Specific Transcription Factors Research Articles

Cang-ai volatile oil alleviates nasal inflammation via Th1/Th2 cell imbalance regulation in a rat model of ovalbumin-induced allergic rhinitis

We previously revealed that Cang-ai volatile oil (CAVO) regulates T-cell activity, enhancing the immune response in people with chronic respiratory diseases. However, the effects of CAVO on allergic rhinitis (AR) have not been investigated. Herein, we established an ovalbumin (OVA)-induced AR rat model to determine these effects. Sprague–Dawley (SD) rats were exposed to OVA for 3 weeks. CAVO or loratadine (positive control) was given orally once daily for 2 weeks to OVA-exposed rats. Behavior modeling nasal allergies was observed. Nasal mucosa, serum, and spleen samples of AR rats were analyzed. CAVO treatment significantly reduced the number of nose rubs and sneezes, and ameliorated several hallmarks of nasal mucosa tissue remodeling: inflammation, eosinophilic infiltration, goblet cell metaplasia, and mast cell hyperplasia. CAVO administration markedly upregulated expressions of interferon-γ, interleukin (IL)-2, and IL-12, and downregulated expressions of serum tumor necrosis factor-α, IL-4, IL-5, IL-6, IL-13, immunoglobulin-E, and histamine. CAVO therapy also increased production of IFN-γ and T-helper type 1 (Th1)-specific T-box transcription factor (T-bet) of the cluster of differentiation-4+ T-cells in splenic lymphocytes, and protein and mRNA expressions of T-bet in nasal mucosa. In contrast, levels of the Th2 cytokine IL-4 and Th2-specific transcription factor GATA binding protein-3 were suppressed by CAVO. These cumulative findings demonstrate that CAVO therapy can alleviate AR by regulating the balance between Th1 and Th2 cells.

Epistasis facilitates functional evolution in an ancient transcription factor.

A protein's genetic architecture - the set of causal rules by which its sequence produces its functions - also determines its possible evolutionary trajectories. Prior research has proposed that the genetic architecture of proteins is very complex, with pervasive epistatic interactions that constrain evolution and make function difficult to predict from sequence. Most of this work has analyzed only the direct paths between two proteins of interest - excluding the vast majority of possible genotypes and evolutionary trajectories - and has considered only a single protein function, leaving unaddressed the genetic architecture of functional specificity and its impact on the evolution of new functions. Here, we develop a new method based on ordinal logistic regression to directly characterize the global genetic determinants of multiple protein functions from 20-state combinatorial deep mutational scanning (DMS) experiments. We use it to dissect the genetic architecture and evolution of a transcription factor's specificity for DNA, using data from a combinatorial DMS of an ancient steroid hormone receptor's capacity to activate transcription from two biologically relevant DNA elements. We show that the genetic architecture of DNA recognition consists of a dense set of main and pairwise effects that involve virtually every possible amino acid state in the protein-DNA interface, but higher-order epistasis plays only a tiny role. Pairwise interactions enlarge the set of functional sequences and are the primary determinants of specificity for different DNA elements. They also massively expand the number of opportunities for single-residue mutations to switch specificity from one DNA target to another. By bringing variants with different functions close together in sequence space, pairwise epistasis therefore facilitates rather than constrains the evolution of new functions.

P62 promotes FSH-induced antral follicle formation by directing degradation of ubiquitinated WT1

In females, the pathophysiological mechanism of poor ovarian response (POR) is not fully understood. Considering the expression level of p62 was significantly reduced in the granulosa cells (GCs) of POR patients, this study focused on identifying the role of the selective autophagy receptor p62 in conducting the effect of follicle-stimulating hormone (FSH) on antral follicles (AFs) formation in female mice. The results showed that p62 in GCs was FSH responsive and that its level increased to a peak and then decreased time-dependently either in ovaries or in GCs after gonadotropin induction in vivo. GC-specific deletion of p62 resulted in subfertility, a significantly reduced number of AFs and irregular estrous cycles, which were same as pathophysiological symptom of POR. By conducting mass spectrum analysis, we found the ubiquitination of proteins was decreased, and autophagic flux was blocked in GCs. Specifically, the level of nonubiquitinated Wilms tumor 1 homolog (WT1), a transcription factor and negative controller of GC differentiation, increased steadily. Co-IP results showed that p62 deletion increased the level of ubiquitin-specific peptidase 5 (USP5), which blocked the ubiquitination of WT1. Furthermore, a joint analysis of RNA-seq and the spatial transcriptome sequencing data showed the expression of steroid metabolic genes and FSH receptors pivotal for GCs differentiation decreased unanimously. Accordingly, the accumulation of WT1 in GCs deficient of p62 decreased steroid hormone levels and reduced FSH responsiveness, while the availability of p62 in GCs simultaneously ensured the degradation of WT1 through the ubiquitin‒proteasome system and autophagolysosomal system. Therefore, p62 in GCs participates in GC differentiation and AF formation in FSH induction by dynamically controlling the degradation of WT1. The findings of the study contributes to further study the pathology of POR.

