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Low iron (Fe) bioavailability can limit the biosynthesis of Fe-containing proteins, which are especially abundant in photosynthetic organisms, thus negatively affecting global primary productivity. Understanding cellular coping mechanisms under Fe limitation is therefore of great interest. We surveyed the temporal responses of Chlamydomonas (Chlamydomonas reinhardtii) cells transitioning from an Fe-rich to an Fe-free medium to document their short and long-term adjustments. While slower growth, chlorosis and lower photosynthetic parameters are evident only after one or more days in Fe-free medium, the abundance of some transcripts, such as those for genes encoding transporters and enzymes involved in Fe assimilation, change within minutes, before changes in intracellular Fe content are noticeable, suggestive of a sensitive mechanism for sensing Fe. Promoter reporter constructs indicate a transcriptional component to this immediate primary response. With acetate provided as a source of reduced carbon, transcripts encoding respiratory components are maintained relative to transcripts encoding components of photosynthesis and tetrapyrrole biosynthesis, indicating metabolic prioritization of respiration over photosynthesis. In contrast to the loss of chlorophyll, carotenoid content is maintained under Fe limitation despite a decrease in the transcripts for carotenoid biosynthesis genes, indicating carotenoid stability. These changes occur more slowly, only after the intracellular Fe quota responds, indicating a phased response in Chlamydomonas, involving both primary and secondary responses during acclimation to poor Fe nutrition.

Fast growth phenotypes are achieved through optimal transcriptomic allocation, in which cells must balance tradeoffs in resource allocation between diverse functions. One such balance between stress readiness and unbridled growth in E. coli has been termed the fear versus greed (f/g) tradeoff. Two specific RNA polymerase (RNAP) mutations observed in adaptation to fast growth have been previously shown to affect the f/g tradeoff, suggesting that genetic adaptations may be primed to control f/g resource allocation. Here, we conduct a greatly expanded study of the genetic control of the f/g tradeoff across diverse conditions. We introduced 12 RNA polymerase (RNAP) mutations commonly acquired during adaptive laboratory evolution (ALE) and obtained expression profiles of each. We found that these single RNAP mutation strains resulted in large shifts in the f/g tradeoff primarily in the RpoS regulon and ribosomal genes, likely through modifying RNAP-DNA interactions. Two of these mutations additionally caused condition-specific transcriptional adaptations. While this tradeoff was previously characterized by the RpoS regulon and ribosomal expression, we find that the GAD regulon plays an important role in stress readiness and ppGpp in translation activity, expanding the scope of the tradeoff. A phylogenetic analysis found the greed-related genes of the tradeoff present in numerous bacterial species. The results suggest that the f/g tradeoff represents a general principle of transcriptome allocation in bacteria where small genetic changes can result in large phenotypic adaptations to growth conditions.IMPORTANCETo increase growth, E. coli must raise ribosomal content at the expense of non-growth functions. Previous studies have linked RNAP mutations to this transcriptional shift and increased growth but were focused on only two mutations found in the proteins central region. RNAP mutations, however, commonly occur over a large structural range. To explore RNAP mutations impact, we have introduced 12 RNAP mutations found in laboratory evolution experiments and obtained expression profiles of each. The mutations nearly universally increased growth rates by adjusting said tradeoff away from non-growth functions. In addition to this shift, a few caused condition-specific adaptations. We explored the prevalence of this tradeoff across phylogeny and found it to be a widespread and conserved trend among bacteria.

The high heritability of amyotrophic lateral sclerosis (ALS) contrasts with its low molecular diagnosis rate post-genetic testing, pointing to potential undiscovered genetic factors. To aid the exploration of these factors, we introduced EpiOut, an algorithm to identify chromatin accessibility outliers that are regions exhibiting divergent accessibility from the population baseline in a single or few samples. Annotation of accessible regions with histone chromatin immunoprecipitation sequencing and Hi-C indicates that outliers are concentrated in functional loci, especially among promoters interacting with active enhancers. Across different omics levels, outliers are robustly replicated, and chromatin accessibility outliers are reliable predictors of gene expression outliers and aberrant protein levels. When promoter accessibility does not align with gene expression, our results indicate that molecular aberrations are more likely to be linked to post-transcriptional regulation rather than transcriptional regulation. Our findings demonstrate that the outlier detection paradigm can uncover dysregulated regions in rare diseases. EpiOut is available at github.com/uci-cbcl/EpiOut.

Acinetobacter baumannii causes severe infections in humans, resists multiple antibiotics, and survives in stressful environmental conditions due to modulations of its complex transcriptional regulatory network (TRN). Unfortunately, our global understanding of the TRN in this emerging opportunistic pathogen is limited. Here, we apply independent component analysis, an unsupervised machine learning method, to a compendium of 139 RNA-seq data sets of three multidrug-resistant A. baumannii international clonal complex I strains (AB5075, AYE, and AB0057). This analysis allows us to define 49 independently modulated gene sets, which we call iModulons. Analysis of the identified A. baumannii iModulons reveals validating parallels to previously defined biological operons/regulons and provides a framework for defining unknown regulons. By utilizing the iModulons, we uncover potential mechanisms for a RpoS-independent general stress response, define global stress-virulence trade-offs, and identify conditions that may induce plasmid-borne multidrug resistance. The iModulons provide a model of the TRN that emphasizes the importance of transcriptional regulation of virulence phenotypes in A. baumannii. Furthermore, they suggest the possibility of future interventions to guide gene expression toward diminished pathogenic potential.IMPORTANCEThe rise in hospital outbreaks of multidrug-resistant Acinetobacter baumannii infections underscores the urgent need for alternatives to traditional broad-spectrum antibiotic therapies. The success of A. baumannii as a significant nosocomial pathogen is largely attributed to its ability to resist antibiotics and survive environmental stressors. However, there is limited literature available on the global, complex regulatory circuitry that shapes these phenotypes. Computational tools that can assist in the elucidation of A. baumanniis transcriptional regulatory network architecture can provide much-needed context for a comprehensive understanding of pathogenesis and virulence, as well as for the development of targeted therapies that modulate these pathways.

The Kaposi's sarcoma-associated herpesvirus (KSHV) genome consists of an approximately 140-kb unique coding region flanked by 30-40 copies of a 0.8-kb terminal repeat (TR) sequence. A gene enhancer recruits transcription-related enzymes by having arrays of transcription factor binding sites. Here, we show that KSHV TR possesses transcription regulatory function with latency-associated nuclear antigen (LANA). Cleavage under targets and release using nuclease demonstrated that TR fragments were occupied by LANA-interacting histone-modifying enzymes in naturally infected cells. The TR was enriched with histone H3K27 acetylation (H3K27Ac) and H3K4 tri-methylation (H3K4me3) modifications and also expressed nascent RNAs. The sites of H3K27Ac and H3K4me3 modifications were also conserved in the KSHV unique region among naturally infected primary effusion lymphoma cells. KSHV origin of lytic replication (Ori-Lyt) showed similar protein and histone modification occupancies with that of TR. In the Ori-Lyt region, the LANA and LANA-interacting proteins colocalized with an H3K27Ac-modified nucleosome along with paused RNA polymerase II. The KSHV transactivator KSHV replication and transcription activator (K-Rta) recruitment sites franked the LANA-bound nucleosome, and reactivation evicted the LANA-bound nucleosome. Including TR fragments in reporter plasmid enhanced inducible viral gene promoter activities independent of the orientations. In the presence of TR in reporter plasmids, K-Rta transactivation was drastically increased, while LANA acquired the promoter repression function. KSHV TR, therefore, functions as an enhancer for KSHV inducible genes. However, in contrast to cellular enhancers bound by multiple transcription factors, perhaps the KSHV enhancer is predominantly regulated by the LANA nuclear body.IMPORTANCEEnhancers are a crucial regulator of differential gene expression programs. Enhancers are the cis-regulatory sequences determining target genes' spatiotemporal and quantitative expression. Here, we show that Kaposi's sarcoma-associated herpesvirus (KSHV) terminal repeats fulfill the enhancer definition for KSHV inducible gene promoters. The KSHV enhancer is occupied by latency-associated nuclear antigen (LANA) and its interacting proteins, such as CHD4. Neighboring terminal repeat (TR) fragments to lytic gene promoters drastically enhanced KSHV replication and transcription activator and LANA transcription regulatory functions. This study, thus, proposes a new latency-lytic switch model in which TR accessibility to the KSHV gene promoters regulates viral inducible gene expression.

Hallmarks of the developmental cycle of the obligate intracellular pathogenic bacterium Chlamydia are the primary differentiation of the infectious elementary body (EB) into the proliferative reticulate body (RB) and the secondary differentiation of RBs back into EBs. The mechanisms regulating these transitions remain unclear. In this report, we developed an effective novel strategy termed dependence on plasmid-mediated expression (DOPE) that allows for the knockdown of essential genes in Chlamydia. We demonstrate that GrgA, a Chlamydia-specific transcription factor, is essential for the secondary differentiation and optimal growth of RBs. We also show that GrgA, a chromosome-encoded regulatory protein, controls the maintenance of the chlamydial virulence plasmid. Transcriptomic analysis further indicates that GrgA functions as a critical regulator of all three sigma factors that recognize different promoter sets at developmental stages. The DOPE strategy outlined here should provide a valuable tool for future studies examining chlamydial growth, development, and pathogenicity.

