Transcriptional Regulatory Mechanisms Research Articles

Transcriptional Regulatory Network of the Embryonic Diapause Termination Process in Artemia

Artemia is a typical animal used for the study of the diapause mechanism. The research on the regulation mechanism of diapause mainly focuses on the occurrence and maintenance of diapause. There are few studies on the mechanism of embryonic pause termination (EDT), especially for its transcriptional regulation mechanism. This study integrated transcriptional regulatory data from ATAC-seq and gene expression data from RNA-seq to explore the transcriptional regulatory mechanisms involved in the EDT process. Through integrated analysis, four important transcription factors (TFs), SVP, MYC, RXR, and SMAD6, were found to play a role in the EDT process, in which SVP, MYC, and RXR were upregulated, while SMAD6 was downregulated in the EDT stage. Through co-expression analysis, a transcription regulatory network for these four TFs was constructed and the functions of the TFs were analyzed. The expression of the TFs was further verified by RT-qPCR. Through functional analysis, SVP was found to be predominantly involved in cell adhesion and signal transduction. MYC probably played a role in protein binding. RXR may function in the process of RNA binding and the transfer of phosphorus-containing groups. Smad6 regulated the signal transduction, cell adhesion, and oxidation–reduction processes. The expression of the key TFs was verified by RT-qPCR. The results of this work provide important clues for the mechanism of transcriptional regulation in the EDT process of Artemia.

Oocyte transcriptomes and follicular fluid proteomics of ovine atretic follicles reveal the underlying mechanisms of oocyte degeneration

BackgroundIn mammals, female fertility is influenced by the result of follicular development (ovulation or atresia). Follicular atresia is a complex physiological process that results in the degeneration of oocytes from the ovary. However, the molecular mechanisms of oocyte degeneration and key protein markers of follicular atresia remain unclear. In this study, we investigated the complex transcriptional regulatory mechanisms and protein profiles in oocytes and follicular fluid in atretic follicle stages using single-cell RNA sequencing and tandem mass tag proteomics.ResultsFirst, through paired analysis of different follicle development stages, we identified 175 atresia-specific genes and eight candidate oocyte-secreted factors, including PKG1, YTHDF2, and MYC. Meanwhile, we also characterized unique features of the oocyte transcriptional landscape in the atretic follicle stage that displayed cell death-related transcriptional changes and mechanisms, such as autophagy (TBK1 and IRS4), necroptosis (PKR), and apoptosis (MARCKS). Moreover, we identified atresia-specific genes, namely FTH1, TF, and ACSL4, which may participate in regulation of oocyte ferroptosis in atretic follicles through a series of mechanisms including ferritinophagy, ferritin transport, and lipid metabolism. Additionally, we uncovered 333 differentially expressed proteins that may coordinate follicular atresia and revealed key pathways, such as negative regulation of angiogenesis, metabolic pathways, and transcription and mRNA splicing, that lead to oocyte degeneration. Finally, by combining transcriptome and proteomics analyses, we identified two oocyte-secreted biomarkers, PGK1 and ANGPT2, that may be associated with follicular atresia.ConclusionsIn conclusion, our work offers a thorough characterization of oocyte transcription mechanism and follicular fluid protein changes in ovine atretic follicles, which offers a crucial reference for analyzing the mechanism of follicular atresia and establishing an oocyte quality assessment system in sheep.

CitGATA7 interact with histone acetyltransferase CitHAG28 to promote citric acid degradation by regulating the glutamine synthetase pathway in citrus

Organic acid is a crucial indicator of fruit quality traits. Citric acid, the predominant organic acid in citrus fruit, directly influences its edible quality and economic value. While the transcriptional regulatory mechanisms of citric acid metabolism have been extensively studied, the understanding about the transcriptional and epigenetic co-regulation mechanisms is limited. This study characterized a transcription factor, CitGATA7, which directly binds to and activates the expression of genes associated with the glutamine synthetase pathway regulating citric acid degradation. These genes include the aconitase encoding gene CitACO3, the isocitrate dehydrogenase encoding gene CitIDH1, and the glutamine synthetase encoding gene CitGS1. Furthermore, CitGATA7 physically interacts with the histone acetyltransferase CitHAG28 to enhance histone 3 acetylation levels near the transcription start site of CitACO3, CitIDH1, and CitGS1, thereby increasing their transcription and promoting citric acid degradation. The findings demonstrate that the CitGATA7-CitHAG28 protein complex transcriptionally regulate the expression of the GS pathway genes, i.e., CitACO3, CitIDH1, and CitGS1, via histone acetylation, thus promoting citric acid catabolism. This study establishes a direct link between transcriptional regulation and histone acetylation regarding citric acid metabolism, providing insights for strategies to manipulate organic acid accumulation in fruit.

