studies. Here, we report a 42.1 and 42.7 mb highly contiguous genome assemblies of N. faginata and N. coccinea, respectively, obtained using Illumina technology. These species share similar gene number counts (12,941 and 12,991) and percentages of predicted genes with assigned functional categories (64 and 65%). Approximately 32% of the predicted proteomes of both species are homologous to proteins involved in pathogenicity, yet N. coccinea shows a higher number of predicted mitogen-activated protein kinase genes, virulence determinants possibly contributing to differences in disease severity between N. faginata and N. coccinea. A wide range of genes encoding for carbohydrate-active enzymes capable of degradation of complex plant polysaccharides and a small number of predicted secretory effector proteins, secondary metabolite biosynthesis clusters and cytochrome oxidase P450 genes were also found. This arsenal of enzymes and effectors correlates with, and reflects, the hemibiotrophic lifestyle of these two fungal pathogens. Phylogenomic analysis and timetree estimations indicated that the N. faginata and N. coccinea species divergence may have occurred at ∼4.1 million years ago. Differences were also observed in the annotated mitochondrial genomes as they were found to be 81.7 kb (N. faginata) and 43.2 kb (N. coccinea) in size. The mitochondrial DNA expansion observed in N. faginata is attributed to the invasion of introns into diverse intra- and intergenic locations. These first draft genomes of N. faginata and N. coccinea serve as valuable tools to increase our understanding of basic genetics, evolutionary mechanisms and molecular physiology of these two nectriaceous plant pathogenic species.

Salgado-Salazar, Catalina↗
DOE OSTI2024

African Swine Fever Virus Protein–Protein Interaction Prediction

The African swine fever virus (ASFV) is an often deadly disease in swine and poses a threat to swine livestock and swine producers. With its complex genome containing more than 150 coding regions, developing effective vaccines for this virus remains a challenge due to a lack of basic knowledge about viral protein function and protein–protein interactions between viral proteins and between viral and host proteins. In this work, we identified ASFV-ASFV protein–protein interactions (PPIs) using artificial intelligence-powered protein structure prediction tools. We benchmarked our PPI identification workflow on the Vaccinia virus, a widely studied nucleocytoplasmic large DNA virus, and found that it could identify gold-standard PPIs that have been validated in vitro in a genome-wide computational screening. We applied this workflow to more than 18,000 pairwise combinations of ASFV proteins and were able to identify seventeen novel PPIs, many of which have corroborating experimental or bioinformatic evidence for their protein–protein interactions, further validating their relevance. Two protein–protein interactions, I267L and I8L, I267L__I8L, and B175L and DP79L, B175L__DP79L, are novel PPIs involving viral proteins known to modulate host immune response.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2015

Bacterial microcompartment assembly: The key role of encapsulation peptides

Bacterial microcompartments (BMCs) are proteinaceous organelles used by a broad range of bacteria to segregate and optimize metabolic reactions. Their functions are diverse, and can be divided into anabolic (carboxysome) and catabolic (metabolosomes) processes, depending on their cargo enzymes. The assembly pathway for the β-carboxysome has been characterized, revealing that biogenesis proceeds from the inside out. The enzymes coalesce into a procarboxysome, followed by encapsulation in a protein shell that is recruited to the procarboxysome by a short (~17 amino acids) extension on the C-terminus of one of the encapsulated proteins. A similar extension is also found on the N- or C-termini of a subset of metabolosome core enzymes. These encapsulation peptides (EPs) are characterized by a primary structure predicted to form an amphipathic α-helix that interacts with shell proteins. In this study, we review the features, function and widespread occurrence of EPs among metabolosomes, and propose an expanded role for EPs in the assembly of diverse BMCs.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2026

metagRoot: a comprehensive database of protein families associated with plant root microbiomes