BackgroundGene expression may be regulated by the DNA methylation of regulatory elements in cis, distal, and trans regions. One method to evaluate the relationship between DNA methylation and gene expression is the mapping of expression quantitative trait methylation (eQTM) loci (also called expression associated CpG loci, eCpG). However, no open-source tools are available to provide eQTM mapping. In addition, eQTM mapping can involve a large number of comparisons which may prevent the analyses due to limitations of computational resources. Here, we describe Torch-eCpG, an open-source tool to perform eQTM mapping that includes an optimized implementation that can use the graphical processing unit (GPU) to reduce runtime.ResultsWe demonstrate the analyses using the tool are reproducible, up to 18 × faster using the GPU, and scale linearly with increasing methylation loci.ConclusionsTorch-eCpG is a fast, reliable, and scalable tool to perform eQTM mapping. Source code for Torch-eCpG is available at https://github.com/kordk/torch-ecpg .

SUMMARY: Cytokinin is central to coordinating plant adaptation to environmental stresses. Here, we first demonstrated the involvement of cytokinin in Arabidopsis responses to arsenite [As(III)] stress. As(III) treatment reduced cytokinin contents, while cytokinin treatment repressed further primary root growth in Arabidopsis plants under As(III) stress. Subsequently, we revealed that the cytokinin signaling members ARR1 and ARR12, the type‐B ARABIDOPSIS RESPONSE REGULATORs, participate in cytokinin signaling‐mediated As(III) responses in plants as negative regulators. A comprehensive transcriptome analysis of the arr1 and arr12 single and arr1,12 double mutants was then performed to decipher the cytokinin signaling‐mediated mechanisms underlying plant As(III) stress adaptation. Results revealed important roles for ARR1 and ARR12 in ion transport, nutrient responses, and secondary metabolite accumulation. Furthermore, using hierarchical clustering and regulatory network analyses, we identified two NODULIN 26‐LIKE INTRINSIC PROTEIN (NIP)‐encoding genes, NIP1;1 and NIP6;1, potentially involved in ARR1/12‐mediated As(III) uptake and transport in Arabidopsis. By analyzing various combinations of arr and nip mutants, including high‐order triple and quadruple mutants, we demonstrated that ARR1 and ARR12 redundantly function as negative regulators of As(III) tolerance by acting upstream of NIP1;1 and NIP6;1 to modulate their function in arsenic accumulation. ChIP–qPCR, EMSA, and transient dual‐LUC reporter assays revealed that ARR1 and ARR12 transcriptionally activate the expression of NIP1;1 and NIP6;1 by directly binding to their promoters and upregulating their expression, leading to increased arsenic accumulation under As(III) stress. These findings collectively provide insights into cytokinin signaling‐mediated plant adaptation to excessive As(III), contributing to the development of crops with low arsenic accumulation. Significance Statement: A genome‐wide transcriptome analysis of mutants defective in the ARR1 and/or ARR12 genes revealed that cytokinin signaling modulates As(III) responses in roots by regulating As(III) transport, stress responses, and secondary metabolism; furthermore, NIP1;1 and NIP6;1 were found to be involved in ARR1/12‐mediated arsenic accumulation in Arabidopsis, directly downstream of ARR1 and ARR12. Our study provides a strategy for manipulating cytokinin signaling to alter As(III) absorption and responses, enabling the development of crops with lower‐arsenic accumulation. [ABSTRACT FROM AUTHOR]

ABSTRACT When a species invades a novel environment, it must bridge the environment–phenotype mismatch in its new range to persist. Contemporary invasion biology research has focused on the role that trait variation and adaptation, and their underlying genomic factors, play in a species' adaptive potential, and thus facilitating invasion. Empirical studies have provided valuable insights into phenotypes that persist and arise in novel environments, coupled with ‘omics tools that further the understanding of the contributions of genomic architecture in species establishment. Particularly, the use of transcriptomics to explore the role of plasticity in the initial stages of an invasion is growing. Here, we assess the role of various mechanisms relating to regulation and functional adaptation (often measured via the transcriptome) that support trait‐specific plasticity in invasive species, allowing phenotypic variability without directly altering genomic diversity. First, we present a comprehensive review of the studies utilising transcriptomics in invasion biology. Second, we collate the evidence for and against the role of a range of regulatory processes in contributing to invasive species plasticity. Finally, we pose open questions in invasion biology where the use of transcriptome data may be valuable, as well as discuss the methodological limitations. [ABSTRACT FROM AUTHOR]

Background: Echinocandin B (ECB) is a key precursor of the antifungal drug anidulafungin and its biosynthesis occurs via ani gene cluster in Aspergillus nidulans NRRL8112. Strain improvement for industrial ECB production has mainly relied on mutation breeding due to the lack of genetic tools. Results: Here, a CRISPR-base-editing tool was developed in A. nidulans NRRL8112 for simultaneous inactivation of the nkuA gene and two marker genes, pryoA and riboB, which enabled efficient genetic manipulation. Then, in-vivo plasmid assembly was harnessed for ani gene expression screening, identifying the rate-limiting enzyme AniA and a pathway-specific transcription factor AniJ. Stepwise titer enhancement was achieved by overexpressing aniA and/or aniJ, and ECB production reached 1.5 g/L during 5-L fed-batch fermentation, an increase of ~ 30-fold compared with the parent strain. Conclusion: This study, for the first time, revealed the regulatory mechanism of ECB biosynthesis and harnessed genetic engineering for the development of an efficient ECB-producing strain. [ABSTRACT FROM AUTHOR]

Precise regulation of gene expression is crucial to development. Enhancers, the core of gene regulation, determine the spatiotemporal pattern of gene transcription. Since many disease-associated mutations are characterized in enhancers, the research on enhancer will provide clues to precise medicine. Rapid advances in high-throughput sequencing technology facilitate the characterization of enhancers at genome wide, but understanding the functional mechanisms of enhancers remains challenging. Herein, we provide a panorama of enhancer characteristics, including epigenetic modifications, enhancer transcripts, and enhancer-promoter interaction patterns. Furthermore, we outline the applications of high-throughput sequencing technology and functional genomics methods in enhancer research. Finally, we discuss the role of enhancers in human disease and their potential as targets for disease prevention and treatment strategies. [ABSTRACT FROM AUTHOR]

SUMMARY Calmodulin7 (CAM7) is a key transcription factor of Arabidopsis seedling development. CAM7 works together with HY5 bZIP protein to promote photomorphogenesis at various wavelengths of light. In this study, we show that AtMYB4, identified from a yeast two‐hybrid screen, physically interacts with CAM7 and works as a positive regulator of photomorphogenesis at various wavelengths of light. CAM7 and HY5 directly bind to the promoter of AtMYB4 to promote its expression for photomorphogenic growth. On the other hand, ARA4, identified from the same yeast two‐hybrid screen, works as a negative regulator of photomorphogenic growth specifically in white light. The double mutant analysis reveals that the altered hypocotyl elongation of atmyb4 and ara4 is either partly or completely suppressed by additional loss of function of CAM7. Furthermore, ARA4 genetically interacts with AtMYB4 in an antagonistic manner to suppress the elongated hypocotyl phenotype of atmyb4. The transactivation studies reveal that while CAM7 activates the promoter of AtMYB4 in association with HY5, ARA4 negatively regulates AtMYB4 expression. Taken together, these results demonstrate that working as a negative regulator of photomorphogenesis, ARA4 plays a balancing act on CAM7 and HY5‐mediated regulation of AtMYB4. [ABSTRACT FROM AUTHOR]

Background: De-novo-designed synthetic transcriptional regulators have great potential as the genetic parts for constructing complex multilayered gene circuits. The design flexibility afforded by advanced nucleic acid sequence design tools vastly expands the repertoire of regulatory elements for circuit design. In principle, the design space of synthetic regulators should allow for the construction of regulatory circuits of arbitrary complexity; still, the orthogonality and robustness of such components have not been fully elucidated, thereby limiting the depth and width of synthetic circuits. Results: In this work, we systematically explored the design strategy of synthetic transcriptional regulators, termed switchable transcription terminators. Specifically, by redesigning key sequence domains, we created a high-performance switchable transcription terminator with a maximum fold change of 283.11 upon activation by its cognate input RNA. Further, an automated design algorithm was developed for these elements to improve orthogonality for a complex multi-layered circuit construction. The resulting orthogonal switchable transcription terminators could be used to construct a three-layer cascade circuit and a two-input three-layer OR gate. Conclusions: We demonstrated a practical strategy for designing standardized regulatory elements and assembling modular gene circuits, ultimately laying the foundation for the streamlined construction of complex synthetic gene circuits. [ABSTRACT FROM AUTHOR]