Identification of co-expressed central genes and transcription factors in acute myocardial infarction and diabetic nephropathy

BackgroundAcute myocardial infarction (AMI) and diabetic nephropathy (DN) are common clinical co-morbidities, but they are challenging to manage and have poor prognoses. There is no research on the bioinformatics mechanisms of comorbidity, and this study aims to investigate such mechanisms.MethodsWe downloaded the AMI data (GSE66360) and DN datasets (GSE30528 and GSE30529) from the Gene Expression Omnibus (GEO) platform. The GSE66360 dataset was divided into two parts: the training set and the validation set, and GSE30529 was used as the training set and GSE30528 as the validation set. After identifying the common differentially expressed genes (DEGs) in AMI and DN in the training set, gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses and protein–protein interaction (PPI) network construction were performed. A sub-network graph was constructed by MCODE, and 15 hub genes were screened by the Cytohubba plugin. The screened hub genes were validated, and the 15 screened hub genes were subjected to GO, KEGG, Gene MANIA analysis, and transcription factor (TF) prediction. Finally, we performed TF differential analysis, enrichment analysis, and TF and gene regulatory network construction.ResultsA total of 46 genes (43 up-regulated and 3 down-regulated) were identified for subsequent analysis. GO functional analysis emphasized the presence of genes mainly in the vesicle membrane and secretory granule membrane involved in antigen processing and presentation, lipopeptide binding, NAD + nucleosidase activity, and Toll-like receptor binding. The KEGG pathways analyzed were mainly in the phagosome, neutrophil extracellular trap formation, natural killer cell-mediated cytotoxicity, apoptosis, Fc gamma R-mediated phagocytosis, and Toll-like receptor signaling pathways. Eight co-expressed hub genes were identified and validated, namely TLR2, FCER1G, CD163, CTSS, CLEC4A, IGSF6, NCF2, and MS4A6A. Three transcription factors were identified and validated in AMI, namely NFKB1, HIF1A, and SPI1.ConclusionsOur study reveals the common pathogenesis of AMI and DN. These common pathways and hub genes may provide new ideas for further mechanistic studies.

A perplexing case report of concomitant multiple sclerosis and myasthenia gravis

Introduction & Importance: The co-occurrence of multiple sclerosis (MS) and myasthenia gravis (MG) within the Middle East and North African Region (MENA) has been scarcely reported in current literature. This rare case report explores the pathophysiological mechanisms and potential avenues of treatment modalities. Such insights can potentially facilitate the development of more efficacious and targeted treatment modalities and perhaps pave the way for disease prevention. Case Description: 29-year-old female patient presented with diplopia of two weeks duration associated with occasional blurred vision in the left eye. On physical examination, she was discovered to have marked left eye ptosis. A visual evoked potential (VEP) test was performed, which revealed asymmetrical delay. MRI imaging revealed a few white matter hyperintense foci noted at both periventricular regions and the corpus callosum with the characteristic appearance of Dawson’s fingers and thus MS was diagnosed. An anti-acetylcholine receptor antibody test returned positive, confirming the diagnosis of concurrent MG. Clinical Discussion: Proposed pathophysiological mechanisms underlying the concurrent manifestation of both diseases include, among others, the involvement of HLA haplotype and non-HLA genotypes, as well as the immunogenetic influence of specific transcription factors. Notable HLA haplotype genes include DRB1 and HLA-DQ5 genes. In contrast, non-HLA genes include the interleukin-4 receptor (IL4RA) and factor forkhead box P3 (FOXP3). Conclusion: Considering the similar immunological background of the two diseases, ideally, a single therapeutic modality could be used for management. This will hopefully simplify the patient’s treatment regimen and may ultimately reduce the treatment cost and patient burden.