The plant root microbiome is vital in plant health, nutrient uptake, and environmental resilience. To explore and harness this diversity, we present metagRoot, a specialized and enriched database focused on the protein families of the plant root microbiome. MetagRoot integrates metagenomic, metatranscriptomic, and reference genome-derived protein data to characterize 71 091 enriched protein families, each containing at least 100 sequences. These families are annotated with multiple sequence alignments, CRISPR elements, hidden Markov models, taxonomic and functional classifications, ecosystem and geolocation metadata, and predicted 3D structures using AlphaFold2. MetagRoot is a powerful tool for decoding the molecular landscape of root-associated microbial communities and advancing microbiome-informed agricultural practices by enriching protein family information with ecological and structural context. The database is available at https://pavlopoulos-lab.org/metagroot/ or https://www.metagroot.org.

Chasapi, Maria N↗
DOE OSTI2012

Metagenomic Exploration of Viruses throughout the Indian Ocean

The characterization of global marine microbial taxonomic and functional diversity is a primary goal of the Global Ocean Sampling Expedition. As part of this study, 19 water samples were collected aboard the Sorcerer II sailing vessel from the southern Indian Ocean in an effort to more thoroughly understand the lifestyle strategies of the microbial inhabitants of this ultra-oligotrophic region. No investigations of whole virioplankton assemblages have been conducted on waters collected from the Indian Ocean or across multiple size fractions thus far. Therefore, the goals of this study were to examine the effect of size fractionation on viral consortia structure and function and understand the diversity and functional potential of the Indian Ocean virome. Five samples were selected for comprehensive metagenomic exploration; and sequencing was performed on the microbes captured on 3.0-, 0.8- and 0.1 µm membrane filters as well as the viral fraction (<0.1 µm). Phylogenetic approaches were also used to identify predicted proteins of viral origin in the larger fractions of data from all Indian Ocean samples, which were included in subsequent metagenomic analyses. Taxonomic profiling of viral sequences suggested that size fractionation of marine microbial communities enriches for specific groups of viruses within the different size classes and functional characterization further substantiated this observation. Functional analyses also revealed a relative enrichment for metabolic proteins of viral origin that potentially reflect the physiological condition of host cells in the Indian Ocean including those involved in nitrogen metabolism and oxidative phosphorylation. A novel classification method, MGTAXA, was used to assess virus-host relationships in the Indian Ocean by predicting the taxonomy of putative host genera, with Prochlorococcus, Acanthochlois and members of the SAR86 cluster comprising the most abundant predictions. This is the first study to holistically explore virioplankton dynamics across multiple size classes and provides unprecedented insight into virus diversity, metabolic potential and virus-host interactions.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2025

Machine learning guided selection of broad-spectrum epitope-specific functional antibodies for "Disease X"

Our project established and demonstrated a transfer learning framework that enables prediction of antibody–antigen interactions across related viruses. The approach focused on three major activities: 1. Conserved region and epitope identification – We compared viral protein structures and sequences to identify shared receptor-binding domains and neutralizing epitope regions across variants and related viruses. These conserved features formed the foundation for discovering broadly functional antibodies. 2. Machine learning model development – We built neural network–based models that integrate epitope features with antibody sequence information. Instead of relying solely on structural or physical properties, the models learned transferable patterns that describe antibody binding potential across different viral families. 3. Transfer learning and validation – Using SARS-CoV-2 and Ebola as source systems, we successfully transferred learned epitope features to predict antibody interactions for SARS CoV-1 and Marburg virus. Iterative cycles of dataset generation, retraining, and evaluation improved generalization and predictive power, ensuring the framework can adapt to new threats.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Proteome-wide association study and functional validation identify novel protein markers for pancreatic ductal adenocarcinoma