Cotton fiber length is basically determined by well‐coordinated gene expression and phosphatidylinositol phosphates (PIPs) accumulation during fiber elongation but the regulatory mechanism governing PIPs transport remains unknown. Here, we report a MYB transcription factor GhMYB30D04 in Gossypium hirsutum that promotes fiber elongation through modulating the expression of PIP transporter gene GhLTPG1. Knockout of GhMYB30D04 gene in cotton (KO) results in a reduction of GhLTPG1 transcripts with lower accumulation of PIPs, leading to shorter fibers and lower fiber yield. Conversely, GhMYB30D04 overexpression (GhMYB30D04‐OE) causes richer PIPs and longer cotton fibers, mimicking the effects of exogenously applying PIPs on the ovules of GhMYB30D04‐KO and wild type. Furthermore, GhMYB30D04 interacts with GhHD1, the crucial transcription factor of fiber initiation, to form an activation complex stabilized by PIPs, both of which upregulate GhLTPG1 expression. Comparative omics‐analysis revealed that higher and extended expressions of LTPG1 in fiber elongation mainly correlate with the variations of the GhMYB30D04 gene between two cotton allotetraploids, contributing to longer fiber in G. babardense. Our work clarifies a mechanism by which GhHD1–GhMYB30D04 form a regulatory module of fiber elongation to tightly control PIP accumulation. Our work still has an implication that GhMYB30D04–GhHD1 associates with development transition from fiber initiation to elongation. [ABSTRACT FROM AUTHOR]

Metastasis suppressor 1 (MTSS1) has been reported to play important roles in suppressing cancer progression. In this study, we investigated the underlying mechanism that regulates MTSS1 expression. We showed that in breast cancer cells, lncRNA-SNHG15-induced cell invasion and proliferation was accompanied with the decreased expression of MTSS1 mRNA. Further study revealed that SNHG15 mediated MTSS1 repression through blocking its promoter activity. Mechanistically, SNHG15 complexes with DDX5 and RTF1 and interacts with the core promoter of the MTSS1 gene to interfere with RNA-Pol-II-directed transcriptional initiation. Association with DDX5 stabilizes SNHG15 while binding to RTF1 allows SNHG15 to carry RTF1 to the core promoter, where RTF1 forms a complex with PNA pl II to enhance transcriptional pausing. Our findings revealed a molecular mechanism by which SNHG15 serves as a regulator to suppresses MTSS1 transcription via interaction with the gene core promoter. [ABSTRACT FROM AUTHOR]

SUMMARY: Cold stress restricts the metabolic and physiological activities of plants, thereby affecting their growth and development. Although broad‐complex, tramtrack, and bric‐à‐brac (BTB) proteins are essential for diverse biological processes and stress responses, the mechanisms underlying BTB‐mediated cold responses remain not fully understood. Here, we characterize the function of the cold‐induced SlBTB19 protein in tomato (Solanum lycopersicum). Overexpression of SlBTB19 resulted in increased plant sensitivity to cold stress, whereas SlBTB19 knockout mutants exhibited a cold‐tolerance phenotype. Further analyses, including protein–protein interaction studies and cell‐free degradation assays, revealed that SlBTB19 interacts with and destabilizes the transcription factor SlWRKY2. Using virus‐induced gene silencing (VIGS) to silence SlWRKY2 in both wild‐type and slbtb19 mutants, we provided genetic evidence that SlWRKY2 acts downstream of SlBTB19 in regulating cold tolerance. Importantly, we demonstrated that SlWRKY2 positively regulates cold tolerance in a CRT/DRE binding factor (CBF)‐dependent manner. Under cold stress, SlWRKY2 binds to the W‐box in the CBF1 and CBF3 promoters, directly activating their expression. In summary, our findings identify a SlBTB19‐SlWRKY2 module that negatively regulates the CBF‐dependent cold tolerance pathway in tomato. Significance Statement: Our findings not only contribute to the broader understanding of the molecular basis of plant responses to cold stress but also highlight the importance of post‐transcriptional mechanisms in regulating these responses. In summary, we proposed an updated model: A cold‐induced BTB family protein SlBTB19 negatively regulates cold tolerance. SlBTB19 interacts with and facilitates the proteolysis of the SlWRKY2 protein. SlWRKY2 is a positive regulator of cold tolerance, which activates cold‐responsive genes SlCBFs. [ABSTRACT FROM AUTHOR]

The TRIM (tripartite motif) family, with TRIM28 as a key member, plays a vital role in regulating health and disease. TRIM28 contains various functional domains essential for transcriptional regulation, primarily through its interaction with KRAB‐ZNF proteins, which influence chromatin remodeling and gene expression. Despite extensive research, the precise mechanisms by which TRIM28 impacts health and disease remain elusive. This review delves into TRIM28's multifaceted roles in maintaining health, contributing to a variety of diseases, and influencing cancer progression. In cancers, TRIM28 exhibits a dual nature, functioning as both a tumor promoter and suppressor depending on the cellular context and cancer type. The review also explores its critical involvement in processes such as DNA repair, cell cycle regulation, epithelial‐to‐mesenchymal transition, and the maintenance of stem cell properties. By uncovering TRIM28's complex roles across different cancers and other diseases, this review underscores its potential as a therapeutic target. The significance of TRIM28 as a versatile regulator opens the door to innovative therapeutic strategies, particularly in cancer treatment and the management of other diseases. Ongoing research into TRIM28 may yield key insights into disease progression and novel treatment options. [ABSTRACT FROM AUTHOR]

Thyroid hormones and their receptors (TRs) play critical roles during vertebrate development. One of the most dramatic developmental processes regulated by thyroid hormones is frog metamorphosis, which mimics the postembryonic (perinatal) period in mammals. Here, we review some of the findings on the developmental functions of thyroid hormones and TRs as well as their associated mechanisms of action obtained from this model system. More than 2 decades ago, a dual function model was proposed for TR in anuran development. During larval development, unliganded receptors recruit corepressors to repress thyroid hormone response genes to prevent premature metamorphic changes. Subsequently, when thyroid hormone levels rise, liganded receptors recruit coactivators to activate thyroid hormone response genes, leading to metamorphic changes. Over the years, molecular and genetic approaches have provided strong support for this model and have shown that it is applicable to mammalian development as well as to understanding the diverse effects of thyroid hormones in normal physiology and diseases caused by thyroid hormone signaling dysfunction. [ABSTRACT FROM AUTHOR]

The intestinal epithelium harbours a unique lymphocyte population, the intraepithelial lymphocytes (IELs). A large fraction of IELs is represented by γδ T cells. Their role in epithelial homeostasis and immune response is well documented, but a conclusive view of their developmental pathway is still missing. In this review, we discuss the existing literature as well as recent advances regarding the tissue adaptation of γδ IELs, both for the characteristic cytotoxic subset and the newly described noncytotoxic subset. We particularly highlight the environmental cues and the transcriptional regulation that equip γδ T cells with their IEL phenotype. [ABSTRACT FROM AUTHOR]

Hypoxia inducible transcription factors (HIFs) mediate the hypoxic response in metazoans. When sufficient O2 is present, Fe(II)/2-oxoglutarate (2OG)-dependent oxygenases (human PHD1-3) promote HIFα degradation via prolyl-hydroxylation. We report crystallographic, spectroscopic, and biochemical characterization of stable and inactive PHD2.Fe(III).2OG complexes. Aerobic incubation of PHD2 with Fe(II) and 2OG enables formation of PHD2.Fe(III).2OG complexes which bind HIF1-2α to give inactive PHD2.Fe(III).2OG.HIF1-2α complexes. The Fe(III) oxidation state in the inactive complexes was shown by EPR spectroscopy. L-Ascorbate hinders formation of the PHD2.Fe(III).2OG.(+/-HIFα) complexes and slowly regenerates them to give the catalytically active PHD2.Fe(II).2OG complex. Crystallographic comparison of the PHD2.Fe(III).2OG.HIF2α complex with the analogous anaerobic Fe(II) complex reveals near identical structures. Exposure of the anaerobic PHD2.Fe(II).2OG.HIF2α crystals to O2 enables in crystallo hydroxylation. The resulting PHD2.product structure, manifests conformational changes compared to the substrate structures. The results have implications for the role of the PHDs in hypoxia sensing and open new opportunities for inhibition of the PHDs and other 2OG dependent oxygenases by promoting formation of stable Fe(III) complexes. [ABSTRACT FROM AUTHOR]