CEBPA restricts alveolar type 2 cell plasticity during development and injury-repair

Cell plasticity theoretically extends to all possible cell types, but naturally decreases as cells differentiate, whereas injury-repair re-engages the developmental plasticity. Here we show that the lung alveolar type 2 (AT2)-specific transcription factor (TF), CEBPA, restricts AT2 cell plasticity in the mouse lung. AT2 cells undergo transcriptional and epigenetic maturation postnatally. Without CEBPA, both neonatal and mature AT2 cells reduce the AT2 program, but only the former reactivate the SOX9 progenitor program. Sendai virus infection bestows mature AT2 cells with neonatal plasticity where Cebpa mutant, but not wild type, AT2 cells express SOX9, as well as more readily proliferate and form KRT8/CLDN4+ transitional cells. CEBPA promotes the AT2 program by recruiting the lung lineage TF NKX2-1. The temporal change in CEBPA-dependent plasticity reflects AT2 cell developmental history. The ontogeny of AT2 cell plasticity and its transcriptional and epigenetic mechanisms have implications in lung regeneration and cancer.

Evolutionary relationship of moso bamboo forms and a multihormone regulatory cascade involving culm shape variation.

Moso bamboo (Phyllostachys edulis) known as Mao Zhu (MZ) in Chinese exhibits various forms with distinct morphological characteristics. However, the evolutionary relationship among MZ forms and the mechanisms of culm shape variation are still lacking. Here, the main differences among MZ forms were identified as culm shape variation, which were confirmed by analysing MZ forms (799 bamboo culms) and MZ (458 bamboo culms) populations. To unravel the genetic basis underlying the morphological variations, 20 MZ forms were subjected to whole-genome resequencing. Further analysis yielded 3 230 107 high-quality SNPs and uncovered low genetic diversity and high genotype heterozygosity associated with MZ forms' formation. By integrating the SNP data of 427 MZ individuals representing 15 geographic regions, the origins of eight MZ forms were successfully traced using the phylogenetic tree and the identified common heterozygous loci. Meanwhile, transcriptomic analysis was performed using shoots from MZ and its two forms with culm shape variation. The results, combined with genomic analyses, demonstrated that hormone signalling related genes played crucial roles in culm variation. Co-expression network analysis uncovered genes associated with multiple plant hormone signal transduction, especially auxin and cytokinin were involved in culm shape variation. Furthermore, the regulatory relationships of a specific transcription factor and their target genes associated with auxin and ethylene signalling were validated by yeast one-hybrid, electrophoretic mobility shift assays, and dual-luciferase reporter. Overall, this study provides important insights into the culm shape variation formation in bamboo, which facilitates to breed new varieties with novel culms.

NAC transcription factor ATAF1 negatively modulates the PIF-regulated hypocotyl elongation under a short-day photoperiod.

Day length modulates hypocotyl elongation in seedlings to optimize their overall fitness. Variations in cell growth-associated genes are regulated by several transcription factors. However, the specific transcription factors through which the plant clock increases plant fitness are still being elucidated. In this study, we identified the no apical meristem, Arabidopsis thaliana-activating factor (ATAF-1/2), and cup-shaped cotyledon (NAC) family transcription factor ATAF1 as a novel repressor of hypocotyl elongation under a short-day (SD) photoperiod. Variations in day length profoundly affected the transcriptional and protein levels of ATAF1. ATAF1-deficient mutant exhibited increased hypocotyl length and cell growth-promoting gene expression under SD conditions. Moreover, ATAF1 directly targeted and repressed the expression of the cycling Dof factor 1/5 (CDF1/5), two key transcription factors involved in hypocotyl elongation under SD conditions. Additionally, ATAF1 interacted with and negatively modulated the effects of phytochrome-interacting factor (PIF), thus inhibiting PIF-promoted gene expression and hypocotyl elongation. Taken together, our results revealed ATAF1-PIF as a crucial pair modulating the expression of key transcription factors to facilitate plant growth during day/night cycles under fluctuating light conditions.