Pancreatic ductal adenocarcinoma (PDAC) remains a lethal malignancy, largely due to the paucity of reliable biomarkers for early detection and therapeutic targeting. Existing blood protein biomarkers for PDAC often suffer from replicability issues, arising from inherent limitations such as unmeasured confounding factors in conventional epidemiologic study designs. To circumvent these limitations, we use genetic instruments to identify proteins with genetically predicted levels to be associated with PDAC risk. Leveraging genome and plasma proteome data from the INTERVAL study, we established and validated models to predict protein levels using genetic variants. By examining 8,275 PDAC cases and 6,723 controls, we identified 40 associated proteins, of which 16 are novel. Functionally validating these candidates by focusing on 2 selected novel protein-encoding genes, GOLM1 and B4GALT1, we demonstrated their pivotal roles in driving PDAC cell proliferation, migration, and invasion. Furthermore, we also identified potential drug repurposing opportunities for treating PDAC.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Up-regulation of miR-122 protects against neuronal cell death in ischemic stroke through the heat shock protein 70-dependent NF-κB pathway by targeting FOXO3

Highlights: • miR-122 was decreased in mouse N2A neuroblastoma (N2A) cells after OGD and mouse brain after MCAO. • Gain-of-miR-122 function significantly suppressed OGD- and MCAO-induced injures in vitro and in vivo. • miR-122 was validated to directly bind to the predicted 3′-untranslated region (3′-UTR) of FOXO3. • The inhibitory effects of miR-122 on ischemic injury in vitro and in vivo were overturned by FOXO3 overexpression and miR-122-FOXO3 axis functioned via the heat shock protein 70 (HSP-70)-mediated NF-κB pathway. • miR-122 inhibits ischemic neuronal death through HSP-70-dependent NF-κB pathway by targeting FOXO3. Dysfunction of the microRNA (miRNA) network has been emerging as a main regulator in ischemic stroke. Recently, studies have linked the deregulation of miR-122 to ischemic stroke. However, the specific role and molecular mechanism of miR-122 in ischemic stroke remain to be further investigated. Here, we found that miR-122 was decreased in mouse N2A neuroblastoma (N2A) cells after oxygen-glucose deprivation (OGD) and mouse brain after transient middle cerebral artery occlusion (MCAO). OGD treatment significantly increased N2A cell death and Caspase-3 activity, and decreased Bcl-2 protein expression. In addition, MCAO treatment induced severe mouse brain infarction, mitochondrial reactive oxygen species (ROS) production, and long-term neurological deficit. Gain-of-miR-122 function significantly suppressed OGD- and MCAO-induced injures in vitro and in vivo. Subsequently, miR-122 was validated to directly bind to the predicted 3′-untranslated region (3′-UTR) of FOXO3 gene, and the inhibitory effects of miR-122 on ischemic injury in vitro and in vivo were overturned by FOXO3 overexpression. Moreover, our results further revealed that miR-122-FOXO3 axis functioned via the heat shock protein 70 (HSP-70)-mediated NF-κB pathway. Collectively, our data suggest that miR-122 inhibits ischemic neuronal death through the HSP-70-dependent NF-κB pathway by targeting FOXO3. These findings raise the possibility that this regulatory net may contribute to the pathogenesis of the ischemic brain injury in stroke.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2014

Protein expression, characterization and activity comparisons of wild type and mutant DUSP5 proteins

Background: The mitogen-activated protein kinases (MAPKs) pathway is critical for cellular signaling, and proteins such as phosphatases that regulate this pathway are important for normal tissue development. Based on our previous work on dual specificity phosphatase-5 (DUSP5), and its role in embryonic vascular development and disease, we hypothesized that mutations in DUSP5 will affect its function. Results: In this study, we tested this hypothesis by generating full-length glutathione-S-transferase-tagged DUSP5 and serine 147 proline mutant (S147P) proteins from bacteria. Light scattering analysis, circular dichroism, enzymatic assays and molecular modeling approaches have been performed to extensively characterize the protein form and function. We demonstrate that both proteins are active and, interestingly, the S147P protein is hypoactive as compared to the DUSP5 WT protein in two distinct biochemical substrate assays. Furthermore, due to the novel positioning of the S147P mutation, we utilize computational modeling to reconstruct full-length DUSP5 and S147P to predict a possible mechanism for the reduced activity of S147P. Conclusion: Taken together, this is the first evidence of the generation and characterization of an active, full-length, mutant DUSP5 protein which will facilitate future structure-function and drug development-based studies.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

