Search Results (956)

N-acetylserotonin methyltransferase (ASMT) is among the key enzymes involved in the final steps of melatonin biosynthesis. Here, we have shown that inactivation of ASMT in A. thaliana results in reduced endogenous melatonin levels, modulating other plant hormone pathways and affecting stress-related responses. Transcriptomic analysis of the asmt-null mutant revealed that the differentially expressed genes were predominantly enriched in terms associated with auxin responses and signalling, as well as with abscisic acid (ABA)-mediated stress responses. In addition, the expression of genes involved in the ethylene, salicylic acid, jasmonic acid and brassinosteroid pathways was altered in the mutant. Assays of a β-glucuronidase (GUS) construct in which a fragment containing 1000 bp upstream of the ASMT start codon was fused to the GUS reporter gene confirmed that ASMT is involved in the responses to ABA, gibberellic and indole acetic acids, trans-zeatin, ethylene and epibrassinolide, which is consistent with the results of the in silico analysis of the ASMT promoter. Furthermore, the expression of a number of genes, such as SLG1, HIS1-3, AtAIRP1 and several LEA genes, whose transcriptional regulation is associated with water management and contributes to impaired tolerance to dehydration stress, was altered in the mutant. The pleiotropic effects of ASMT gene disruption facilitate the identification of new potential melatonin targets and provide insights into the specific mechanisms of melatonin action. Full article

Strigolactones (SLs), as a class of novel plant hormones, play important roles in the regulation of plant branching. However, their function in branch development of wintersweet remains unclear. In this study, a gene involved in SLs biosynthesis, CpD27, was identified and isolated from wintersweet. The sequence characteristics, expression patterns, subcellular localization, and functional analysis through heterologous expression in Arabidopsis thaliana were investigated. Multiple sequence alignment showed that CpD27 contains the conserved D27 protein domain DUF4033. Quantitative real-time PCR analysis revealed that CpD27 is expressed in various vegetative organs of wintersweet, with the highest expression in leaves, followed by axillary buds. It is also expressed in all floral organs, with the highest expression level in the outer petals. CpD27 expression is induced by hormones (ABA and ACC) and low temperature (4 °C). Subcellular localization analysis indicated that CpD27 is localized in the chloroplasts of Arabidopsis. Heterologous expression of CpD27 in Arabidopsis delayed bolting. The number of both rosette branches and cauline branches in transgenic plants was reduced compared with wild-type plants. In addition, the expression of AtBRC1 was significantly upregulated in transgenic lines, suggesting that CpD27 has a function similar to that of its homolog in Arabidopsis. Overall, these results indicate that CpD27 plays a conserved role in the SLs-mediated branching pathway, which regulates branch development in wintersweet. This study provides a molecular and theoretical basis for further understanding branch development in wintersweet. Full article

Nuclear factor erythroid 2-related factor 2 (NRF2) has traditionally been framed as a Kelch-like ECH-associated protein 1 (KEAP1)-regulated stress-response transcription factor, but three observations now require a broader framework: NRF2 turnover is controlled by parallel E3 ligase systems; transcriptional output can be limited by coactivator assembly despite unchanged NRF2 abundance; and NRF2 activation can be beneficial or harmful depending on disease context, as illustrated by lung cancer models in which NRF2 paradoxically promotes metastasis through BTB and CNC homology 1 (BACH1) stabilization. We synthesize these observations into an NRF2 partner-code framework in which NRF2 acts as a context-dependent transcriptional platform assembled through four partly independent modules: a degradation module (KEAP1; β-transducin repeat-containing protein, β-TrCP; HMG-CoA reductase degradation protein 1/synoviolin 1, Hrd1/SYVN1; WD repeat-containing protein 23/DDB1- and CUL4-associated factor 11, WDR23/DCAF11); a cytoplasmic scaffold module (p62/sequestosome 1, p62/SQSTM1; IQ motif-containing GTPase-activating protein 1, IQGAP1; type I phosphatidylinositol 4-phosphate 5-kinase γ/heat shock protein 27, PIPKIγ–HSP27; peptidyl-prolyl cis-trans isomerase NIMA-interacting 1, PIN1; peptidyl-prolyl isomerase A/cyclophilin A, PPIA); a nuclear coactivator module at Neh4/5 (CREB-binding protein/p300, CBP/p300; receptor-associated coactivator 3/steroid receptor coactivator 3, RAC3/SRC-3; protein arginine methyltransferase 1/coactivator-associated arginine methyltransferase 1, PRMT1/CARM1; Mediator complex subunit 16, MED16); and a DNA/chromatin module at Neh1 (small musculoaponeurotic fibrosarcoma [Maf] proteins, BACH1, and chromodomain helicase DNA-binding protein 6, CHD6). Mapping 22 partners onto the Neh-domain architecture identifies approximately 25 pharmacologically addressable interfaces, stratified into four translational tiers. The framework reframes NRF2 pharmacology around one principle: the most actionable target is often a partner rather than NRF2 itself, with disease context dictating the direction of modulation. We close with five testable hypotheses and a partner-code decision matrix linking disease, biomarker, and candidate target. Full article