The effect of miR-23b-3p on regulating GH by targeting POU1F1 in Yanbian yellow cattle

This study aimed to evaluate the effect of miR-23b-3p on growth hormone (GH) in pituitary cells of Yanbian yellow cattle. The mRNA and protein levels of GH and miR-23b-3p target genes were measured by real time fluorescence quantitative PCR (qPCR) and Western blot, respectively. The target relationship of miR-23b-3p was validated by double luciferase reporter gene system. The results showed that GH mRNA and protein levels in pituitary cells of Yanbian yellow cattle were significantly lower in the miR-23b-3p-mi group than in the NC group (P<0.01), while GH mRNA and protein levels were higher in the miR-23b-3p-in group than in the iNC group (P<0.05). The result of bioinformatics analysis and double luciferase reporter gene system validation proved that miR-23b-3p targeted 3′UTR of pituitary specific transcription factor 1 (POU1F1). POU1F1 mRNA and protein levels were lower miR-23b-3p-mi group than in the NC group (P<0.01), while POU1F1 mRNA and protein levels were higher in the miR-23b-3p-in group than in the iNC group (P<0.01). These results demonstrated that miR-23b-3p could regulate GH expression in pituitary cells by regulating POU1F1 gene.

Signalling: Growth Promoters and Inhibitors

Plant growth and development are intricately regulated by a multitude of signaling pathways that respond to internal and external cues. Central to these regulatory mechanisms are the signaling pathways of growth promoters and inhibitors. Various growth-promoting hormones, including auxins, cytokinins, gibberellins, and brassinosteroids, stimulate cell division, elongation, and differentiation, playing vital roles in shaping plant architecture and influencing various developmental processes. The pathways of these growth promoters involve specific receptors and transcription factors that orchestrate growth in response to dynamic signals. In contrast, growth inhibitors, represented by hormones like abscisic acid, ethylene, jasmonic acid, and salicylic acid, act as crucial checkpoints that restrict growth during stress, pathogen attacks, or other adverse conditions. These signaling pathways trigger growth inhibition and activate defense mechanisms, enabling plants to adapt to challenging environments. Balancing the interplay between growth promoters and inhibitors allows plants to allocate resources efficiently and ensure their survival under changing circumstances. Understanding these signaling pathways is not only fundamental to plant biology but also essential for applications in agriculture, horticulture, and environmental management. This abstract provides a concise overview of these essential processes. This knowledge enables us to manipulate plant growth and development for improved crop yields, disease resistance, and stress tolerance. Ongoing research continues to unravel the complexities of these pathways, offering valuable insights into how plants delicately balance growth promotion and inhibition to thrive in an ever-evolving world.

Genome-wide DNA methylation changes in human spermatogenesis.

Sperm production and function require the correct establishment of DNA methylation patterns in the germline. Here, we examined the genome-wide DNA methylation changes during human spermatogenesis and its alterations in disturbed spermatogenesis. We found that spermatogenesis is associated with remodeling of the methylome, comprising a global decline in DNA methylation in primary spermatocytes followed by selective remethylation, resulting in a spermatids/sperm-specific methylome. Hypomethylated regions in spermatids/sperm were enriched in specific transcription factor binding sites for DMRT and SOX family members and spermatid-specific genes. Intriguingly, while SINEs displayed differential methylation throughout spermatogenesis, LINEs appeared to be protected from changes in DNA methylation. In disturbed spermatogenesis, germ cells exhibited considerable DNA methylation changes, which were significantly enriched at transposable elements and genes involved in spermatogenesis. We detected hypomethylation in SVA and L1HS in disturbed spermatogenesis, suggesting an association between the abnormal programming of these regions and failure of germ cells progressing beyond meiosis.

Interplay of RUNX2 and KLF4 in initial commitment of odontoblast differentiation.

Odontoblast differentiation is a key process in dentin formation. Mouse dental papilla cells (mDPCs) are pivotal in dentinogenesis through their differentiation into odontoblasts. Odontoblast differentiation is intricately controlled by transcription factors (TFs) in a spatiotemporal manner. Previous research explored the role of RUNX2 and KLF4 in odontoblast lineage commitment, respectively. Building on bioinformatics analysis of our previous ATAC-seq profiling, we hypothesized that KLF4 potentially collaborates with RUNX2 to exert its biological role. To investigate the synergistic effect of multiple TFs in odontoblastic differentiation, we first examined the spatiotemporal expression patterns of RUNX2 and KLF4 in dental papilla at the bell stage using immunostaining techniques. Notably, RUNX2 and KLF4 demonstrated colocalization in preodontoblast. Further, immunoprecipitation and proximity ligation assays verified the interaction between RUNX2 and KLF4 in vitro. Specifically, the C-terminus of RUNX2 was identified as the interacting domain with KLF4. Functional implications of this interaction were investigated using small hairpin RNA-mediated knockdown of Runx2, Klf4, or both. Western blot analysis revealed a marked decrease in DSPP expression, an odontoblast differentiation marker, particularly in the double knockdown condition. Additionally, alizarin red S staining indicated significantly reduced mineralized nodule formation in this group. Collectively, our findings highlight the synergistic interaction between RUNX2 and KLF4 in promoting odontoblast differentiation from mDPCs. This study contributes to a more comprehensive understanding of the regulatory network of TFs governing odontoblast differentiation.