Inhibition of triglyceride metabolism-associated enhancers alters lipid deposition during adipocyte differentiation.

Triglyceride (TG) metabolism is a complex and highly coordinated biological process regulated by a series of genes, and its dysregulation can lead to the occurrence of disorders in lipid metabolism. However, the transcriptional regulatory mechanisms of crucial genes in TG metabolism mediated by enhancer-promoter interactions remain elusive. Here, we identified candidate enhancers regulating the Agpat2, Dgat1, Dgat2, Pnpla2, and Lipe genes in 3T3-L1 adipocytes by integrating epigenomic data (H3K27ac, H3K4me1, and DHS-seq) with chromatin three-dimensional interaction data. Luciferase reporter assays revealed that 11 enhancers exhibited fluorescence activity. The repression of enhancers using the dCas9-KRAB system revealed the functional roles of enhancers of Dgat2 and Pnpla2 in regulating their expression and TG metabolism. Furthermore, transcriptome analyses revealed that inhibition of Dgat2-En4 downregulated pathways associated with lipid metabolism, lipid biosynthesis, and adipocyte differentiation. Additionally, overexpression and motif mutation experiments of transcription factor found that two TFs, PPARG and RXRA, regulate the activity of Agpat2-En1, Dgat2-En4, and Pnpla2-En5. Our study identified functional enhancers regulating TG metabolism and elucidated potential regulatory mechanisms of TG deposition from enhancer-promoter interactions, providing insights into understanding lipid deposition.

Glucocorticoid receptor suppresses GATA6-mediated RNA polymerase II pause release to modulate classical subtype identity in pancreatic cancer

BackgroundPancreatic ductal adenocarcinoma (PDAC) is a highly lethal cancer with a 5-year survival rate of 12%. It has two major molecular subtypes: classical and basal, regulated by the master transcription...

Reconstruction and computer analysis of the structural and functional organization of the gene network regulating cholesterol biosynthesis in humans and the evolutionary characteristics of the genes involved in the network

Cholesterol is an essential structural component of cell membranes and a precursor of vitamin D, as well as steroid hormones. Humans and other animal species can absorb cholesterol from food. Cholesterol is also syn­thesized de novo in the cells of many tissues. We have previously reconstructed the gene network regulating intra­cellular cholesterol levels, which included regulatory circuits involving transcription factors from the SREBP (Sterol Regulatory Element-Binding Proteins) subfamily. The activity of SREBP transcription factors is regulated inversely depending on the intracellular cholesterol level. This mechanism is implemented with the participation of proteins SCAP, INSIG1, INSIG2, MBTPS1/S1P and MBTPS2/S2P. This group of proteins, together with the SREBP factors, is designated as “cholesterol sensor”. An elevated cholesterol level is a risk factor for the development of cardiovas­cular diseases and may also be observed in obesity, diabetes and other pathological conditions. Systematization of information about the molecular mechanisms controlling the activity of SREBP factors and cholesterol biosyn­thesis in the form of a gene network and building new knowledge about the gene network as a single object is extremely important for understanding the molecular mechanisms underlying the predisposition to diseases. With a computer tool, ANDSystem, we have built a gene network regulating cholesterol biosynthesis. The gene network included data on: (1) the complete set of enzymes involved in cholesterol biosynthesis; (2) proteins that function as part of the “cholesterol sensor”; (3) proteins that regulate the activity of the “cholesterol sensor”; (4) genes encod­ing proteins of these groups; (5) genes whose transcription is regulated by SREBP factors (SREBP target genes). The gene network was analyzed and feedback loops that control the activity of SREBP factors were identified. These feedback loops involved the PPARG, NR0B2/SHP1, LPIN1, and AR genes and the proteins they encode. Analysis of the phylostratigraphic age of the genes showed that the ancestral forms of most human genes encoding the enzymes of cholesterol biosynthesis and the proteins of the “cholesterol sensor” may have arisen at early evolutionary stages (Cellular organisms (the root of the phylostratigraphic tree) and the stages of Eukaryota and Metazoa divergence). However, the mechanism of gene transcription regulation in response to changes in cholesterol levels may only have formed at later evolutionary stages, since the phylostratigraphic age of the genes encoding the transcription factors SREBP1 and SREBP2 corresponds to the stage of Vertebrata divergence.