SUMMARY Land plants have evolved a hydrophobic cuticle on the surface of aerial organs as an adaptation to ensure survival in terrestrial environments. Cuticle is mainly composed of lipids, namely cutin and intracuticular wax, with epicuticular wax deposited on plant surface. The composition and permeability of cuticle have a large influence on its ability to protect plants against drought stress. However, the regulatory mechanisms underlying cuticular wax biosynthesis in response to drought stress have not been fully elucidated. Here, we identified three AP2/ERF transcription factors (DREB26/ERF12, ERF13 and ERF14) involved in the regulation of water permeability of the plant surface. Transmission electron microscopy revealed thicker cuticle on the leaves of DREB26‐overexpressing (DREB26OX) plants, and thinner cuticle on the leaves of transgenic plants expressing SRDX repression domain‐fused DREB26 (DREB26SR). Genes involved in cuticular wax formation were upregulated in DREB26OX and downregulated in DREB26SR. The levels of very‐long chain (VLC) alkanes, which are a major wax component, increased in DREB26OX leaves and decreased in DREB26SR leaves. Under dehydration stress, water loss was reduced in DREB26OX and increased in DREB26SR. The erf12/13/14 triple mutant showed delayed growth, decreased leaf water content, and reduced drought‐inducible VLC alkane accumulation. Taken together, our results indicate that the DREB26/ERF12 and its closed family members, ERF13 and ERF14, play an important role in cuticular wax biosynthesis in response to drought stress. The complex transcriptional cascade involved in the regulation of cuticular wax biosynthesis under drought stress conditions is discussed. [ABSTRACT FROM AUTHOR]

Optical regulation of transcription using chemical compounds is an effective strategy to manipulate gene expression spatiotemporally. Conventional caging approaches with photoremovable protecting groups may require intense UV‐light exposure and release potentially toxic byproducts. To address these problems, here we developed a light‐mediated transcriptional regulation system by combining a caging‐group‐free photoactivatable dye PaX560 and a multidrug‐binding transcriptional regulator QacR. The cationic dye generated from PaX560 through traceless photoconversion bound QacR and reduced its repressor function, resulting in transcriptional activation. Importantly, this system allowed transcriptional activation with a large dynamic range under mild visible light exposure and simultaneous detection of the state of the photoactivated effector. This module was integrated into the T7 RNA polymerase expression system to demonstrate light‐activated transcription in vitro and in living cells. [ABSTRACT FROM AUTHOR]

SUMMARY Fiber elongation rate is an essential characteristic of cotton fiber in the textile industry, yet it has been largely overlooked in genetic studies. Gibberellins (GAs) and auxin (IAA) are recognized for their role in directing numerous developmental processes in plants by influencing cell differentiation and elongation. However, the degree to which GA–IAA interaction governs cellular elongation in cotton fiber cells remains to be fully understood. In this study, we identified a causal gene, Gibberellic Acid‐Stimulated in Arabidopsis 24 (GhGASA24), that appears to be responsible for fiber elongation rate via regulating fiber cell wall thickness. Subsequent experiments revealed that GhGASA24 influences cell wall formation by promoting the expression of GhCesA8 and GhCesA10. Our findings suggest that Auxin Response Factor 2 (GhARF2) regulates fiber elongation rate by directly binding to the AuxRE elements in GhGASA24 promoter. In addition, we identified Growth Regulation Factor 4 (GhGRF4) as a transcription factor that interacts with GhARF2 to form a heterodimer complex, which also transcriptionally activates GhGASA24. Intriguingly, GhGRF4 regulates GhARF2 expression by directly binding to its promoter, thereby acting as a cascade regulator to enhance the transcriptional levels of GhGASA24. We propose that the GhGRF4/GhARF2‐GhGASA24‐GhCesAs module may contribute to fiber cell wall thickness by modulating cellulose biosynthesis, and provide a theoretical basis for improvement of fiber quality. [ABSTRACT FROM AUTHOR]

Retinoic acid is crucial for vertebrate embryogenesis, influencing anterior‐posterior patterning and organogenesis through its interaction with nuclear hormone receptors comprising heterodimers of retinoic acid receptors (RARα, β, or γ) and retinoid X receptors (RXRα, β, or γ). Tissue retinoic acid levels are tightly regulated since both its excess and deficiency are deleterious. Dehydrogenase/reductase 3 (DHRS3) plays a critical role in this regulation by converting retinaldehyde to retinol, preventing excessive retinoic acid formation. Mutations in DHRS3 can result in embryonic lethality and congenital defects. This study shows that mouse Dhrs3 expression is responsive to vitamin A status and is directly regulated by the RAR/RXR complex through cis‐regulatory elements. This highlights a negative feedback mechanism that ensures retinoic acid homeostasis. [ABSTRACT FROM AUTHOR]

Goat milk is abundant in nutrients, particularly in milk fats, which confer health benefits to humans. Exploring the regulatory mechanism of fatty acid synthesis is highly important to understand milk composition manipulation. In this study, we used chromatin immunoprecipitation sequencing (ChIP-seq) on goat mammary glands at different lactation stages which revealed a novel lactation regulatory factor: cell death-inducing DFFA-like effector B (CIDEB). RT-qPCR results revealed that CIDEB was significantly upregulated during lactation in dairy goats. CIDEB overexpression significantly increased the expression levels of genes involved in fatty acid synthesis (ACACA, SCD1, p < 0.05; ELOVL6, p < 0.01), lipid droplet formation (XDH, p < 0.05), and triacylglycerol (TAG) synthesis (DGAT1, p < 0.05; GPAM, p < 0.01) in goat mammary epithelial cells (GMECs). The contents of lipid droplets, TAG, and cholesterol were increased (p < 0.05) in CIDEB-overexpressing GMECs, and knockdown of CIDEB led to the opposite results. In addition, CIDEB knockdown significantly decreased the proportion of C16:0 and total C18:2. Luciferase reporter assays indicated that X-box binding protein 1 (XBP1) promoted CIDEB transcription via XBP1 binding sites located in the CIDEB promoter. Furthermore, CIDEB knockdown attenuated the stimulatory effect of XBP1 on lipid droplet accumulation. Collectively, these findings elucidate the critical regulatory roles of CIDEB in milk fat synthesis, thus providing new insights into improving the quality of goat milk. [ABSTRACT FROM AUTHOR]

SUMMARY: Geraniol is one of the most abundant aromatic compounds in fresh tea leaves and contributes to the pleasant odor of tea products. Additionally, it functions as an airborne signal that interacts with other members of the ecosystem. To date, the regulation of the geraniol biosynthesis in tea plants remains to be investigated. In this study, a correlation test of the content of geraniol and its glycosides with gene expression data revealed that nudix hydrolase, CsNudix26, and its transcription factor, CsbHLH133 are involved in geraniol biosynthesis. In vitro enzyme assays and metabolic analyses of genetically modified tea plants confirmed that CsNudix26 is responsible for the formation of geraniol. Yeast one‐hybrid, dual‐luciferase reporter, and EMSA assays were used to verify the binding of CsbHLH133 to the CsNudix26 promoter. Overexpression of CsbHLH133 in tea leaves enhanced CsNudix26 expression and geraniol accumulation, whereas CsbHLH133 silencing reduced CsNudix26 transcript levels and geraniol content. Interestingly, CsbHLH133‐AS, produced by alternative splicing, was discovered and proved to be the primary transcript expressed in response to various environmental stresses. Furthermore, geraniol release was found to be affected by various factors that alter the expression patterns of CsbHLH133 and CsbHLH133‐AS. Our findings indicate that distinct transcript splicing patterns of CsbHLH133 regulate geraniol biosynthesis in tea plants in response to different regulatory factors. Significance Statement: Geraniol is one of the most abundant aromatic compounds in fresh tea leaves and contributes to the pleasant odor of tea products. The transcription factor CsbHLH133 regulates geraniol biosynthesis by activating the expression of CsNudix26 in tea plants. Our results delineate the transcriptional regulatory network involved in geraniol biosynthesis in Camellia sinensis. Alternative splicing serves as a mechanism for plant stress tolerance, secondary metabolism, and developmental plasticity. Changes in the transcription of CsbHLH133 through alternative splicing modulate its abundance and translation efficiency, enabling tea plants to adapt better to adverse environmental conditions. [ABSTRACT FROM AUTHOR]

Rapeseed (Brassica napus L.) is a major oilseed crop in the middle and lower reaches of the Yangtze River in China. However, it is susceptible to waterlogging stress. This study aimed to investigate the physiological characteristics, cellular changes, and gene expression patterns of rapeseed under waterlogging stress, with the goal of providing a foundation for breeding waterlogging-tolerant rapeseed. The results revealed that waterlogging-tolerant rapeseed exhibited higher levels of soluble sugars and antioxidant enzyme activity, particularly in the roots. Conversely, waterlogging-sensitive rapeseed displayed greater changes in malondialdehyde, proline, and hydrogen peroxide levels. Cellular observations showed that after experiencing waterlogging stress, the intercellular space of rapeseed leaf cells expanded, leading to disintegration of mitochondria and chloroplasts. Moreover, the area of the root xylem increased, the number of vessels grew, and there were signs of mitochondrial disintegration and vacuole shrinkage, with more pronounced changes observed in waterlogging-sensitive rapeseed. Furthermore, significant differences were found in the transcription levels of genes related to anaerobic respiration and flavonoid biosynthesis, and different varieties demonstrated varied responses to waterlogging stress. In conclusion, there are differences in the response of different varieties to waterlogging stress at the levels of morphology, physiological characteristics, cell structure, and gene transcription. Waterlogging-tolerant rapeseed responds to waterlogging stress by regulating its antioxidant defense system. This study provides valuable insights for the development of waterlogging-tolerant rapeseed varieties. [ABSTRACT FROM AUTHOR]

