Quinoa (Chenopodium quinoa) is a nutrient-rich pseudocereal with diverse specialized metabolites, yet the genetic basis of this metabolic diversity is poorly understood. Here we integrate whole-genome sequencing and multi-tissue metabolic profiling of 603 quinoa accessions. We detected 4688 metabolic features and identified over 1000 metabolites in seeds, leaves, and roots. Using multi-tissue genome-wide association, we mapped the genetic architecture of quinoa metabolome by identifying 584 quantitative trait loci (QTL) and prioritized 219 candidate genes across 58 major QTL governing saponin, betalain, and flavonoid biosynthesis. Moreover, we constructed a drought-responsive multi-omics regulatory network and uncovered additional key genes involved in quinoa stress signaling and metabolic pathways. Finally, we cloned and functional validated the roles of cytochrome P450 76AD1 (CYP76AD1) in betalamate accumulation, UDP-glycosyltransferase (UGT91C1) in flavonoid glycosylation, and CYP72A154 and soyasapogenol B glucuronide galactosyltransferase in saponin biosynthesis. This multi-omic framework provides a high-resolution map of the quinoa metabolome and a foundation for breeding nutrient-rich and stress-resilient quinoa cultivars.

RNA covalent modifications (RCMs) influence RNA stability and translation efficiency, and they thus play critical roles in eukaryotic growth and development. However, their role in regulating plant performance under abiotic stress remains largely unexplored. Here, we integrated multi-omics data in 6 Sorghum bicolor accessions under water-limiting conditions in the field to explore the relationship between RCMs and drought response. Within a stress- and photosynthesis-associated gene co-expression module, we identified SbDUS2, a member of a family of enzymes conserved across eukaryotes, that catalyzes the reduction of uracil to dihydrouridine (DHU) on RNA molecules. DHU-modified transcripts in this module were enriched for photosynthetic functions and showed strong correlation with photosynthetic traits. To elucidate the function of this RCM, we characterized loss-of-function dus2 mutants in Arabidopsis thaliana. Under control conditions, these DHU-deficient mutants exhibited impaired germination and delayed development. Furthermore, under water-limiting or heat conditions, these mutants showed significantly reduced net CO2 assimilation and survival. Using multiple transcriptome-wide RNA stability assays, we demonstrated that transcripts associated with lower DHU levels in a dus2 background generally exhibited increased stability compared to Col-0 controls. Particularly, lack of DUS2 led to the hyperstability of photosynthesis-related transcripts, impeding their turnover and likely preventing proper photosynthetic acclimation during stress. We propose a model where DHU acts as a critical post-transcriptional regulator marking mRNAs for rapid turnover under stress, highlighting an overlooked regulatory layer contributing to plant resilience.

Plastids are dynamic organelles that remodel their composition, ultrastructure, and function according to developmental and environmental demands. The synthetic molecule X57 induces the conversion of leaf chloroplasts into tocopherol-rich plastids lacking thylakoids and containing proliferating plastoglobules. Removal of X57 triggers chloroplast redifferentiation, enabling precise spatial-temporal dissection of these transitions. X57 directly binds and inhibits the phosphatase SAL1, causing accumulation of its substrate 3 '-phosphoadenosine 5 '-phosphate (PAP), a retrograde signal that modulates nuclear gene expression. SAL1 inhibition activates a cascade that depletes cytokinins and down-regulates GOLDEN2-LIKE1 (GLK1) and other transcription factors involved in chloroplast biogenesis. SAL1-defective mutants fail to undergo this signaling pathway. The SAL1-PAP-mediated weakening of chloroplast identity preconditions plastids for their eventual conversion into storage-type organelles upon X57-promoted SAL1-independent accumulation of tocopherols. After X57 withdrawal, photosynthetic gene expression and chloroplast functions are restored. This framework identifies key molecular mechanisms underlying chloroplast plasticity, a central process in biology.