Spermatogenesis is a highly coordinated biological process in which diploid spermatogonia undergo mitotic expansion, meiotic division, and terminal differentiation into haploid spermatozoa. This process is tightly regulated by intrinsic germ cell programs and extrinsic signals from Sertoli cells within the seminiferous epithelium. Among the signaling pathways governing male germ cell development, all-trans retinoic acid (RA), a bioactive metabolite of vitamin A, has emerged as a master regulator of meiotic initiation and spermatogonial differentiation in mammals. RA functions through nuclear retinoic acid receptors (RARs) and retinoid X receptors (RXRs), which regulate transcriptional networks essential for germ cell progression, including the activation of Stimulated by Retinoic Acid 8 (STRA8), a key determinant of meiotic entry. Intratesticular RA homeostasis is maintained by a balance between synthesis via aldehyde dehydrogenase (ALDH) enzymes and degradation by cytochrome P450 family 26 (CYP26) enzymes, ensuring precise temporal and spatial control of germ cell development. While rodent models have defined core mechanisms of RA signaling, the canine testis provides a valuable comparative and translational system due to its physiological similarity to human spermatogenesis and relevance to reproductive management. Recent studies highlight conserved RA signaling pathways in dogs, including receptor-mediated transcriptional regulation, feedback control of RA metabolism, and post-transcriptional modulation via microRNAs. Importantly, pharmacological manipulation of RA signaling can reversibly disrupt spermatogenesis, supporting its potential applications in non-hormonal male contraception. This review integrates molecular, developmental, pharmacological, and comparative evidence and presents RA signaling as a central regulatory axis of spermatogenesis with important translational applications. Full article

Background/Objectives: Barley malting is an agro-industrial process that produces malt, an essential ingredient for the brewing and distilling industries. Previously, tran-scriptome profiling has revealed mRNA changes during malting but less is known about their regulation. Methods: The spring 2-row barley variety ‘Conrad’ was sampled at five stages of malt-ing. Using small RNA (sRNA)-sequencing and degradome-sequencing data from these malting stages, de novo discovery of mature microRNA (miRNA), as well as cognate mRNAs targeted for slicing, was identified. ShortStack v4.1.0 was used to map sRNA reads to the Hordeum vulgare Morex V3 genome. Results: In total, 33 expressed MIRs were identified, six of which may be novel. Using the degradome-sequencing data from the same malting stages, CleaveLand4 v4.5 pre-dicted 64 sliced mRNA targets, predominantly transcription factors associated with root development. Conclusions: This study provides an overview of post-transcriptional modulations of miRNAs-cognate mRNA targets, as well as plausible interactions between miRNAs during barley malting. Full article