Yellowhorn (Xanthoceras sorbifolium Bunge) is an economically important tree species in northern China, mainly distributed in arid and semi-arid areas where water resources are scarce. Drought affects its yield and the expansion of its suitable growth area. It was found that the wax content in yellowhorn leaves varied significantly among different germplasms, which had a strong correlation with the drought resistance of yellowhorn. In this study, XsMYB30 was isolated from 'Zhongshi 4' of yellowhorn, a new highly waxy variety. DAP-Seq technology revealed that the pathways associated with fatty acids were significantly enriched in the target genes of XsMYB30. Moreover, the results of electrophoretic mobility shift assay, yeast one hybrid assay and dual-luciferase assay demonstrated that XsMYB30 could directly and specifically bind with the promoters of genes involved in wax biosynthesis (XsFAR4 , XsCER1 and XsKCS1), lipid transfer (XsLTPG1 and XsLTP1) and fatty acid synthesis (XsKASIII), thus enhancing their expression. In addition, the overexpression of XsMYB30 in poplar promoted the expression levels of these target genes and increased the wax deposition on poplar leaves leading to a notable improvement in the plant's ability to withstand drought. These findings indicate that XsMYB30 is an important regulatory factor in cuticular wax biosynthesis and the drought resistance of yellowhorn. [ABSTRACT FROM AUTHOR]

Summary: Transcriptional regulation mechanisms underlying chilling injury (CI) development have been widely investigated in model plants and cold‐sensitive fruits, such as banana (Musa acuminata). However, unlike the well‐known NAC and WRKY transcription factors (TFs), the function and deciphering mechanism of heat shock factors (HSFs) involving in cold response are still fragmented. Here, we showed that hot water treatment (HWT) alleviated CI in harvested banana fruits accomplishing with reduced reactive oxygen species (ROS) accumulation and increased antioxidant enzyme activities. A cold‐inducible but HWT‐inhibited HSF, MaHsf24, was identified. Using DNA affinity purification sequencing (DAP‐seq) combined with RNA‐seq analyses, we found three heat shock protein (HSP) genes (MaHSP23.6, MaHSP70‐1.1 and MaHSP70‐1.2) and three antioxidant enzyme genes (MaAPX1, MaMDAR4 and MaGSTZ1) were the potential targets of MaHsf24. Subsequent electrophoretic mobility shift assay (EMSA), chromatin immunoprecipitation coupled with quantitative PCR (ChIP‐qPCR) and dual‐luciferase reporter (DLR) analyses demonstrated that MaHsf24 repressed the transcription of these six targets via directly binding to their promoters. Moreover, stably overexpressing MaHsf24 in tomatoes increased cold sensitivity by suppressing the expressions of HSPs and antioxidant enzyme genes, while HWT could recover cold tolerance, maintaining higher levels of HSPs and antioxidant enzyme genes, and activities of antioxidant enzymes. In contrast, transiently silencing MaHsf24 by virus‐induced gene silencing (VIGS) in banana peels conferred cold resistance with the upregulation of MaHSPs and antioxidant enzyme genes. Collectively, our findings support the negative role of MaHsf24 in cold tolerance, and unravel a novel regulatory network controlling bananas CI occurrence, concerning MaHsf24‐exerted inhibition of MaHSPs and antioxidant enzyme genes. [ABSTRACT FROM AUTHOR]

Drought is a primary abiotic stress that inhibits rice (Oryza sativa L.) growth and development, and during the reproductive stage it has a negative impact on the rice seed-setting rate. This research study examined two rice lines, La-96 (drought sensitive) and La-163 (drought resistant), for drought stress treatment (with soil moisture at 20% for 7 days) and control (normal irrigation and kept soil moisture ≥40%). To elucidate the photosynthesis and molecular mechanisms underlying drought tolerance in rice, leaf photosynthetic traits and transcriptome sequencing were used to compare differences between two contrasting recombinant inbred lines (RIL) during drought and subsequent recovery at the booting stage. The rice line La-96 showed a significant decrease in seed-setting rate after being treated for seven days' drought stress (from 86.64% to 22.75%), while La-163 was slightly affected (from 89.04% to 79.33%). The photosynthetic activities of both lines significantly decreased under the drought treatment, and these traits of La-163 recovered to a comparable level with the control after three days of rewatering. The transcriptome of both lines in three treatments (the control, drought stress, and subsequent recovery) were tested, and a total of 16,051 genes were identified, among which 10,566 genes were differentially expressed in various treatments and rice lines. Comprehensive gene expression profiles revealed that the specifically identified DEGs were involved in the ribosome synthesis and the metabolic pathway of photosynthesis, starch, and sucrose metabolism. The DEGs that are activated and respond quickly, as seen during recovery in the tolerant rice line, may play essential roles in regulating subsequent growth and development. This study uncovered the molecular genetic pathways of drought tolerance and extended our understanding of the drought tolerance mechanisms and subsequent recovery regulation in rice. [ABSTRACT FROM AUTHOR]

Lactose assimilation is a relatively rare trait in yeasts, and Kluyveromyces yeast species have long served as model organisms for studying lactose metabolism. Meanwhile, the metabolic strategies of most other lactose-assimilating yeasts remain unknown. In this work, we have elucidated the genetic determinants of the superior lactose-growing yeast Candida intermedia. Through genomic and transcriptomic analyses, we identified three interdependent gene clusters responsible for the metabolism of lactose and its hydrolysis product galactose: the conserved LAC cluster (LAC12, LAC4) for lactose uptake and hydrolysis, the conserved GAL cluster (GAL1, GAL7, and GAL10) for galactose catabolism through the Leloir pathway, and a "GALLAC" cluster containing the transcriptional activator gene LAC9, second copies of GAL1 and GAL10, and a XYL1 gene encoding an aldose reductase involved in carbon overflow metabolism. Bioinformatic analysis suggests that the GALLAC cluster is unique to C. intermedia and has evolved through gene duplication and divergence, and deletion mutant phenotyping proved that the cluster is indispensable for C. intermedia's growth on lactose and galactose. We also show that the regulatory network in C. intermedia, governed by Lac9 and Gal1 from the GALLAC cluster, differs significantly from the galactose and lactose regulons in Saccharomyces cerevisiae, Kluyveromyces lactis, and Candida albicans. Moreover, although lactose and galactose metabolism are closely linked in C. intermedia, our results also point to important regulatory differences. IMPORTANCE This study paves the way to a better understanding of lactose and galactose metabolism in the non-conventional yeast C. intermedia. Notably, the unique GALLAC cluster represents a new, interesting example of metabolic network rewiring and likely helps to explain how C. intermedia has evolved into an efficient lactose-assimilating yeast. With the Leloir pathway of budding yeasts acting like a model system for understanding the function, evolution, and regulation of eukaryotic metabolism, this work provides new evolutionary insights into yeast metabolic pathways and regulatory networks. In extension, the results will facilitate future development and use of C. intermedia as a cell-factory for conversion of lactose-rich whey into value-added products. [ABSTRACT FROM AUTHOR]

Complementation remains a foundation for demonstrating molecular Koch's postulates. While this is frequently achieved using plasmids, limitations such as increased gene copy number and the need for antibiotic supplementation to avoid plasmid loss can restrict their use. Chromosomal integration systems using the Tn7 transposon provide an alternative to plasmids for complementation and facilitate the stable insertion of genes at the chromosomal attTn7 site without the need for selection pressure. Here, we enhanced the utility of mini-Tn7 insertion vectors by the addition of inducible (Pcym) and constitutive (PcL and PrpsM) promoters, allowing different ial transcriptional control of genes integrated into the chromosome. We validated the utility of these promoters by cloning the gfp gene, encoding green fluorescent protein, downstream of each promoter and integrating a mini-Tn7 construct harboring these elements into the attTn7 site on the chromosome of the Escherichia coli K-12 strain MG1655. The PcL and PrpsM promoters provided equivalent levels of GFP expression and offered flexibility based on the target host strain. Activation of the tightly regulated Pcym promoter with its inducer cumate resulted in tunable expression of GFP in a dose-dependent manner. We further demonstrated the tight control of the Pcym promoter using the toxic impCAB genes, and the expression of which is detrimental to E. coli viability. Together, these modified mini-Tn7 vectors allowing differential control of genes integrated into the chromosome at a conserved site offer an efficient system for complementation where plasmid use is restricted. IMPORTANCE Chromosomal integration using mini-Tn7 vectors provides an efficient means to insert genes into the chromosome of many gram-negative bacteria. Insertion occurs at a conserved site and allows for the stable integration of genes in single copy. While this system has multiple benefits for enabling complementation, a cornerstone for fulfilling molecular Koch's postulates, greater flexibility for controlled gene expression would enhance its utility. Here, we have added to the function of mini-Tn7 vectors by the addition of inducible and constitutive promoters and demonstrated their capacity to drive the controlled expression of target genes integrated into the chromosome. In addition to complementation, these modified vectors offer broad application for other approaches including chromosomal tagging, in vivo expression, metabolic engineering, and synthetic biology. [ABSTRACT FROM AUTHOR]