Cyclic adenosine monophosphate (cAMP) is a fundamental second messenger involved in diverse signaling pathways across both animals and plants. While the role of 3 ',5 '-cAMP has been extensively characterized, the biological significance of its structural isomer, 2 ',3 '-cAMP, remains largely unexplored, particularly in plants. Here, we show that 2 ',3 '-cAMP and 3 ',5 '-cAMP represent parallel signaling systems in Arabidopsis thaliana, with different enzymatic origins and largely distinct downstream effects. In vitro enzymatic assays show that plant adenylate cyclases (ACs), including AFB5 and HpAC1, produce specifically 3 ',5 '-cAMP from ATP, whereas the TIR domain of protein L7 also catalyzes the formation of 2 ',3 '-cAMP from RNA. Comprehensive multiomics analyses reveal that two isomers elicit distinct yet partially overlapping metabolic, proteomic, and transcriptional response: 2 ',3 '-cAMP activates broad, stress-adaptive gene expression reprogramming, while 3 ',5 '-cAMP fine-tunes responses related to nutrient status and cellular homeostasis. Our findings establish the existence of dual cAMP signaling systems in plants, each with specialized functions and provide insights into the complex regulatory networks governing plant physiology.

Protein-metabolite interactions (PMIs) are critical regulators of cellular processes, yet their roles in plant-pathogen interactions remain poorly understood. Here, we applied PROMIS (Protein-Metabolite Interactions using Size separation) to map the Arabidopsis protein-metabolite interactome during infection with Pseudomonas syringae pv. tomato DC3000 (Pto DC3000). PROMIS resolved elution profiles for >5,700 proteins and 192 annotated metabolites across infected, systemic, and control tissues, revealing infection-associated remodeling of protein assemblies and metabolite co-fractionation patterns. Integrating co-fractionation with coronatine-induced transcriptional responses, we identified NATA1, a polyamine N-acetyltransferase, as a coronatine-associated host protein binder. Biochemical assays confirmed direct coronatine-NATA1 interaction (Kd = 586 nm), whereas the homolog NATA2 did not bind coronatine. Coronatine did not strongly inhibit NATA1 catalytic turnover under our assay conditions but consistently depressed NATA1 dimer formation in vitro and in planta. In silico docking together with targeted mutational analyses identified a coronatine-sensitive interface on NATA1, revealing residue-specific coupling between ligand binding, assembly state, and acetyl-CoA engagement. Collectively, these findings establish PROMIS as a robust platform for infection-state PMI discovery and uncover a coronatine-responsive regulatory node that links small-molecule signaling to host polyamine acetylation.

Mineral nutrient uptake and deposition profoundly influence plant development, stress resilience, and productivity. Silicon (Si), though classified as a non-essential element, significantly influences a plant's physiology, particularly in fortifying defense responses and mitigating stress. While the genetic and molecular mechanisms of Si uptake and transport are well studied in monocots, particularly rice, their role in dicot species, such as soybean, remains unclear at the cellular and molecular levels. In this study, we utilized single-nucleus RNA sequencing (snRNA-seq) to dissect cellular responses to Si accumulation in soybean leaves. We identified distinct cellular populations, including a unique Si-induced or Si-associated cell cluster within vascular cells, suggesting a specialized mechanism of Si distribution. Si treatment notably induced the expression of defense-related genes, with a pronounced enrichment in vascular cells, underscoring their pivotal role in activating plant defense mechanisms. Moreover, Si modulated the expression of genes involved in phytoalexin biosynthesis, salicylic acid, and immune receptor signaling, suggesting transcriptional priming of genes involved in defense responses. Further investigation of Si transporters revealed precise expression of an Si efflux gene in epidermal cells in response to Si treatment. We also validated the role of efflux Si transporters using a Xenopus oocyte assay and CRISPR/Cas9 genome editing of composite soybean plant roots. This study provides critical insights into the biotic stress regulatory networks influenced by Si treatment in soybean leaves at the single-cell level, thus laying the foundation for enhancing stress tolerance through optimized mineral nutrient uptake.