SRF and CBP jointly regulate integrin β6 overexpression in head and neck squamous cell carcinomas.

Overexpression of integrin β6 (ITGB6) is crucially linked to the invasion and metastasis of head and neck squamous cell carcinoma (HNSCC). The molecular mechanisms driving ITGB6 upregulation in HNSCC are not well understood. Our study comprehensively analyzed the transcriptional regulation and epigenetic modification mechanisms affecting ITGB6 transcription. We retrospectively evaluated ITGB6 expression using immunohistochemistry on a tissue microarray. Elevated ITGB6 expression in HNSCC specimens correlates with poor clinical prognosis. Using a luciferase reporter assay, site-directed mutagenesis, RNA interference, chromatin immunoprecipitation assay, and a 4-nitroquinoline 1-oxide (4NQO)-induced murine HNSCC model, we have demonstrated that the transcription factor Serum Response Factor (SRF) upregulates ITGB6 transcription. Our results further demonstrated that the histone acetyltransferase (HAT) CBP mediates the hyperacetylation of histones H3 and H4, facilitating their recruitment to the ITGB6 promoter. This recruitment strengthens SRF binding to the ITGB6 promoter. These findings suggest that SRF and CBP-mediated histone hyperacetylation are crucial for ITGB6 overexpression in HNSCC. Epigenetic mechanisms play a critical role in the active transcriptional expression of ITGB6 in HNSCC cells.

Analysis of Functional Cis-regulatory Elements Reveals Novel Transcriptional Regulatory Mechanisms in Gonadal Development.

Recent studies have demonstrated that the production of bidirectional enhancer-derived transcripts (eRNAs) is a characteristic of an active Cis-regulatory element (CRE). Higher levels of eRNA synthesis correlate with the activation of histone modifications, a potentially valuable tool for deciphering the complexity of the gene regulatory network. To understand the changes of CREs during gonadal development in mice, we collected gonadal WT1-positive cells from the piggyBac-Wt1-mCherry-2A-EGFP (PBWt1-RG) reporter strain at E13.5, E16.5, and P0 in both sexes and conducted Cap Analysis of Gene Expression analysis (CAGE) which is capable to capture transcriptional starting site (TSS). We compared the levels of intergenic bidirectional RNA, i.e, potentially eRNA, according to sex at each stage (testis somatic cells vs ovary somatic cells at E13.5, E16.5, and P0) and stage in each sex (E13.5 vs E16.5, E16.5 vs P0, and E13.5 vs P0 in testis somatic cells or ovary somatic cells). Intergenic RNAs with significant changes (|Log2FC| > 1, p < 0.05) were selected. The TSS profile of intergenic RNA changed more profoundly in testis somatic cells than in ovary somatic cells, suggesting embryonic testicular development is driven by larger changes in a transcriptional regulatory network than ovarian development. Based on the profiles of the predicted transcription factors (TFs) that would bind to the active CREs during gonadal development, the NR4A, EGR, and TCF3 families would be novel TFs to play pivotal roles in gonadal development. Identifying active CREs using eRNAs would provide a means to comprehensively understand the transcriptional regulatory system, leading to valuable insights into the gonadal development of male and female individuals.

Tgfβ1a/vegfa gene expression and methylation in response to acute hypoxia in Japanese flounder (Paralichthys olivaceus).