Caveolin-1 is a scaffolding protein of caveolae, flask-shaped membrane microdomains involved in diverse cellular processes. Caveolae are primarily localized to the plasma membrane, the trans-Golgi network, and mitochondria-associated endoplasmic reticulum (ER) membranes (MAMs). Most enzymes involved in cholesterol biosynthesis reside in the ER, and although caveolin-1 avidly binds cholesterol, its role in cholesterol trafficking remains unclear. Acyl-coenzyme A:cholesterol acyltransferases (ACAT1 and ACAT2) convert free cholesterol into cholesteryl esters for storage, with ACAT1 serving as the predominant isoenzyme in most cell types. ACAT1 is an ER-resident protein, with a fraction associated with specialized ER subdomains, including the MAM. Here, we report that a subset of caveolin-1 molecules appears to be associated with a fraction of ACAT1 in ER subdomains. Using immunoprecipitation under detergent conditions, immunoadsorption of MAM-enriched membranes under detergent-free conditions, and electron microscopy, we provide evidence consistent with an association between a subset of caveolin-1 molecules and ACAT1. Functionally, in mouse embryonic fibroblasts, we show that genetic ablation of caveolin-1 significantly increases the esterification of low-density lipoprotein-derived cholesterol, suggesting that caveolin-1 may attenuate ACAT1 activity. Collectively, these findings indicate that caveolin-1 may modulate cholesterol esterification and contribute to the regulation of cholesterol distribution among cellular membranes. Full article

Newborn piglets in intensive pig farms are often prone to pulmonary diseases due to underdeveloped fetal lung neural systems, and ATRA is often regarded as an important morphogen that displays pleiotropic functions during embryonic development. However, information about the effect of maternal administration with ATRA on the peripheral neural system of fetal lungs is still scare. Fifteen pregnant sows were assigned to the ATRA0 (0 mg/kg diet), ATRA4, ATRA8, ATRA16 and ATRA32 treatment groups and offered their own feed from d12 to d95 after artificial insemination; then two neonatal pigs with birth weights similar to the average birth weight were taken out from each litter for the collection of lung samples, and samples were subjected to immunofluorescence staining, RNA-seq and RT-qPCR assays. Results indicated that compared to newborn piglets from the ATRA0 treatment group, newborn piglets from the ATRA4 treatment group had higher percentages of GFAP-positive astrocyte cells (p < 0.05) and GFAP-TUBB3 colocalization (p < 0.05) in the lungs. KEGG (Kyoto Encyclopedia of Genes and Genomes) analysis showed that differential expressed genes are mainly enriched in the pathways of neuroactive ligand–receptor interaction, GABAergic synapses and cell adhesion molecules. Addition of ATRA at 4 mg/kg to the diet of pregnant sows can enhance the healthy development of the pulmonary nervous systems of fetal pigs. Full article

All-trans retinoic acid (atRA) is known to regulate lipid metabolism, adipocyte differentiation, and the immune system in mammals and other aquatic species. However, studies on atRA in crustaceans, especially in Eriocheir sinensis, are still scarce. The present study aimed to investigate the regulatory effects of dietary or injected atRA on female crabs during the fattening period. In the dietary regulation experiment, 270 female crabs were fed diets containing different doses of atRA (0, 150, 300, 600, 1200, and 2400 mg/kg) for a total of 49 days. In the in vivo injection experiment, 90 females were divided into an experimental group (injected with a 0.3 μg/g dose of atRA) and a control group (injected with the same amount of DMSO solvent). Injections were given weekly throughout the 35-day experimental period. Results: Both dietary atRA and atRA injection promoted ovarian development, as evidenced by increased GSI, elevated serum Vg levels, decreased GIH, and upregulated expression of vg, vgr, and rxr genes. In terms of mechanism, dietary atRA promoted ovarian development via the upregulation of pyrimidine nucleotides and dehydroepiandrosterone, which supplied nucleic acid precursors and hormonal support. Furthermore, RXR was identified as a potential key target of atRA in inducing ovarian development, as molecular docking revealed that atRA could spontaneously bind to RXR. Moreover, following atRA injection, the expression of rxr, along with key genes involved in ovarian development, lipid synthesis, and lipid transport, was significantly upregulated. In addition, the atRA diet created a favorable microenvironment for ovarian development by reducing pro-inflammatory lipid levels in the ovary. Transcriptomic and metabolomic analyses revealed that atRA modulates energy and lipid metabolism by activating the AMPK pathway. In terms of the bacterial community structure, the atRA diet significantly decreased Fusobacterium abundance and enriched Parabacteroides as the signature beneficial bacterium. In terms of nutritional quality, the atRA diet markedly reduced saturated and trans-fatty acids while increasing monounsaturated fatty acids and various key essential amino acids. Conclusions: This study revealed that atRA plays a key role in promoting ovarian development, improving nutritional quality, and modulating the structure of the microbiota, thereby providing theoretical support for healthy aquaculture of female crabs during the fattening period. Full article