Diketopiperazines (DKPs) are chemically and functionally diverse cyclic dipeptides associated primarily with microbes. Few DKPs have been reported from plants and animals; the best characterized is cyclo(His-Pro), found in the mammalian central nervous system, where it arises from the proteolytic cleavage of a thyrotropin-releasing tripeptide hormone. Herein, we report the identification of cyclo(His-Pro) in Arabidopsis (Arabidopsis thaliana), where its levels increase upon abiotic stress conditions, including high salt, heat, and cold. To screen for potential protein targets, we used isothermal shift assays, which examine changes in protein-melting stability upon ligand binding. Among the identified proteins, we focused on the glycolytic enzyme, cytosolic glyceraldehyde-3-phosphate dehydrogenase (GAPC1). Binding between the GAPC1 protein and cyclo(His-Pro) was validated using nano-differential scanning fluorimetry and microscale thermophoresis, and we could further demonstrate that cyclo(His-Pro) inhibits GAPC1 activity with an IC50 of ∼200 μm. This inhibition was conserved in human GAPDH. Inhibition of glyceraldehyde-3-phosphate dehydrogenase activity has previously been reported to reroute carbon from glycolysis toward the pentose phosphate pathway. Accordingly, cyclo(His-Pro) supplementation augmented NADPH levels, increasing the NADPH/NADP+ ratio. Phenotypic screening revealed that plants supplemented with cyclo(His-Pro) were more tolerant to high-salt stress, as manifested by higher biomass, which we show is dependent on GAPC1/2. Our work reports the identification and functional characterization of cyclo(His-Pro) as a modulator of glyceraldehyde-3-phosphate dehydrogenase in plants.

In their natural environment, plants experience temperature fluctuations, including intensified heat waves driven by climate change, which pose significant threats to their productivity. To adapt, plants have evolved diverse mechanisms to withstand heat stress (HS), minimizing potential damage and ensuring survival. One such adaptation is acquired thermotolerance (AT), where prior exposure to HS primes plants to withstand subsequent severe HS. AT can persist for several days and involves a recovery period during which plants establish heat stress memory (HSM), reorganize cellular processes, and strengthen stress resilience. The molecular mechanisms underlying HSM remain the subject of active investigation. In this study, we employ a high-throughput comparative multi-omics approach to unravel the transcriptome, metabolome, and proteome of Arabidopsis thaliana seedlings during distinct intervals of the HSM phase. Our findings provide insights into the intricacies of HS recovery and the memory process. Notably, distinct temporal responses emerge at both the transcriptional and protein levels during the early and late recovery phases. Transcripts associated with HSM are upregulated during the early HS recovery phase, indicating a rapid response crucial for initial memory formation, while corresponding protein levels remain elevated throughout the recovery period, supporting memory consolidation. Additionally, metabolite profiles reveal distinctive patterns across the HS memory phase. This marks the first detailed multi-omic analysis of the HSM phase in Arabidopsis seedlings, providing insights into the multifaceted nature of this complex process. These comprehensive datasets hold promise in elucidating regulators of HS resilience, thereby enhancing efforts in breeding HS-tolerant crops ### Competing Interest Statement The authors have declared no competing interest.

Mitochondrial disease encompasses a group of genetically inherited disorders hallmarked by an inability of the respiratory chain to produce sufficient ATP. These disorders present with multisystemic pathologies that predominantly impact highly energetic tissues such as skeletal muscle. There is no cure or effective treatment for mitochondrial disease. We have discovered a small molecule known as oxybutynin that can bypass complex III mitochondrial dysfunction in primary murine and human skeletal muscle progenitor cells (MPCs). Oxybutynin administration improves MPC proliferative capacity, enhances cellular glycolytic function, and improves myotube formation. Mechanistically, results from our isothermal shift assay indicate that oxybutynin interacts with a suite of proteins involved in mRNA processing, which then trigger the upregulation of biological pathways to circumvent CIII mitochondrial dysfunction. Taken together, we provide evidence for the small molecule oxybutynin as a potential therapeutic candidate for the future treatment of CIII mitochondrial dysfunction.NEW & NOTEWORTHY Mitochondrial disease currently has no cure and affects highly energetic tissues such as skeletal muscle. Under disease conditions, the stem cell pool of the tissue is depleted and has reduced regenerative capacity, resulting in functional decline of the muscle. Here, we have identified the small molecule oxybutynin as a potential treatment option, as it improved the regenerative capacity of skeletal muscle stem cells harboring a complex III mitochondrial disease mutation.