Background and aims: Myofibroblasts, the primary producers of extracellular matrix, primarily originate from hepatic stellate cells (HSCs), and their activation plays a pivotal role in liver fibrosis. This study aimed to investigate the function of CXC motif ligand 14 (CXCL14) in the progression of liver fibrosis. Approach and results: CXCL14 knockdown significantly reduced the extent of liver fibrosis. Using Ingenuity pathway analysis and qRT‐PCR, activating transcription factor 3 (ATF3) was identified as a key upstream regulator of CXCL14 expression. Mechanistically, ATF3 was shown to bind to the CXCL14 promoter, promoting its transactivation by TGF‐β in HSCs. Notably, both global CXCL14 deletion (CXCL14−/−) and HSC/myofibroblast‐specific CXCL14 knockdown significantly attenuated liver fibrosis in mice. RNA‐seq comparisons between CXCL14−/− and WT mice highlighted Jak2 as the most significantly downregulated gene, implicating its role in the antifibrotic effects of CXCL14 suppression on HSC inactivation. Moreover, Jak2 overexpression reversed the inhibition of liver fibrosis caused by CXCL14 knockdown in vivo. Conclusions: These findings unveil a novel ATF3/CXCL14/Jak2 signalling axis in liver fibrosis, presenting potential therapeutic targets for the disease. [ABSTRACT FROM AUTHOR]

Skeletal muscle plasticity enables an enormous potential to adapt to various internal and external stimuli and perturbations. Most notably, changes in contractile activity evoke a massive remodeling of biochemical, metabolic and force-generating properties. In recent years, a large number of signals, sensors, regulators and effectors have been implicated in these adaptive processes. Nevertheless, our understanding of the molecular underpinnings of training adaptation remains rudimentary. Specifically, the mechanisms that underlie signal integration, output coordination, functional redundancy and other complex traits of muscle adaptation are unknown. In fact, it is even unclear how stimulus-dependent specification is brought about in endurance or resistance exercise. In this review, we will provide an overview on the events that describe the acute perturbations in single endurance and resistance exercise bouts. Furthermore, we will provide insights into the molecular principles of long-term training adaptation. Finally, current gaps in knowledge will be identified, and strategies for a multi-omic and –cellular analyses of the molecular mechanisms of skeletal muscle plasticity that are engaged in individual, acute exercise bouts and chronic training adaptation discussed. [Display omitted] • Exercise training elicits massive molecular adaptations in skeletal muscle. • The response to an acute exercise bout differs from long-term training adaptation. • Untrained and trained muscles respond differently to an exercise bout. • The expression of few genes is persistently changed in trained muscle at rest. • The mechanisms of specification of endurance and resistance training are mysterious. [ABSTRACT FROM AUTHOR]

Infectious laryngotracheitis virus (ILTV) exhibits a cascade expression pattern of encoded genes, and ICP4 is the only immediate-early gene of ILTV, which plays a crucial role in initiating the subsequent viral genes. Therefore, studying the transcriptional regulation mechanism of ICP4 holds promise for effectively blocking ILTV infection and spread. Host transcriptional factors p53 and Fos are proven to regulate a variety of viral infections, and our previous studies have demonstrated their synergistic effects in regulating ILTV infection. In this study, we constructed eukaryotic expression vectors for p53 and Fos as well as their specific siRNAs and transfected them into a chicken hepatoma cell line. The results showed that knocking down p53 or Fos significantly inhibited ICP4 transcription, while overexpressing p53 or Fos had an opposite effect. A further CoIP and ChIP-qPCR assay suggested p53 and Fos physically interacted with each other, and jointly bound to the upstream transcriptional regulatory region of ICP4. To elucidate the specific mechanisms of p53 and Fos in regulating ICP4 transcription, we designed p53 and Fos protein mutants by mutating their DNA binding domains, which significantly reduced their binding ability to DNA without affecting their interaction. The results showed that Fos directly bound to the promoter region of ICP4 as a binding target of p53, and the p53–Fos protein complex acted as a transcriptional co-regulator of ICP4. Studying the transcriptional process and regulatory pattern of ICP4 is of great significance for understanding the molecular mechanism of ILTV infection, and thus for finding effective methods to control and prevent it. [ABSTRACT FROM AUTHOR]

Background: The role of Mediator complex subunit 1 (MED1), a pivotal transcriptional coactivator implicated in diverse biological pathways, remains unexplored in the context of oral squamous cell carcinoma (OSCC). This study aims to elucidate the contributory mechanisms and potential impact of MED1 on the progression of OSCC. Methods: The expression and clinical significance of MED1 in OSCC tissues were evaluated through the bioinformatics analyses. The effects of MED1 on the biological behavior of OSCC cancer cells were assessed both in vitro and in vivo. Dual-luciferase reporter assay, chromatin immunoprecipitation (ChIP) assay, bioinformatic analysis, CD8+ T cell isolation experiment, coculture experiment, enzyme-linked immunosorbent assay (ELISA), and flow cytometric analysis were employed to elucidate the underlying mechanism through which MED1 operates in the progression of OSCC. Results: MED1 exhibited upregulation in both OSCC tissues and multiple OSCC cell lines, which correlated with decreased overall survival in patients. In vitro experiments demonstrated that knockdown of MED1 in metastatic OSCC cell lines SCC-9 and UPCI-SCC-154 hindered cell migration and invasion, while overexpression of MED1 promoted these processes. Whereas, MED1 knockdown had no impact on proliferation of cell lines mentioned above. In vivo studies further revealed that downregulation of MED1 effectively suppressed distant metastasis in OSCC. Mechanistically, MED1 enhanced the binding of transcription factors c-Jun and c-Fos to the matrix metalloprotein 9 (MMP9) promoters, resulting in a significant upregulation of MMP9 transcription. This process contributes to the migration and invasion of SCC-9 and UPCI-SCC-154 cells. Furthermore, MED1 modulated the expression of programmed death-ligand 1 (PD-L1) through the Notch signaling pathway, consequently impacting the tumor-killing capacity of CD8+ T cells in the tumor microenvironment. Conclusions: Our findings indicate that MED1 plays a pivotal role in OSCC progression through the activation of MMP9 transcription and suppression of CD8+ T cell antitumor immunity, suggesting that MED1 may serve as a novel prognostic marker and therapeutic target in OSCC. [ABSTRACT FROM AUTHOR]

Renal tubular epithelial cells (TECs) undergo an energy‐related metabolic shift from fatty acid oxidation to glycolysis during chronic kidney disease (CKD) progression. However, the mechanisms underlying this burst of glycolysis remain unclear. Herein, a new critical glycolysis regulator, the transcription factor forkhead box protein K1 (FOXK1) that is expressed in TECs during renal fibrosis and exhibits fibrogenic and metabolism‐rewiring capacities is reported. Genetic modification of the Foxk1 locus in TECs alters glycolytic metabolism and fibrotic lesions. A surge in the expression of a set of glycolysis‐related genes following FOXK1 protein activation contributes to the energy‐related metabolic shift. Nuclear‐translocated FOXK1 forms condensate through liquid‐liquid phase separation (LLPS) to drive the transcription of target genes. Core intrinsically disordered regions within FOXK1 protein are mapped and validated. A therapeutic strategy is explored by targeting the Foxk1 locus in a murine model of CKD by the renal subcapsular injection of a recombinant adeno‐associated virus 9 vector encoding Foxk1‐short hairpin RNA. In summary, the mechanism of a FOXK1‐mediated glycolytic burst in TECs, which involves the LLPS to enhance FOXK1 transcriptional activity is elucidated. [ABSTRACT FROM AUTHOR]