Identification and functional analysis of the DOF gene family in Populus simonii: implications for development and stress response.

Populus simonii, a notable native tree species in northern China, demonstrates impressive resistance to stress, broad adaptability, and exceptional hybridization potential. DOF family is a class of specific transcription factors that only exist in plants, widely participating in plant growth and development, and also playing an important role in abiotic stress response. To date, there have been no reported studies on the DOF gene family in P. simonii, and the expression levels of this gene family in different tissues of poplar, as well as its expression patterns under cold, heat, and other stress conditions, remain unclear. In this study, DOF gene family were identified from the P. simonii genome, and various bioinformatics data on the DOF gene family, gene structure, gene distribution, promoters and regulatory networks were analyzed. Quantitative real time PCR technology was used to verify the expression patterns of the DOF gene family in different poplar tissues. This research initially pinpointed 41 PSDOF genes in P. simonii genome. The chromosomal localization results revealed that the PSDOF genes is unevenly distributed among 19 chromosomes, with the highest number of genes located on chromosomes 4, 5, and 11. A phylogenetic tree was constructed based on the homology between Arabidopsis thaliana and P. simonii, dividing the 41 PSDOF genes into seven subgroups. The expression patterns of PSDOF genes indicated that specific genes are consistently upregulated in various tissues and under different stress conditions, suggesting their pivotal involvement in both plant development and response to stress. Notably, PSDOF35 and PSDOF28 serve as pivotal hubs in the interaction network, playing a unique role in coordinating with other genes within the family. The analysis enhances our comprehension of the functions of the DOF gene family in tissue development and stress responses within P. simonii. These findings provide a foundation for future exploration into the biological roles of DOF gene family.

Transcriptomic responses to drought stress in the Patagonian southern beech Nothofagus alpina

BackgroundDeciphering the genetic architecture of drought tolerance could allow the candidate genes identification responding to water stress. In the Andean Patagonian forest, the genus Nothofagus represents an ecologically relevant species to be included in different genomic studies. These studies are scarce in South American ecosystems however represent an important source of genomic data in order to interpret future climate-change environment scenarios of these emblematic forests. Here, we achieved the assemblage of the transcriptome of N. alpina while searching for key genes of activated or suppressed metabolic pathways in response to drought stress.ResultsDe novo transcriptome assembly resulted in 104,030 transcripts. Following confirmation of drought conditions, based on reduction of leaf water potential and stomatal conductance, a differential gene expression analysis resulted in 2720 significantly expressed genes (1601 up-regulated and 1119 down-regulated). Enrichment analysis (over-representation analysis and gene set enrichment analysis) resulted in more than one hundred stress-responsive term ontologies (i.e. biological processes) and pathways. Terms such as response to abscisic acid and pathways such as plant hormone signal transduction or starch and sucrose metabolism were over-represented. Protein–protein interaction assessment resulted in networks with significantly expressed top common hub gene clusters (e.g. plant-type cell wall biogenesis among down-regulated or ABA-signalling among up-regulated). These networks evidenced important regulators at gene expression such as transcriptional factors.ConclusionsResponses of N. alpina seedlings to drought stress were evidenced by the activation of several genes linked to GO biological processes and KEGG pathways, which were mainly based on over-expression of specific protein kinases, phosphatases, synthases and transcription factors. This suggests an up-regulation of signalling pathways, triggered through plant hormones such as abscisic acid or auxin, which could counteract the osmotic stress created as a probable immediate response to drought. On the other hand, groups of carbon fixation genes related to the galactose metabolism, photosynthesis, secondary wall biogenesis, and fatty acid biosynthesis degradation were down-regulated under drought. Overall, our results provide new genomic data for understanding how non-model long-lived trees of Patagonian forests would acclimate to environmental changes.