Covalent RNA modifications (RCMs) are post-transcriptional changes to the chemical composition of RNA. RCMs influence mRNA stability, regulate transcription and translation efficiency, and play critical roles in the growth and development of eukaryotes. However, their role in plants remains poorly understood, particularly as they relate to performance in the field under stress conditions. In this study, we grew a panel of six diverse sorghum (S. bicolor) accessions in the field during the summer in central Arizona and examined their physiological and molecular responses to drought and heat stress over time. We then explored the molecular features that contributed to plant performance under stress. To do so, we combined genomic, transcriptomic, epitranscriptomic, physiological, and metabolomic data in a systems-level approach. Co-expression network analyses uncovered two modules of interest, one controlled primarily by a single stress-responsive transcription factor, SbCDF3, and the other by an RCM, dihydrouridine. While the CDF3 module largely contained a set of stress response and photosynthesis-associated genes that were positively correlated with plant performance, the dihydrouridine-associated module was largely comprised of photosynthesis and metabolism genes, including SbPPDK1, which is integral to C4 photosynthesis in the grasses. In addition, the transcript encoding the enzyme responsible for this RCM, dihydrouridine synthase (SbDUS2), was also present in this module, and its abundance was positively correlated with photosynthetic traits and SbPPDK1 abundance. Given that this highly conserved RCM has never been characterized in plants, we examined loss of function mutants for the DUS2 enzyme in Arabidopsis, demonstrating decreases in plant growth and performance under heat stress in this background. Our work highlights both a key transcription factor, CDF3, for breeding in the Poaceae. In addition, for the first time in plants, we reveal a role for the RCM dihydrouridine in modifying conserved, core metabolic and photosynthesis-associated transcripts in plants. ### Competing Interest Statement The authors have declared no competing interest.

RNA Covalent Modifications (RCMs) are post-transcriptional chemical alterations that influence RNA stability and translation efficiency, thus play critical roles in eukaryotic growth and development. However, their role in regulating plant performance under abiotic stress remain largely unexplored. Here, we integrated multi-omics data in six Sorghum bicolor accessions under water-limiting conditions in the field to explore the relationship between RCMs and drought response. Within a stress and photosynthesis-associated gene co-expression module, we identified SbDUS2, a member of family of enzymes, conserved across eukaryotes, which catalyzes the reduction of uracil to dihydrouridine (DHU) on RNA molecules. DHU-modified transcripts in this module were enriched for photosynthetic functions and showed strong correlation with photosynthetic traits. To elucidate the function of this RCM, we characterized loss of function dus2 mutants in the genetic model, Arabidopsis thaliana. Under control conditions, these DHU-deficient mutants exhibited impaired germination and delayed development. Furthermore, when exposed to heat or water-limiting conditions, these mutants showed significantly reduced net CO2 assimilation and survival. Using multiple transcriptome-wide RNA stability assays, we demonstrated that transcripts associated with lower DHU level in a dus2 background generally exhibited increased stability compared to Col-0 controls. Particularly, lack of DUS2 led to the hyperstability of photosynthesis-related transcripts, impeding their turnover and likely preventing proper photosynthetic acclimation during stress. We propose a model based on these data where DHU acts as a critical post-transcriptional regulator marking mRNAs for rapid turnover under stress, highlighting an overlooked regulatory layer contributing to plant resilience.