The physiological response and molecular mechanism of the immune response of Japanese flounder under hypoxia stress remain unclear. In this study, we examined the immune-related physiological indexes and the molecular mechanisms of Japanese flounders under acute hypoxia stress. The results showed that the levels of serum ALT, ALP, AST and LDH in hypoxia stress group were significantly increased (P<0.01). Through quantitative real-time PCR and double in situ hybridization, we found acute hypoxia stress induced immune response of skeletal muscle and gill filaments. The transcriptional regulation mechanism of this immune signaling pathway was investigated by dual luciferase assay. In addition, DNA methylation levels of genes were detected to explore epigenetic modifications of this pathway. As a transcription factor, Foxo1a can interact tgfβ1a(AGATGTTTTTT) and vegfa(TTCTTTTTATA, TACTGTTGCTA) sequences of the promoter regions. The DNA methylation levels of tgfβ1a and vegfa genes were significantly affected by hypoxia and negatively correlated with their expression. These experiments showed that the expression of immune related genes tgfβ1a and vegfa were regulated by transcription factor Foxo1a and DNA methylation. Our study provides theoretical foundations for acute hypoxia stress induced immune response of Japanese flounder.

WRKY transcription factors: Hubs for regulating plant growth and stress responses.

As sessile organisms, plants must directly face various stressors. Therefore, plants have evolved a powerful stress resistance system and can adjust their growth and development strategies appropriately in different stressful environments to adapt to complex and ever-changing conditions. Nevertheless, prioritizing defensive responses can hinder growth; this is a crucial factor for plant survival but is detrimental to crop production. As such, comprehending the impact of adverse environments on plant growth is not only a fundamental scientific inquiry but also imperative for the agricultural industry and for food security. The traditional view that plant growth is hindered during defense due to resource allocation trade-offs is challenged by evidence that plants exhibit both robust growth and defensive capabilities through human intervention. These findings suggest that the growth‒defense trade-off is not only dictated by resource limitations but also influenced by intricate transcriptional regulatory mechanisms. Hence, it is imperative to conduct thorough investigations on the central genes that govern plant resistance and growth in unfavorable environments. Recent studies have consistently highlighted the importance of WRKY transcription factors in orchestrating stress responses and plant-specific growth and development, underscoring the pivotal role of WRKYs in modulating plant growth under stressful conditions. Here, we review recent advances in understanding the dual roles of WRKYs in the regulation of plant stress resistance and growth across diverse stress environments. This information will be crucial for elucidating the intricate interplay between plant stress response and growth and may aid in identifying gene loci that could be utilized in future breeding programs to develop crops with enhanced stress resistance and productivity.

Collaboration between DELLA proteins and the Mediator subunit MED15 to regulate transcription in plants.

Gibberellins (GA) are diterpenoids that are categorized as one of main hormones that promote major developmental responses such as germination and stem elongation in plants. DELLA proteins act as the key repressors of GA responses. They interact with hundreds of different proteins. While the functioning of DELLA as transcriptional coactivators has also been reported earlier, the actual mechanism still remains elusive. One recent report describes interaction of DELLA with Mediator subunit MED15 as one of the mechanisms contributing to its transcription activation capability (Hernández-García et al. 2024). Interestingly, this mechanism of DELLA-MED15 module-mediated transcription regulation seems to be very ancient conserved from bryophyte Marchantia polymorpha to dicot Arabidopsis thaliana.

Identification of a Subpopulation of Astrocyte Progenitor Cells in the Neonatal Subventricular Zone: Evidence that Migration is Regulated by Glutamate Signaling