Longitudinal proteomics analysis in the immediate microenvironment of islet allografts during progression of rejection

The applicability and benefits of pancreatic islet transplantation are limited due to various issues including the need to avoid immune-mediated rejection. In the present study, we used our experimental platform of allogeneic islet transplant in the anterior chamber of the eye (ACE-platform) to longitudinally monitor the progress of rejection in mice and obtain aqueous humor samples representative of the microenvironment of the graft for accurately-timed proteomic analyses. LC-MS/MS-based proteomics performed on such mass-limited samples (~5 μL) identified a total of 1296 proteins. Various analyses revealed distinct protein patterns associated with the mounting of the inflammatory and immune responses and their evolution with the progression of the rejection. Pathway analyses indicated predominant changes in cytotoxic functions, cell movement, and innate and adaptive immune responses. Furthermore, network prediction analyses revealed transition from humoral to cellular immune response and exacerbation of pro-inflammatory signaling. One of the proteins identified by this localized proteomics as a candidate biomarker of islet rejection, Cystatin 3, was further validated by ELISA in the aqueous humor. This study provides (1) experimental evidence demonstrating the feasibility of longitudinal localized proteomics using small aqueous humor samples and (2) proof-of-concept for the discovery of biomarkers of impending immune attack from the immediate local microenvironment of ACE-transplanted islets.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Comparative Assessment of Pose Prediction Accuracy in RNA–Ligand Docking

Structure-based virtual high-throughput screening is used in early-stage drug discovery. Over the years, docking protocols and scoring functions for protein–ligand complexes have evolved to improve the accuracy in the computation of binding strengths and poses. In the past decade, RNA has also emerged as a target class for new small-molecule drugs. However, most ligand docking programs have been validated and tested for proteins and not RNA. Here, we test the docking power (pose prediction accuracy) of three state-of-the-art docking protocols on 173 RNA–small molecule crystal structures. The programs are AutoDock4 (AD4) and AutoDock Vina (Vina), which were designed for protein targets, and rDock, which was designed for both protein and nucleic acid targets. AD4 performed relatively poorly. For RNA targets for which a crystal structure of a bound ligand used to limit the docking search space is available and for which the goal is to identify new molecules for the same pocket, rDock performs slightly better than Vina, with success rates of 48% and 63%, respectively. However, in the more common type of early-stage drug discovery setting, in which no structure of a ligand–target complex is known and for which a larger search space is defined, rDock performed similarly to Vina, with a low success rate of ~27%. Further, Vina was found to have bias for ligands with certain physicochemical properties, whereas rDock performs similarly for all ligand properties. Thus, for projects where no ligand–protein structure already exists, Vina and rDock are both applicable. However, the relatively poor performance of all methods relative to protein–target docking illustrates a need for further methods refinement.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Thermophilic Chassis-Enabled High-Throughput Selection of a Thermostable Fluorogenic Reporter