Climate change and especially the concomitant increasing frequency of drought and salinity induced water-deficit stresses, is a major limiting factor of plant growth and crop productivity worldwide. Understanding the adaptive responses of plants to water-deficit stress has therefore become a central challenge in plant biotechnology. Considerable evidence indicates that late embryogenesis abundant (LEA) proteins contribute to water-deficit stress tolerance and the stabilization of metabolic enzymes, yet the molecular basis of these responses remains unknown. To date, limited direct evidence for specific in vivo interactions between LEA proteins and their molecular targets has been reported. Here, we identify for the first time the native in vivo interactome of three Arabidopsis dehydrin LEA proteins - LEA4 (COR47), LEA5 (ERD10), and LEA10 (ERD14) - expressed under their own promoters and in response to salt stress. Our results show that LEA4, LEA5, and LEA10 dehydrins interact with each other, and share six common candidate protein interactors, suggesting that they form a core protein complex in salt stress conditions. We confirmed the direct protein-protein interaction between LEA5 with LEA4, LEA10, OSCA3, PLD α 1, PIP2B and OST1/AHA1 at the biochemical level. Phenotypic analyses of loss-of-function genetic mutants revealed that the interactions of LEA5 with OSCA3 and PIP2B promote seed germination under salt stress. LEA5 interaction with PLDα1 promotes root growth under salt stress, while its interaction with PIP2B enhances root growth under osmotic stress, indicating distinct stress-specific functional roles for each interaction. In conclusion, this study identifies the native in vivo interactions of three dehydrin proteins, while uncovering the functional relevance of these interactions under salt and osmotic stress conditions, thus providing novel mechanistic insights into the role of LEA proteins in water-deficit stress adaptation. ### Competing Interest Statement The authors have declared no competing interest. Ministerio de Ciencia, Innovación y Universidades, https://ror.org/05r0vyz12, RYC2022-038173-I

Climate change and especially the concomitant increasing frequency of drought and salinity induced water-deficit stresses, is a major limiting factor of plant growth and crop productivity worldwide. Understanding the adaptive responses of plants to water-deficit stress has therefore become a central challenge in plant biotechnology. Considerable evidence indicates that late embryogenesis abundant (LEA) proteins contribute to water-deficit stress tolerance and the stabilization of metabolic enzymes, yet the molecular basis of these responses remains unknown. To date, limited direct evidence for specific in vivo interactions between LEA proteins and their molecular targets has been reported. Here, we identify for the first time the native in vivo interactome of three Arabidopsis dehydrin LEA proteins - LEA4 (COR47), LEA5 (ERD10), and LEA10 (ERD14) - expressed under their own promoters and in response to salt stress. Our results show that LEA4, LEA5, and LEA10 dehydrins interact with each other, and share six common candidate protein interactors, suggesting that they form a core protein complex in salt stress conditions. We confirmed the direct protein-protein interaction between LEA5 with LEA4, LEA10, OSCA3, PLD α 1, PIP2B and OST1/AHA1 at the biochemical level. Phenotypic analyses of loss-of-function genetic mutants revealed that the interactions of LEA5 with OSCA3 and PIP2B promote seed germination under salt stress. LEA5 interaction with PLDα1 promotes root growth under salt stress, while its interaction with PIP2B enhances root growth under osmotic stress, indicating distinct stress-specific functional roles for each interaction. In conclusion, this study identifies the native in vivo interactions of three dehydrin proteins, while uncovering the functional relevance of these interactions under salt and osmotic stress conditions, thus providing novel mechanistic insights into the role of LEA proteins in water-deficit stress adaptation.