In mice engineered to express enhanced green fluorescent protein (eGFP) under the control of the entire glutamate transporter 1 (GLT1) gene, eGFP is found in all ‘adult’ cortical astrocytes. However, when 8.3 kilobases of the human GLT1/EAAT2 promoter is used to control expression of tdTomato (tdT), tdT is only found in a subpopulation of these eGFP-expressing astrocytes. The eGFP mice have been used to define mechanisms of transcriptional regulation using astrocytes cultured from cortex of 1–3 day old mice. Using the same cultures, we were never able to induce tdT+ expression. We hypothesized that these cells might not have migrated into the cortex by this age. In this study, we characterized the ontogeny of tdT+ cells, performed single-cell RNA sequencing (scRNA-seq), and tracked their migration in organotypic slice cultures. At postnatal day (PND) 1, tdT+ cells were observed in the subventricular zone and striatum but not in the cortex, and they did not express eGFP. At PND7, tdT+ cells begin to appear in the cortex with their numbers increasing with age. At PND1, scRNA-seq demonstrates that the tdT+ cells are molecularly heterogeneous, with a subpopulation expressing astrocytic markers, subsequently validated with immunofluorescence. In organotypic slices, tdT+ cells migrate into the cortex, and after 7 days they express GLT1, NF1A, and eGFP. An ionotropic glutamate receptor (iGluR) antagonist reduced by 50% the distance tdT+ cells migrate from the subventricular zone into the cortex. The pan-glutamate transport inhibitor, TFB-TBOA, increased, by sixfold, the number of tdT+ cells in the cortex. In conclusion, although tdT is expressed by non-glial cells at PND1, it is also expressed by glial progenitors that migrate into the cortex postnatally. Using this fluorescent labeling, we provide novel evidence that glutamate signaling contributes to the control of glial precursor migration.

A unified framework for cell-type-specific eQTL prioritization by integrating bulk and scRNA-seq data.

Genome-wide association studies (GWASs) have identified numerous genetic variants associated with complex traits, yet the biological interpretation remains challenging, especially for variants in non-coding regions. Expression quantitative trait locus (eQTL) studies have linked these variations to gene expression, aiding in identifying genes involved in disease mechanisms. Traditional eQTL analyses using bulk RNA sequencing (bulk RNA-seq) provide tissue-level insights but suffer from signal loss and distortion due to unaddressed cellular heterogeneity. Recently, single-cell RNA-seq (scRNA-seq) has provided higher resolution, enabling cell-type-specific eQTL (ct-eQTL) analyses. However, these studies are limited by their smaller sample sizes and technical constraints. In this paper, we present a statistical framework, IBSEP, which integrates bulk RNA-seq and scRNA-seq data for enhanced ct-eQTL prioritization. Our method employs a hierarchical linear model to combine summary statistics from both data types, overcoming the limitations while leveraging the advantages associated with each technique. Through extensive simulations and real data analyses, including peripheral blood mononuclear cells and brain cortex datasets, IBSEP demonstrated superior performance in identifying ct-eQTLs compared to existing methods. Our approach unveils transcriptional regulatory mechanisms specific to cell types, offering deeper insights into the genetic basis of complex diseases at a cellular resolution.

Epibrassinolide Regulates Lhcb5 Expression Though the Transcription Factor of MYBR17 in Maize.

Photosynthesis, which is the foundation of crop growth and development, is accompanied by complex transcriptional regulatory mechanisms. Research has established that brassinosteroids (BRs) play a role in regulating plant photosynthesis, with the majority of research focusing on the physiological level and regulation of rate-limiting enzymes in the dark reactions of photosynthesis. However, studies on their effects on maize photosynthesis, specifically on light-harvesting antenna proteins, have yet to be conducted. The peripheral light-harvesting antenna protein Lhcb5 is crucial for capturing and dissipating light energy. Herein, by analyzing the transcriptomic data of maize seedling leaves treated with 24-epibrassinolide (EBR) and verifying them using qPCR experiments, we found that the MYBR17 transcription factor may regulate the expression of the photosynthetic light-harvesting antenna protein gene. Further experiments using protoplast transient expression and yeast one-hybrid tests showed that the maize transcription factor MYBR17 responds to EBR signals and binds to the promoter of the light-harvesting antenna protein Lhcb5, thereby upregulating its expression. These results were validated using an Arabidopsis mybr17 mutant. Our results offer a theoretical foundation for the application of BRs to enhance the photosynthetic efficiency of maize.

Novel target and cofactor repertoire for the transcriptional regulator JTY_0672 from Mycobacterium bovis BCG.