Thermostable proteins show increased shelf life and performance at elevated temperatures and under harsh conditions, resulting in lower costs for various industrial and biotechnological applications. However, due to a limited understanding of the relationship between stability and function, protein stabilization remains primarily a trial-and-error approach. Therefore, building a combinatorial library of mutations predicted to improve stability, followed by experimental testing, represents a markedly improved methodology. However, the lack of high-throughput approaches to screen even a moderately sized library presents a major bottleneck in the field. Here, in this study, we use a thermophile, Parageobacillus thermoglucosidasius (Ptherm) to rapidly screen combinatorial libraries consisting of rationally designed thermostabilizing mutations (∼10 3 –10 4 ) of a mesophilic fluorescent reporter, Y-FAST. On a Petri dish, microbial growth at an elevated temperature and exposure to fluorogen yielded several colonies of Ptherm that showed distinct fluorescence at 55 and 68 °C in our two sequentially generated libraries using Rosetta and ProteinMPNN, respectively. The Y-FAST variants isolated from fluorescent colonies were brighter than Y-FAST and showed higher resistance to thermal and chemical denaturation. AlphaFold-predicted structures and MD simulations revealed stability-enhancing salt bridges and hydrogen bond networks in the isolated FAST variants. The moderately thermostable FAST (tsFAST) and hyperstable FAST (hsFAST) were then demonstrated as translation reporters for protein expression and folding at elevated temperatures, such as 55 and 68 °C. Our approach of combinatorial library generation and high-throughput screening in a thermophilic chassis could, in principle, be extended to other proteins fused to these translation reporters. Furthermore, the hsFAST protein is small─half the size of the green fluorescent protein─and does not require oxygen for maturation, making it ideal for engineering extremophilic anaerobes for biosensing and bioconversion.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2014

Investigating the roles of MicroRNAs in biotic stress response induced by Rhizoctonia solani in rice

Sheath blight disease, caused by Rhizoctonia solani 1802/KB was screened on two rice varieties, Oryza sativaindica cultivar MR219 and Oryza sativa indica cultivar UKMRC9. The disease symptom was severe in MR219 compared to UKMRC9. Total RNA from R. solani 1802/KB, infected rice leaves of MR219 and infected rice leaves of UKMRC9 were extracted using TRIzol reagent, purified and sent for small RNA sequencing. Three miRNA libraries were generated and analyzed. The libraries generated 65 805, 78 512 and 81 325 known miRNAs respectively. The structure of miRNA of these samples was predicted. The up-regulated and down-regulated of miRNAs target gene prediction and its target functions were discovered and were mainly related to the growth and development of metabolism, protein transport, transcriptional regulation, stress response, and hormone signaling and electron transfer. Sheath blight-induced differential expression of known miRNAs tends to targetMYB transcription factor, F-box proteins, NBS-LRR, leucine-rich repeat receptor protein kinases and zinc finger proteins. Detecting new miRNAs and measuring the expression profiles of known miRNAs is an important tasks required for a better understanding of various biological conditions. Therefore, further analysis using Gene Ontology Slim will be conducted to deduce some biological information from the datasets obtained.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2016

Proteome Remodeling in Response to Sulfur Limitation in “ Candidatus Pelagibacter ubique”

“ Ca . Pelagibacter ubique” is a key driver of marine biogeochemistry cycles and a model for understanding how minimal genomes evolved in free-living anucleate organisms. This study explores the unusual sulfur acquisition strategy that has evolved in these cells, which lack assimilatory sulfate reduction and instead rely on reduced sulfur compounds found in oxic marine environments to meet their cellular quotas. Our findings demonstrate that the sulfur acquisition systems are constitutively expressed but the enzymatic steps leading to the essential sulfur-containing amino acid methionine are regulated by a unique array of riboswitches and genes, many of which are encoded in a rapidly evolving genome region. These findings support mounting evidence that streamlined cells have evolved regulatory mechanisms that minimize transcriptional switching and, unexpectedly, localize essential sulfur acquisition genes in a genome region normally associated with adaption to environmental variation.