Toward characterization of protein-metabolite interactomes, we recently introduced PROMIS, a co-fractionation-based mass spectrometry approach. However, the challenge lies in distinguishing true interactors from coincidental co-elution when a metabolite co-fractionates with numerous proteins. To address this, we integrated two chromatographic techniques-size exclusion and ion exchange-to enhance the mapping of protein-metabolite interactions (PMIs) in Escherichia coli. This integration aims to refine the PMI network by considering size and charge characteristics, resulting in 994 interactions involving 51 metabolites and 465 proteins. The PMI network is enriched for known and predicted interactions, providing validation. Furthermore, analyzing protein targets for different metabolites revealed functional insights, such as a connection between proteinogenic dipeptides and fatty acid biosynthesis. Notably, we uncovered an inhibitory interaction between the riboflavin degradation product lumichrome and orotate phosphoribosyltransferase, a key enzyme in de novo pyrimidine synthesis affecting biofilm formation. In summary, our integrated chromatographic approach significantly advances PMI mapping.

Dipeptides are structurally diverse small molecules with varying modes of biogenesis and function. In plants, dipeptides were historically associated with nitrogen storage and mobilization; however, they are also reported to act as antioxidants, signaling molecules, protein regulators, and modulators of microbial communities. In this review, we discuss the structural diversity and biogenesis of dipeptides, with respect to (i) proteinogenic dipeptides that are products of protein degradation, (ii) non-proteinogenic amino acid dipeptides, such as those containing a γ-glutamyl group, and (iii) cyclic dipeptides, largely derived from microorganisms. Highlighted are recent examples of dipeptides that were shown to display plant health-promoting activities, including boosting growth and improving stress resilience against pathogens, salinity, chilling, and heat, making the case that these overlooked compounds are more than what meets the eye.

Drought stress substantially impacts crop physiology resulting in alteration of growth and productivity. Understanding the genetic and molecular crosstalk between stress responses and agronomically important traits such as fibre yield is particularly complicated in the allopolyploid species, upland cotton (Gossypium hirsutum), due to reduced sequence variability between A and D subgenomes. To better understand how drought stress impacts yield, the transcriptomes of 22 genetically and phenotypically diverse upland cotton accessions grown under well-watered and water-limited conditions in the Arizona low desert were sequenced. Gene co-expression analyses were performed, uncovering a group of stress response genes, in particular transcription factors GhDREB2A-A and GhHSFA6B-D, associated with improved yield under water-limited conditions in an ABA-independent manner. DNA affinity purification sequencing (DAP-seq), as well as public cistrome data from Arabidopsis, were used to identify targets of these two TFs. Among these targets were two lint yield-associated genes previously identified through genome-wide association studies (GWAS)-based approaches, GhABP-D and GhIPS1-A. Biochemical and phylogenetic approaches were used to determine that GhIPS1-A is positively regulated by GhHSFA6B-D, and that this regulatory mechanism is specific to Gossypium spp. containing the A (old world) genome. Finally, an SNP was identified within the GhHSFA6B-D binding site in GhIPS1-A that is positively associated with yield under water-limiting conditions. These data lay out a regulatory connection between abiotic stress and fibre yield in cotton that appears conserved in other systems such as Arabidopsis.

Methylerythritol cyclodiphosphate (MEcPP) is an intermediate in the biosynthesis of isoprenoids in plant plastids and in bacteria, and acts as a stress signal in plants. Here, we show that MEcPP regulates biofilm formation in Escherichia coli K-12 MG1655. Increased MEcPP levels, triggered by genetic manipulation or oxidative stress, inhibit biofilm development and production of fimbriae. Deletion of fimE, encoding a protein known to downregulate production of adhesive fimbriae, restores biofilm formation in cells with elevated MEcPP levels. Limited proteolysis-coupled mass spectrometry (LiP-MS) reveals that MEcPP interacts with the global regulatory protein H-NS, which is known to repress transcription of fimE. MEcPP prevents the binding of H-NS to the fimE promoter. Therefore, our results indicate that MEcPP can regulate biofilm formation by modulating H-NS activity and thus reducing fimbriae production. Further research is needed to test whether MEcPP plays similar regulatory roles in other bacteria. Methylerythritol cyclodiphosphate is an intermediate in the biosynthesis of isoprenoids in plants and bacteria, and acts as a stress signal in plants. Here, Guo et al. show that, in addition, the metabolite can inhibit biofilm formation in Escherichia coli by modulating the activity of the DNA-binding protein H-NS, thus downregulating the production of adhesive fimbriae.