Mycobacterium tuberculosis (Mtb) is the pathogenic agent of tuberculosis (TB). Intracellular survival plays a central role in the pathogenesis of Mtb in a manner that is dependent on an array of transcriptional regulators for Mtb. However, the functionality of JTY_0672, a member of the TetR family of transcriptional regulators, remains unknown. In this study, EMSA, BIL, ChlP-PCR and animal models were used to investigate the regulation function of this protein. We found that the transcriptional regulator JTY_0672 is a broad-spectrum transcriptional regulatory protein and can directly regulate JTY_3148, both in vitro and in vivo. Cofactors containing V B1, V B3, V B6, V C , His, Cys, Asp, Glu, Fe3+, Pb2+, Cu2+, and Li+ were found to inhibit binding between JTY_0672 and the promoter of JTY_3148. JTY_0672 enhanced TAG production and increased Isoniazid (INH) resistance. Besides, this protein either promoted recalcitrance to the host immune response and induced pathological injury and inflammation. In summary, this research identified new targets and cofactors of JTY_0672 and deciphered the physiological functionality of JTY_0672. Our findings will provide an important theoretical basis for understanding the Mtb transcriptional regulatory mechanism.

Asymmetry of Motif Conservation Within Their Homotypic Pairs Distinguishes DNA-Binding Domains of Target Transcription Factors in ChIP-Seq Data

(1) Background: Transcription factors (TFs) are the main regulators of eukaryotic gene expression. The cooperative binding of at least two TFs to genomic DNA is a major mechanism of transcription regulation. Massive analysis of the co-occurrence of overrepresented pairs of motifs for different target TFs studied in ChIP-seq experiments can clarify the mechanisms of TF cooperation. (2) Methods: We categorized the target TFs from M. musculus ChIP-seq and A. thaliana ChIP-seq/DAP-seq experiments according to the structure of their DNA-binding domains (DBDs) into classes. We studied homotypic pairs of motifs, using the same recognition model for each motif. Asymmetric and symmetric pairs consist of motifs of remote and close recognition scores. (3) Results: We found that asymmetric pairs of motifs predominate for all TF classes. TFs from the murine/plant ‘Basic helix–loop–helix (bHLH)’, ‘Basic leucine zipper (bZIP)’, and ‘Tryptophan cluster’ classes and murine ‘p53 domain’ and ‘Rel homology region’ classes showed the highest enrichment of asymmetric homotypic pairs of motifs. Pioneer TFs, despite their DBD types, have a higher significance of asymmetry within homotypic pairs of motifs compared to other TFs. (4) Conclusion: Asymmetry within homotypic CEs is a promising new feature decrypting the mechanisms of gene transcription regulation.

CDK8 mediated inflammatory microenvironment aggravates osteoarthritis progression.

Cyclin-Dependent Kinase 8 (CDK8), a CDK family member, regulates the development of inflammatory processes through transcriptional activation. The involvement of CDK8 in osteoarthritis (OA) progression is not yet understood. This study aims to investigate whether CDK8, through its transcriptional regulatory functions, collaborates with NF-κB in chondrocytes to regulate the transcription of senescence-associated secretory phenotype (SASP) genes, thereby exacerbating the inflammatory microenvironment in the progression of osteoarthritis (OA), and to explore the specific mechanisms involved. The effects of CDK8 silencing or overexpression will be assessed by measuring OA pathological markers through H&E staining, immunoblotting, Western blot, qRT-PCR, immunofluorescence and ELISA. The DMM surgery mouse model will be used as the OA model, and the PAM and Von Frey tests will be employed to measure the pain threshold in mice. Luciferase and ChIP assays will be conducted to explore the transcriptional regulation and elongation mechanisms of CDK8. CDK8 influences OA advancement by being recruited to the SASP promoter region in cooperation with NF-κB, leading to the elongation phosphorylation of Rpb1 CTD within the context of NF-κB-induced gene specificity, thereby regulating SASP transcription. The SASP secreted by chondrocytes during this process promotes the inflammatory microenvironment in the joint and drives macrophage differentiation into osteoclasts, further worsening the severity of osteoarthritis. The SASP secreted by chondrocytes during the OA process plays a crucial role in worsening the severity of the disease. Inhibiting CDK8 expression can decrease its secretion by downregulating the transcription levels of SASP, which are co-regulated by CDK8 and NF-κB. This could offer a new target for osteoarthritis treatment.