Cell culture is foundational to biomedical advancements, yet its widespread clinical and practical distribution is severely constrained by the high infrastructural costs of cryogenic logistics and the physical stressors of liquid-phase transit. Herein, we propose a proof-of-concept cryogen-free cell transportation strategy leveraging a rapid reversible thermoresponsive collagen (RRTC) hydrogel regulated by simulated body fluid (SBF). Operating via temperature-driven physical network assembly and disassembly rather than chemical crosslinking or chemical modifications, the RRTC system undergoes a rapid sol-to-gel transition within 60 s at 37 °C for efficient cell encapsulation, and completely reverses to a free-flowing sol state within 60 s at 4 °C to facilitate enzyme-free, non-destructive cell retrieval. Using L929 fibroblasts as a standardized benchmarking cell model, the biophysical protection of the matrix was systematically evaluated under both static simulated transit (48 h and 120 h) and real-world trans-city courier transportation (an approximate 50 h round trip via SF Express) within a passively temperature-shield configuration. The SBF-regulated 3D physical confinement successfully shielded cells from manual handling, multi-axis shipping vibrations, and environmental thermal fluctuations. Post-transport evaluations demonstrated that the encapsulated cells maintained a high viability above 90% and a stable recovery yield of approximately 78%, while exhibiting robust subsequent 2D re-adhesion and sustained re-culture capacity. This thermoresponsive matrix provides a potential matrix for short-term cryogen-free cell transportation and post-transport recovery, while further studies using additional cell types, longer transportation periods, and functional assays are required to evaluate its broader applicability. Full article

The PIN-LIKES (PILS) gene family is crucial for regulating auxin homeostasis and stress adaptation in plants; nevertheless, a comprehensive study on this family in alfalfa (Medicago sativa) remains insufficient. This research found 46 MsPILS genes within the tetraploid alfalfa genome and categorized them into four subfamilies. The genes are irregularly allocated throughout 16 chromosomes, with tandem duplications acting as a primary catalyst for family expansion. Analysis indicated that all MsPILS proteins contain the conserved Mem_trans domain. The promoter study revealed that MsPILS genes had many cis-elements that respond to abiotic stressors and hormones. qRT-PCR research indicated that MsPILS genes exhibit variable expression across several tissues and respond to multiple abiotic stressors. Protein–protein interaction (PPI) research revealed PIN3, PIN5, and PIN6 as principal interacting partners of the MsPILS proteins. Subcellular localization studies indicated that MsPILS1c is in the nucleus, plasma membrane, and endoplasmic reticulum (ER). This research offers significant genetic resources and a theoretical framework for elucidating the activities of PILS genes and for molecular breeding aimed at improving stress tolerance in alfalfa. Full article