ABSTRACT All sulfur transfer pathways have generally a l -cysteine desulfurase as an initial sulfur-mobilizing enzyme in common, which serves as a sulfur donor for the biosynthesis of numerous sulfur-containing biomolecules in the cell. In Escherichia coli , the housekeeping l -cysteine desulfurase IscS has several interaction partners, which bind at different sites of the protein. So far, the interaction sites of IscU, Fdx, CyaY, and IscX involved in iron-sulfur (Fe-S) cluster assembly have been mapped, in addition to TusA, which is required for molybdenum cofactor biosynthesis and mnm 5 s 2 U34 tRNA modifications, and ThiI, which is involved in thiamine biosynthesis and s 4 U8 tRNA modifications. Previous studies predicted that the sulfur acceptor proteins bind to IscS one at a time. E. coli TusA has, however, been suggested to be involved in Fe-S cluster assembly, as fewer Fe-S clusters were detected in a ∆ tusA mutant. The basis for this reduction in Fe-S cluster content is unknown. In this work, we investigated the role of TusA in iron-sulfur cluster assembly and iron homeostasis. We show that the absence of TusA reduces the translation of fur , thereby leading to pleiotropic cellular effects, which we dissect in detail in this study. IMPORTANCE Iron-sulfur clusters are evolutionarily ancient prosthetic groups. The ferric uptake regulator plays a major role in controlling the expression of iron homeostasis genes in bacteria. We show that a ∆tusA mutant is impaired in the assembly of Fe-S clusters and accumulates iron. TusA, therefore, reduces fur mRNA translation leading to pleiotropic cellular effects.

Plant homeodomain leucine zipper IV (HD-Zip IV) transcription factors (TFs) contain an evolutionarily conserved steroidogenic acute regulatory protein (StAR)-related lipid transfer (START) domain. While the START domain is required for TF activity, its presumed role as a lipid sensor is not clear. Here we used tandem affinity purification from Arabidopsis cell cultures to demonstrate that PROTODERMAL FACTOR2 (PDF2), a representative member that controls epidermal differentiation, recruits lysophosphatidylcholines (LysoPCs) in a START-dependent manner. Microscale thermophoresis assays confirmed that a missense mutation in a predicted ligand contact site reduces lysophospholipid binding. We additionally found that PDF2 acts as a transcriptional regulator of phospholipid- and phosphate (Pi) starvation-related genes and binds to a palindromic octamer with consensus to a Pi response element. Phospholipid homeostasis and elongation growth were altered in pdf2 mutants according to Pi availability. Cycloheximide chase experiments revealed a role for START in maintaining protein levels, and Pi starvation resulted in enhanced protein destabilization, suggesting a mechanism by which lipid binding controls TF activity. We propose that the START domain serves as a molecular sensor for membrane phospholipid status in the epidermis. Our data provide insights toward understanding how the lipid metabolome integrates Pi availability with gene expression.

Mitochondrial disease encompasses a group of genetically inherited disorders hallmarked by an inability of the respiratory chain to produce sufficient ATP. These disorders present with multisystemic pathologies that predominantly impact highly energetic tissues such as skeletal muscle. There is no cure or effective treatment for mitochondrial disease. We have discovered a small molecule known as oxybutynin that can bypass Complex III mitochondrial dysfunction in primary murine and human skeletal muscle progenitor cells (MPCs). Oxybutynin administration improves MPC proliferative capacity, enhances cellular glycolytic function, and improves myotube formation. Mechanistically, results from our isothermal shift assay indicates that oxybutynin interacts with a suite of proteins involved in mRNA processing which then trigger the upregulation biological pathways to circumvent CIII mitochondrial dysfunction. Taken together, we provide evidence for the small molecule oxybutynin as a potential therapeutic candidate for the future treatment of CIII mitochondrial dysfunction. ### Competing Interest Statement The authors have declared no competing interest.