Transcriptional regulation of Rankl by Txnip-Ecd in aging and diabetic related osteoporosis.

Bone homeostasis between osteoclast bone resorption and osteoblastic bone formation is tightly regulated by a series of factors such as the receptor activator of nuclear factor-κB ligand (RANKL). Denosumab that neutralizes RANKL is effective and widely applied in the treatment of postmenopausal osteoporosis. However, factors that participated in the RANKL-related bone remodeling process in primary and secondary osteoporosis are less known. Revealing the novel transcriptional regulatory mechanism of RANKL is of great significance for the treatment of osteoporosis. After differential expression genes (DEGs) intersection and screening, we generated Thioredoxin-interacting protein (Txnip) bone marrow-derived mesenchymal stromal cells (BMSCs) genetic knockout mice and performed bone histomorphometry and histological analysis. RNA-Sequencing, Western blotting and immunofluorescence staining verified Rankl downregulation. Co-immunoprecipitation and immunofluorescence staining were used for Rankl regulation mechanism exploration. A specific inhibitor was selected for treatment effect verification. Txnip knockout in BMSCs impaired its osteogenic differentiation, suppressed Rankl expression and subsequent osteoclast formation and thus led to increased bone mass. The regulatory function of Txnip on Rankl expression was revealed for the first time through the novel transcription-related Ecdysoneless (Ecd)-P300 axis. Pharmacological inhibition of Txnip can effectively prevent bilateral ovariectomy (OVX)-induced osteoporosis. Inhibition of Txnip is an alternative way to suppress Rankl-mediated osteoblast and osteoclast crosstalk. This interesting finding rendered Txnip an ideal therapeutic target for the treatment of both ovariectomy-induced and diabetes-induced osteoporosis.

Dimerization among multiple NAC proteins mediates secondary cell wall cellulose biosynthesis in cotton fibers

SUMMARYCotton fibers, essentially cellulosic secondary cell walls (SCWs) when mature, are the most important raw material for natural textiles. SCW cellulose biosynthesis determines fiber thickness and industrially important fiber quality parameters, such as fiber strength and fiber length. However, transcriptional regulatory networks controlling fiber SCW cellulose formation remain incomplete. Here, we identify eight NAC domain proteins (GhNACs) that are involved in fiber SCW cellulose synthesis. These eight GhNACs can form pairwise heterodimers that may act as dimers, or perhaps even as an octameric protein complex, to transactivate GhCesA expression. Moreover, heterodimerization of GhNACs can in different combinations synergistically activate GhCesA genes. Through our analyses of transcription factor—DNA and transcription factor—transcription factor interactions, we propose a multi‐layered transcriptional regulatory network in which the regulation of SCW cellulose biosynthesis in cotton fiber is mediated by multiple NAC protein dimers. These findings enhance our understanding of the roles of NAC proteins in SCW formation and offer new insights into fiber‐specific transcriptional regulatory mechanisms of cellulose synthesis.

BarH1 regulates the expression of conserved odorant-binding protein 22 from Dastarcus helophoroides.

Chemical signals are pivotal in establishing tritrophic interactions among host plants, herbivorous insects, and natural enemies. Previous studies have shown that evolutionarily conserved MaltOBPs in Monochamus alternatus and DhelOBPs in Dastarcus helophoroides contribute to the establishment of pine -pest - natural enemy tritrophic interactions by recognizing the same volatile emitted by the host during crucial developmental stages. We hypothesized that the transcriptional regulatory mechanisms of evolutionarily conserved OBPs respectively from pests and enemies are similar. In this study, we identified the promoter region of DhelOBP22 through chromosome walking and discovered that transcription factor BarH1, which have been proved to regulate the expression of MaltOBP19 in M. alternatus, regulates the expression of DhelOBP22 by binding to its promoter region, as evidenced by dual-luciferase assays and electrophoretic mobility shift assay (EMSA). When the BarH1 gene was silenced using RNAi, the expression of DhelOBP22 was inhibited, leading to the disappearance of the attracted behavior of mated female D. helophoroides adults towards camphene. This study underscores that conserved transcriptional regulation mechanisms play a role in regulating the expression of evolutionarily conserved OBPs respectively from herbivores and natural enemies.