Interleukin-6 (IL-6) is a cytokine with multiple biological effects. It plays a complex and seemingly paradoxical central role in both the physiological homeostasis and pathological processes of skeletal muscle. Under physiological conditions, particularly during acute exercise, IL-6 produced and secreted by the contracting skeletal muscle itself acts as an important “myokine.” It operates in an autocrine, paracrine, or endocrine manner to regulate systemic energy metabolism, insulin sensitivity, muscle regeneration, and adaptive hypertrophy. This function is crucial for the health benefits conferred by exercise. However, under various pathological conditions—such as cancer cachexia, sepsis, muscular dystrophy, denervation, disuse atrophy, and chronic inflammatory diseases—persistently elevated systemic or local IL-6 levels become a key mediator driving skeletal muscle atrophy, metabolic disorders, and functional decline. This review systematically elaborates on the dual role of IL-6 in skeletal muscle. It provides an in-depth analysis of its downstream signaling pathways (e.g., JAK/STAT, gp130, MAPK, PI3K-Akt) and upstream regulatory mechanisms (e.g., the Piezo1/KLF15 axis, calcium signaling, mitochondrial function, oxidative stress). A particular focus is placed on discussing the distinct biological effects of classical IL-6 signaling versus trans-signaling. Furthermore, we address current challenges in research and practice, including the cell specificity of IL-6 signaling, the complexity of its temporal regulation, the definition of physiological versus pathological concentrations, discrepancies between animal models and human diseases, and the plasticity of its function across different pathological contexts. Finally, this review explores the potential of targeting the IL-6 signaling pathway as a therapeutic strategy for skeletal muscle atrophy and related metabolic diseases. Potential interventions include IL-6/IL-6R monoclonal antibodies, JAK/STAT inhibitors, gp130 modulators, exercise interventions, and nutritional strategies. This aims to provide a theoretical foundation and novel perspectives for future translational research and clinical interventions. Full article

Background: The regulation of inorganic phosphate (Pi) homeostasis is predominantly mediated by the Pi transporters belonging to the SLC34 and SLC20 families of solute carriers. However, not all Pi handling can be explained by these transporters. In this study, we sought to identify novel Pi transporters in accordance with prior findings on inhibition patterns. Methods: We have performed a functional screening of new Pi carriers using the Xenopus laevis oocyte expression system, focusing on the SLC26 family, and corroboration in cell culture. Results: Both SLC26A7 and SLC26A9 have been shown to express sodium-activated Pi uptakes with approximately 200 µmol/L Pi affinity. In both cases, Pi transport is inhibited by increasing pH and by phosphonoformate, arsenate, bicarbonate, sulfate, the chloride channel inhibitor 5-nitro-2-[(3-phenylpropyl)amino]-benzoate, and several transport site and translocation inhibitors of bicarbonate exchangers. In addition, the CFTR inhibitor GlyH-101 and the SLC4 inhibitors DIDS, SITS, and phloretin exhibited partial inhibition of SLC26A9-mediated Pi uptake. The endogenous expressions of both SLC26A7 and SLC26A9 in the renal cell lines LLC-PK1 and MDCK were primarily intracellular, colocalizing with endosomes, lysosomes, and the trans-Golgi network markers. Conversely, plasma membrane expression was found to be minimal. Pi transport in MDCK cells was sodium-independent, but when either SLC26A7 or SLC26A9 was overexpressed, sodium-activated Pi uptake was observed, along with increased expressions of SLC26A7 or SLC26A9 in the plasma membrane. Conclusions: Sodium-activated Pi transport is a novel function of the SLC26A7 and SLC26A9 multifunctional anion transporters. Further research is necessary to ascertain the relevance to Pi homeostasis in vivo. Full article