Heparan sulfate proteoglycans (HSPGs) regulate a wide range of biological activities in both physiological and pathological conditions. Altered expression or deregulated function of HSPGs and their heparan sulfate (HS) chains significantly contribute to carcinogenesis as well and crucially depends on the functioning of the complex system of HS biosynthetic/modifying enzymes termed as "GAGosome". Here, we aimed at investigating the expression profile of the system in a cell culture model of stroma-epithelial crosstalk and searching for transcription factors potentially related to the regulation of expression of the genes involved. Coculture of BjTERT-fibroblasts with normal PNT2 human prostate epithelial cells resulted in significant downregulation (2-4-fold) of transcriptional activity of HS metabolism-involved genes (EXT1/2, NDST1/2, GLCE, HS2ST1, HS3ST1/2, HS6ST1/2, SULF1/2, HPSE) in both cell types, whereas coculture with prostate cancer cells (LNCaP, PC3, DU145) demonstrated no significant interchanges. Human Transcription Factor RT2 Profiler PCR array and manual RT-PCR verification supposed FOS, MYC, E2F, SRF, NR3C1 as potential candidates for regulation and/or coordination of HS biosynthesis. Taken together, transcriptional activity of HS biosynthetic system in normal fibroblasts and prostate epithelial cells during their coculture might be controlled by their intercellular communication, reflecting of adaptation of these cells to each other. The regulation is attenuated or abrogated if normal fibroblasts interact with prostate cancer cells making the cancer cells independent of the limiting effects of fibroblasts, thus contributing to possibility of unlimited growth and progression. Overall, these data demonstrate an ability of cell-cell interactions to affect transcriptional activity of HS biosynthesis-involved genes. [ABSTRACT FROM AUTHOR]

Cellular differentiation allows cells to transition between different functional states and adapt to various environmental cues. The diversity and plasticity of this process is beautifully exemplified by T cells responding to pathogens, which undergo highly specialized differentiation tailored to the ongoing infection. Such antigen‐induced T cell differentiation is regulated at the transcriptional level by DNA‐binding proteins and at the post‐transcriptional level by RNA‐binding proteins. Although traditionally defined as separate protein classes, a growing body of evidence indicates an overlap between these two groups of proteins, collectively coined DNA/RNA‐binding proteins (DRBPs). In this review, we describe how DRBPs might bind both DNA and RNA, discuss the putative functional relevance of this dual binding, and provide an exploratory analysis into characteristics that are associated with DRBPs. To exemplify the significance of DRBPs in T cell biology, we detail the activity of several established and putative DRBPs during the T cell response. Finally, we highlight several methodologies that allow untangling of the distinct functionalities of DRBPs at the DNA and RNA level, including key considerations to take into account when applying such methods. [ABSTRACT FROM AUTHOR]

SUMMARY: The bilateral‐to‐radial symmetry transition occurring during the development of the Arabidopsis thaliana female reproductive organ (gynoecium) is a crucial biological process linked to plant fertilization and seed production. Despite its significance, the cellular mechanisms governing the establishment and breaking of radial symmetry at the gynoecium apex (style) remain unknown. To fill this gap, we employed quantitative confocal imaging coupled with MorphoGraphX analysis, in vivo and in vitro transcriptional experiments, and genetic analysis encompassing mutants in two bHLH transcription factors necessary and sufficient to promote transition to radial symmetry, SPATULA (SPT) and INDEHISCENT (IND). Here, we show that defects in style morphogenesis correlate with defects in cell‐division orientation and rate. We showed that the SPT‐mediated accumulation of auxin in the medial‐apical cells undergoing symmetry transition is required to maintain cell‐division‐oriented perpendicular to the direction of organ growth (anticlinal, transversal cell division). In addition, SPT and IND promote the expression of specific core cell‐cycle regulators, CYCLIN‐D1;1 (CYC‐D1;1) and CYC‐D3;3, to support progression through the G1 phase of the cell cycle. This transcriptional regulation is repressed by auxin, thus forming an incoherent feed‐forward loop mechanism. We propose that this mechanism fine‐tunes cell division rate and orientation with the morphogenic signal provided by auxin, during patterning of radial symmetry at the style. Significance Statement: Our work reveals the cellular basis underpinning a bilateral‐to‐radial symmetry transition occurring during the development of the Arabidopsis female reproductive organ. To promote the medial‐apical fusion of the developing carpels, two bHLH transcription factors, SPATULA and INDEHISCENT, orchestrate auxin accumulation in specific cells to maintain anticlinal cell‐division orientation and fine‐tune progression through the G1 cell‐cycle phase via an Incoherent type‐I Feed‐Forward Loop (I‐IFFL). [ABSTRACT FROM AUTHOR]

SUMMARY: Pear is a widely cultivated fruit crop, but its distribution and sustainable production are significantly limited by salt stress. This study used RNA‐Seq time‐course analysis, WGCNA, and functional enrichment analysis to uncover the molecular mechanisms underlying salt stress tolerance in Pyrus ussuriensis. We identified an ABA‐related regulatory module, PbGBF3‐PbAPL2‐PbSDH1, as crucial in this response. PbGBF3, a bZIP transcription factor, enhances salt tolerance by upregulating PbAPL2 and PbSDH1. Overexpression of PbGBF3 improved salt tolerance in Pyrus communis calli and Arabidopsis, while silencing it reduced tolerance in Pyrus betulifolia. Functional assays showed that PbGBF3 binds to the promoters of PbAPL2 and PbSDH1, increasing their expression. PbAPL2 and PbSDH1, key enzymes in starch synthesis and the sorbitol pathway, respectively, enhance salt tolerance by increasing AGPase activity, soluble sugar content, and SDH activity, improving ROS scavenging and ion balance. Our findings suggest that the PbGBF3‐PbAPL2 and PbGBF3‐PbSDH1 modules positively regulate salt tolerance by enhancing ABA signaling and reducing ABA‐mediated growth inhibition. These insights provide a foundation for developing salt‐tolerant pear cultivars. [ABSTRACT FROM AUTHOR]

Epilepsy is a neurological disease characterised by recurrent seizures with complex aetiology. Temporal lobe epilepsy, the most common form in adults, can be acquired following brain insults including trauma, stroke, infection or sustained status epilepticus. The mechanisms that give rise to the formation and maintenance of hyperexcitable networks following acquired insults remain unknown, yet an extensive body of literature points towards persistent gene and epigenomic dysregulation as a potential mediator of this dysfunction. While much is known about the function of specific classes of epigenetic regulators (writers and erasers) in epilepsy, much less is known about the enzymes, which read the epigenome and modulate gene expression accordingly. Here, we explore the potential role for the epigenetic reader bromodomain and extra‐terminal domain (BET) proteins in epilepsy. Using the intra‐amygdala kainic acid model of temporal lobe epilepsy, we initially identified widespread dysregulation of important epigenetic regulators including EZH2 and REST as well as altered BRD4 expression in chronically epileptic mice. BRD4 activity was also notably affected by epilepsy‐provoking insults as seen by elevated binding to and transcriptional regulation of the immediate early gene Fos. Despite influencing early aspects of epileptogenesis, blocking BET protein activity with JQ1 had no overt effects on epilepsy development in mice but did alter glial reactivity and influence gene expression patterns, promoting various neurotransmitter signalling mechanisms and inflammatory pathways in the hippocampus. Together, these results confirm that epigenetic reader activity is affected by epilepsy‐provoking brain insults and that BET activity may exert cell‐specific actions on inflammation in epilepsy. [ABSTRACT FROM AUTHOR]

Sirtuin 1 (SIRT1) is a key upstream regulator of lipid metabolism; however, the molecular mechanisms by which SIRT1 regulates milk fat synthesis in dairy goats remain unclear. This study aimed to investigate the regulatory roles of SIRT1 in modulating lipid metabolism in goat mammary epithelial cells (GMECs) and its impact on the adipose triglyceride lipase (ATGL) promoter activity using RNA interference (RNAi) and gene overexpression techniques. The results showed that SIRT1 is significantly upregulated during lactation compared to the dry period. Additionally, SIRT1 knockdown notably increased the expressions of genes related to fatty acid synthesis (SREBP1, SCD1, FASN, ELOVL6), triacylglycerol (TAG) production (DGAT2, AGPAT6), and lipid droplet formation (PLIN2). Consistent with the transcriptional changes, SIRT1 knockdown significantly increased the intracellular contents of TAG and cholesterol and the lipid droplet abundance in the GMECs, while SIRT1 overexpression had the opposite effects. Furthermore, the co-overexpression of SIRT1 and Forkhead box protein O1 (FOXO1) led to a more pronounced increase in ATGL promoter activity, and the ability of SIRT1 to enhance ATGL promoter activity was nearly abolished when the FOXO1 binding sites (FKH1 and FKH2) were mutated, indicating that SIRT1 enhances the transcriptional activity of ATGL via the FKH element in the ATGL promoter. Collectively, our data reveal that SIRT1 enhances the transcriptional activity of ATGL through the FOXO1 binding sites located in the ATGL promoter, thereby regulating lipid metabolism. These findings provide novel insights into the role of SIRT1 in fatty acid metabolism in dairy goats. [ABSTRACT FROM AUTHOR]