Smith, Daniel P.↗
DOE OSTI2022

Phage K gp102 Drives Temperature-Sensitive Antibacterial Activity on USA300 MRSA

There is widespread interest in using obligately lytic bacteriophages (“phages”) to treat human bacterial infections. Among Staphylococcus aureus infections, the USA300 lineage is a frequent cause of invasive disease. We observed that phage K, a model S. aureus myophage, exhibits temperature-sensitive growth on USA300 strains, with the wild-type phage providing poorer growth suppression in broth and forming smaller and fainter plaques at 37 °C vs. 30 °C. We isolated 65 mutants of phage K that had improved plaquing characteristics at 37 °C when compared to the parental phage. In all 65 mutants, this phenotype was attributable to loss-of-function (LoF) mutations in gp102, which encodes a protein of unknown function that has homologs only among the Herelleviridae (SPO1-like myophages infecting gram-positive bacteria). Additional experiments with representative mutants consistently showed that the temperature-sensitive plaque phenotype was specific to USA300 MRSA strains and that Gp102 disruption was correlated with improved suppression of bacterial growth in broth and improved antibacterial activity in a mouse model of upper respiratory tract infection. The same genotype and in vitro phenotypes could be replicated in close relatives of phage K. Gp102 disruption did not have a detectable effect on adsorption but did delay cell culture lysis relative to wild-type under permissive infection conditions, suggesting that gp102 conservation might be maintained by selective pressure for more rapid replication. Expression of gp102 on a plasmid was toxic to both an MSSA and a USA300 MRSA strain. Molecular modeling predicts a protein with two helix-turn-helix domains that displays some similarity to DNA-binding proteins such as transcription factors. While its function remains unclear, gp102 is a conserved gene that is important to the infection process of Kayvirus phages, and it appears that the manner in which USA300 strains defend against them at 37 °C can be overcome by gp102 LoF mutations.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2013

mir-30d Regulates multiple genes in the autophagy pathway and impairs autophagy process in human cancer cells

Highlights: ► Gene set enrichment analysis indicated mir-30d might regulate the autophagy pathway. ► mir-30d represses the expression of BECN1, BNIP3L, ATG12, ATG5 and ATG2. ► BECN1, BNIP3L, ATG12, ATG5 and ATG2 are direct targets of mir-30d. ► mir-30d inhibits autophagosome formation and LC3B-I conversion to LC3B-II. ► mir-30d regulates the autophagy process. -- Abstract: In human epithelial cancers, the microRNA (miRNA) mir-30d is amplified with high frequency and serves as a critical oncomir by regulating metastasis, apoptosis, proliferation, and differentiation. Autophagy, a degradation pathway for long-lived protein and organelles, regulates the survival and death of many cell types. Increasing evidence suggests that autophagy plays an important function in epithelial tumor initiation and progression. Using a combined bioinformatics approach, gene set enrichment analysis, and miRNA target prediction, we found that mir-30d might regulate multiple genes in the autophagy pathway including BECN1, BNIP3L, ATG12, ATG5, and ATG2. Our further functional experiments demonstrated that the expression of these core proteins in the autophagy pathway was directly suppressed by mir-30d in cancer cells. Finally, we showed that mir-30d regulated the autophagy process by inhibiting autophagosome formation and LC3B-I conversion to LC3B-II. Taken together, our results provide evidence that the oncomir mir-30d impairs the autophagy process by targeting multiple genes in the autophagy pathway. This result will contribute to understanding the molecular mechanism of mir-30d in tumorigenesis and developing novel cancer therapy strategy.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2023

A chloroplast protein atlas reveals punctate structures and spatial organization of biosynthetic pathways

Chloroplasts are eukaryotic photosynthetic organelles that drive the global carbon cycle. Despite their importance, our understanding of their protein composition, function, and spatial organization remains limited. Here, we determined the localizations of 1,034 candidate chloroplast proteins using fluorescent protein tagging in the model alga Chlamydomonas reinhardtii. The localizations provide insights into the functions of poorly characterized proteins; identify novel components of nucleoids, plastoglobules, and the pyrenoid; and reveal widespread protein targeting to multiple compartments. We discovered and further characterized cellular organizational features, including eleven chloroplast punctate structures, cytosolic crescent structures, and unexpected spatial distributions of enzymes within the chloroplast. We also used machine learning to predict the localizations of other nuclear-encoded Chlamydomonas proteins. The strains and localization atlas developed here will serve as a resource to accelerate studies of chloroplast architecture and functions.

59 BASIC BIOLOGICAL SCIENCES↗