Background: Brain microvascular endothelial cells (BMECs) form the blood–brain barrier (BBB), a highly selective interface that restricts paracellular diffusion and regulates the transport of nutrients and drugs into the central nervous system via specialized transporters and receptors. Tight junction-associated protein 1 (Tjap1), also termed protein incorporated later into tight junctions (Pilt), has been localized to tight junctions (TJs) in epithelial cells and to the trans-Golgi network in fibroblasts; however, its expression, subcellular localization, and functional significance in BMECs are still unknown. Methods: We characterized Tjap1 subcellular localization in mouse and human BMEC cell lines as well as primary mouse BMECs by immunofluorescence with and without pharmacological Golgi disruption by treatment with Brefeldin A, Golgicide A or Pitstop 2. CRISPR/Cas9-mediated Tjap1 knockout cells were generated and examined with regard to their Golgi morphology using immunostaining. Tjap1 mRNA localization was examined by RNAscope in situ hybridization. Quantitative real-time PCR and Western blot was performed to assess the expression of BBB-associated efflux transporters, solute carrier transporters, and cellular receptors in control and Tjap1 knockout cells. Results: Tjap1 predominantly localized to the cis-Golgi compartment, co-localizing with Gm130 rather than Tgn38, and was absent from TJs in BMECs. Tjap1 knockout induced pronounced Golgi fragmentation BMECs. Importantly, Tjap1 knockout significantly downregulated mRNA-expression of Abcb1a, Abcb1b, Abcc4, Slc2a1, Slc7a1, Slc7a5 and Tfrc, while Abcg2 was upregulated. At the protein level, a decrease in the protein levels of Abcb1, Abcc4, Slc2a1, Slc7a1, and Tfrc was observed in Tjap1 knockout cEND cells. Conclusions: In BMECs, Tjap1 is a cis-Golgi-associated protein required for the structural integrity of the Golgi apparatus. Its deletion is associated with Golgi fragmentation and significant alterations in the mRNA and protein expression of drug transporters and receptors at the BBB. These findings identify Tjap1 as a candidate regulator of both Golgi architecture and the BBB transporter profile in vitro, with potential implications for modulating drug transport across the BBB. Full article

Methyl jasmonate (MeJA) is a key regulator of plant defense and abiotic stress responses, while terpenoids are important secondary metabolites. However, the effects of MeJA on floral volatiles in Lagerstroemia indica and the underlying mechanisms remain unclear. In L. indica ‘Whit III’, MeJA treatment rapidly increased the emission of monoterpenes (e.g., citronellol) and sesquiterpenes (e.g., trans-farnesol) and advanced the peak emission time. We identified 34 LiTPS and 22 LiMYC genes in the genome, with promoter regions enriched in JA-responsive cis-elements. Expression analysis showed that LiMYC genes encoding putative JA repressors were transiently upregulated, whereas LiTPS genes located in a chromosome 11 cluster and LiTPS13 (7.33-fold induction) were strongly activated. These results suggest that MeJA may promote an early scent production through the coordinated activation of specific LiMYC and LiTPS gene sets, pointing to a potential mechanism by which jasmonate signaling modulates floral volatile emission. Full article

Background/Objectives: CTLA-4 is a key checkpoint of peripheral immune regulation, yet its biology cannot be reduced to inhibitory signaling alone. This review discusses CTLA-4 as a dynamic regulatory pathway shaped by ligand handling, intracellular trafficking, recycling, and cell-type-specific function, and examines how these features link molecular mechanism to human disease and therapy. Methods: We synthesized the structural, mechanistic, translational, and clinical literature spanning CTLA-4 molecular biology, cell-type-specific function, inborn errors of immunity, polygenic autoimmunity, transplantation, cancer immunotherapy, and immune-related adverse events. Results: CTLA-4 function depends on surface availability, trans-endocytosis of CD80/CD86, and tight control of endosomal trafficking. These features help explain why CTLA-4 haploinsufficiency, LRBA deficiency, and DEF6 deficiency converge clinically despite different upstream lesions, and why subtler CTLA-4 variation contributes to polygenic autoimmunity. Therapeutic studies also provide mechanistic insight. Abatacept can partly replace pathway function in monogenic disease, whereas belatacept highlights the limits of ligand blockade when endogenous coinhibition is also lost. In oncology, anti-CTLA-4 antibodies act through a more complex interplay involving checkpoint blockade, Fc biology, intratumoral Treg depletion, and receptor recycling. Emerging next-generation agents aim to retain antitumor activity while reducing systemic toxicity through more selective use of these mechanisms. Conclusions: Rather than a static inhibitory receptor, CTLA-4 is better viewed as a context-dependent regulatory pathway whose function depends on trafficking, surface availability, and cellular context. This perspective links molecular mechanism to clinical phenotype and supports more precise CTLA-4-targeted therapy. Full article