The transcription factor WRKYs play pivotal roles in the adapting to adverse environments in plants. Prior research has demonstrated the involvement of OsWRKY70 in resistance against herbivores and its response to abiotic stress. Here, we reported the functional analysis of OsWRKY70 in immunity against fungal diseases and cold tolerance. The results revealed that OsWRKY70 was induced by various Magnaporthe oryzae strains. Knock out mutants of OsWRKY70, which were generated by the CRISPR/Cas9 system, exhibited enhanced resistance to M. oryzae. This was consistent with fortifying the reactive oxygen species (ROS) burst after inoculation in the mutants, elevated transcript levels of defense-responsive genes (OsPR1b, OsPBZ1, OsPOX8.1 and OsPOX22.3) and the observation of the sluggish growth of invasive hyphae under fluorescence microscope. RNA sequencing (RNA-seq) and quantitative real-time PCR (qRT-PCR) validations demonstrated that differentially expressed genes were related to plant-pathogen interactions, hormone transduction and MAPK cascades. Notably, OsbHLH6, a key component of the JA signaling pathway, was down-regulated in the mutants compared to wild type plants. Further investigation confirmed that OsWRKY70 bound to the promoter of OsbHLH6 by semi-in vivo chromatin immunoprecipitation (ChIP). Additionally, the loss-function of OsWRKY70 impaired cold tolerance in rice. The enhanced susceptibility in the mutants characterized by excessive ROS production, elevated ion leakage rate and increased malondialdehyde content, as well as decreased activity of catalase (CAT) and peroxidase (POD) under low temperature stress was, which might be attributed to down-regulation of cold-responsive genes (OsLti6b and OsICE1). In conclusion, our findings indicate that OsWRKY70 negatively contributes to blast resistance but positively regulates cold tolerance in rice, providing a strategy for crop breeding with tolerance to stress. [ABSTRACT FROM AUTHOR]

Food-gene interaction has been identified as a leading risk factor for inflammatory bowel disease (IBD) and colorectal cancer (CRC). Accordingly, nutrigenomics emerges as a new approach to identify biomarkers and therapeutic targets for these two strongly associated gastrointestinal diseases. Recent studies in stem cell biology have further shown that diet and nutrition signal to intestinal stem cells (ISC) by altering nutrient-sensing transcriptional activities, thereby influencing barrier integrity and susceptibility to inflammation and tumorigenesis. This review recognizes the dietary factors related to both CRC and IBD and investigates their impact on the overlapping transcription factors governing stem cell activities in homeostasis and post-injury responses. Our objective is to provide a framework to study the food-gene regulatory network of disease-contributing cells and inspire new nutrigenomic approaches for detecting and treating diet-related IBD and CRC. [ABSTRACT FROM AUTHOR]

RhizoBindingSites is a de novo depurified database of conserved DNA motifs potentially involved in the transcriptional regulation of the Rhizobium, Sinorhizobium, Bradyrhizobium, Azorhizobium, and Mesorhizobium genera covering 9 representative symbiotic species, deduced from the upstream regulatory sequences of orthologous genes (O-matrices) from the Rhizobiales taxon. The sites collected with O-matrices per gene per genome from RhizoBindingSites were used to deduce matrices using the dyad-Regulatory Sequence Analysis Tool (RSAT) method, giving rise to novel S-matrices for the construction of the RizoBindingSites v2.0 database. A comparison of the S-matrix logos showed a greater frequency and/or re-definition of specific-position nucleotides found in the O-matrices. Moreover, S-matrices were better at detecting genes in the genome, and there was a more significant number of transcription factors (TFs) in the vicinity than O-matrices, corresponding to a more significant genomic coverage for S-matrices. O-matrices of 3187 TFs and S-matrices of 2754 TFs from 9 species were deposited in RhizoBindingSites and RhizoBindingSites v2.0, respectively. The homology between the matrices of TFs from a genome showed inter-regulation between the clustered TFs. In addition, matrices of AraC, ArsR, GntR, and LysR ortholog TFs showed different motifs, suggesting distinct regulation. Benchmarking showed 72%, 68%, and 81% of common genes per regulon for O-matrices and approximately 14% less common genes with S-matrices of Rhizobium etli CFN42, Rhizobium leguminosarum bv. viciae 3841, and Sinorhizobium meliloti 1021. These data were deposited in RhizoBindingSites and the RhizoBindingSites v2.0 database (http://rhizobindingsites.ccg.unam.mx/). [ABSTRACT FROM AUTHOR]

Background: Alternative splicing is a pivotal mechanism of post-transcriptional modification that contributes to the transcriptome plasticity and proteome diversity in metazoan cells. Although many splicing regulations around the exon/intron regions are known, the relationship between promoter-bound transcription factors and the downstream alternative splicing largely remains unexplored. Results: In this study, we present computational approaches to unravel the regulatory relationship between promoter-bound transcription factor binding sites (TFBSs) and the splicing patterns. We curated a fine dataset that includes DNase I hypersensitive site sequencing and transcriptomes across fifteen human tissues from ENCODE. Specifically, we proposed different representations of TF binding context and splicing patterns to examine the associations between the promoter and downstream splicing events. While machine learning models demonstrated potential in predicting splicing patterns based on TFBS occupancies, the limitations in the generalization of predicting the splicing forms of singleton genes across diverse tissues was observed with carefully examination using different cross-validation methods. We further investigated the association between alterations in individual TFBS at promoters and shifts in exon splicing efficiency. Our results demonstrate that the convolutional neural network (CNN) models, trained on TF binding changes in the promoters, can predict the changes in splicing patterns. Furthermore, a systemic in silico substitutions analysis on the CNN models highlighted several potential splicing regulators. Notably, using empirical validation using K562 CTCFL shRNA knock-down data, we showed the significant role of CTCFL in splicing regulation. Conclusion: In conclusion, our finding highlights the potential role of promoter-bound TFBSs in influencing the regulation of downstream splicing patterns and provides insights for discovering alternative splicing regulations. [ABSTRACT FROM AUTHOR]

Phosphorus is an essential macronutrient for plant growth and development. In response to phosphate (Pi) deficiency, plants rapidly produce a substitutive amount of root hairs; however, the mechanisms underlying Pi supply for root hair growth remain unclear. Here, we observed that soybean (Glycine max) plants maintain a consistent level of Pi within root hairs even under external Pi deficiency. We therefore investigated the role of vacuole‐stored Pi, a major Pi reservoir in plant cells, in supporting root hair growth under Pi‐deficient conditions. Our findings indicated that two vacuolar Pi efflux (VPE) transporters, GmVPE1 and GmVPE2, remobilize vacuolar stored Pi to sustain cytosolic Pi content in root hair cells. Genetic analysis showed that double mutants of GmVPE1 and GmVPE2 exhibited reduced root hair growth under low Pi conditions. Moreover, GmVPE1 and GmVPE2 were highly expressed in root hairs, with their expression levels significantly upregulated by low Pi treatment. Further analysis revealed that GmRSL2 (ROOT HAIR DEFECTIVE 6‐like 2), a transcription factor involved in root hair morphogenesis, directly binds to the promoter regions of GmVPE1 and GmVPE2, and promotes their expressions under low Pi conditions. Additionally, mutants lacking both GmRSL2 and its homolog GmRSL3 exhibited impaired root hair growth under low Pi stress, which was rescued by overexpressing either GmVPE1 or GmVPE2. Taken together, our study has identified a module comprising vacuolar Pi exporters and transcription factors responsible for remobilizing vacuolar Pi to support root hair growth in response to Pi deficiency in soybean. [ABSTRACT FROM AUTHOR]

Cellular senescence refers to the stable state of cell cycle arrest in which cells lose the ability of division and proliferation. Various intracellular and extracellular stimuli can induce cell senescence. Senescent cells exhibit multiple hallmarks, such as the upregulation of cell cycle inhibitor proteins p16INK4a and p21Cipl, DNA damage responses, and structural and metabolic alterations. Another major hallmark of senescent cells is that they express and secret a variety of factors, including cytokines, chemokines, growth factors, proteases, and other bioactive molecules, defined as the senescence- associated secretory phenotype (SASP). SASP factors exert multiple biological functions through the cell autonomous autocrine manner or cell non-autonomous paracrine fashion. In this review, we summarize the composition of SASP, and point out its high heterogeneity and dynamics. We then summarize the regulatory mechanisms of SASP at various levels including transcription, post-transcription, translation, post-translational modifications, and epigenetics. Afterwards, we summarize the various biological functions of SASP, including its beneficial effects in tumor suppression, tissue repair, and embryonic development, as well as its detrimental effects in inducing cell senescence, promoting tumor occurrence and development, age-related diseases, and organismal aging. We further discuss the potential applications of the SASP, which overview the senolytic therapy for selectively clearing senescent cells and senomorphic therapy for inhibiting SASP to intervene age and age-related diseases. Finally, we outline several challenges in identifying and detecting senescent cells and SASP factors in vivo, and provide some practical recommendations and new techniques to address these challenges. [ABSTRACT FROM AUTHOR]

The LysR-type transcriptional regulators (LTTRs) are DNA-binding proteins present in bacteria, archaea, and in algae. Knowledge about their distribution, abundance, evolution, structural organization, transcriptional regulation, fundamental roles in free life, pathogenesis, and bacteria–plant interaction has been generated. This review focuses on these aspects and provides a current picture of LTTR biology. [ABSTRACT FROM AUTHOR]