Crosstalk between long non-coding RNAs and miRNAs regulates transferase activities in response to salt stress in Pistachio (Pistacia vera L.)

Long non-coding RNA (lncRNA) molecules are important regulators of a broad spectrum of biological processes, including adaptation to diverse stresses in plants. However, how plant lncRNAs function during salt stress conditions is largely unknown. In our study, we identified 48 conserved mature Pistacia vera L. (P. vera) miRNAs among pistachio miRNA precursors and used RNA sequencing data to predict and validate the interaction relationship between lncRNAs and miRNAs. We indicated that the regulatory networks of pve-miRNA-lncRNA-mRNAs were more complex in the salt-tolerant (Ghazvini) compared to the salt-sensitive (Sarakhs) pistachio cultivar under NaCl treatment. We also found that 7 and 8 salt-responsive lncRNAs play a role as competing endogenous target mimics (eTMs) for 7 pve-miRNA families in Ghazvini (mir164, mir169, mir397, and mir827) and Sarakhs (mir156, mir159, mir169, and mir482), respectively. The GO-term enrichment analysis showed that protein-coding target genes of pve-miRNAs that regulated with target mimic relationship mainly enriched in transferase activity include: 7-deoxyloganetin glucosyltransferase, Arginine N-methyltransferase, Glutathione transferase, Flavonol sulfotransferase, and Beta-glucuronosyltransferase. Their activity regulates alkaloids and polysulfated flavonols biosynthesis, sequesters sodium, improves antioxidant capacity and controls epigenetic elements in Ghazvini under salt treatment. In Sarakhs, the analysis of GO-term enrichment in pve-miRNAs protein-coding target genes of lncRNA-related mimic relationship enriched in response to wounding, cell death, and jasmonic acid biosynthetic process. Compared to Sarakhs, Ghazvini had a more effective ceRNA network for regulating transferase enzymes and specific transcription factors including MYB, WRKY, and ABR1 in response to salt stress, which led to more salt tolerance in the Ghazvini cultivar. The present study provides new information about the complex lncRNA-miRNA-mRNA regulatory networks in response to plant stress conditions in pistachio. Furthermore, our findings can be used for different breeding programs and the introduction of salt-tolerant pistachio rootstocks.

Regulating Schwann Cell Growth by Nanosecond Pulsed Electric Field for Peripheral Nerve Regeneration In Vitro.

Schwann cells (SCs) are myelinating cells of the peripheral nervous system, playing a crucial role in peripheral nerve regeneration. Nanosecond Pulse Electric Field (nsPEF) is an emerging method applicable in nerve electrical stimulation that has been demonstrated to be effective in stimulating cell proliferation and other biological processes. Aiming to assess whether SCs undergo significant changes under nsPEF and help explore the potential for new peripheral nerve regeneration methods, cultured RSC96 cells were subjected to nsPEF stimulation at 5 kV and 10 kV, followed by continued cultivation for 3-4 days. Subsequently, some relevant factors expressed by SCs were assessed to demonstrate the successful stimulation, including the specific marker protein, neurotrophic factor, transcription factor, and myelination regulator. The representative results showed that nsPEF significantly enhanced the proliferation and migration of SCs and the ability to synthesize relevant factors that contribute positively to the regeneration of peripheral nerves. Simultaneously, lower expression of GFAP indicated the benign prognosis of peripheral nerve injuries. All these outcomes show that nsPEF has great potential as an efficient treatment method for peripheral nerve injuries by stimulating SCs.

Abstract PO3-24-01: Mapping inter- and intra-tumor heterogeneity in Ductal Carcinoma in situ and invasive breast cancer using integrative multi-omic profiling

Abstract Background: Molecular profiling of ductal carcinoma in situ (DCIS) has shown some prognostic utility in the clinic. However, there is still an incomplete understanding of the diversity of molecular mechanisms by which DCIS progresses to invasive breast cancer (IBC).Here, we utilize integrative multi-omic profiling of co-occurring DCIS and IBC in humans as a model for mapping the relationship between tumor mutations, global gene expression and morphological changes in DCIS and IBC. Methods: We performed targeted panel DNA sequencing (Tempus xT), whole transcriptome profiling and hyperplex immunofluorescence (Lunaphore COMET) on a cohort of 39 pairs of co-occurring DCIS and IBC lesions. We performed an integrative analysis of the above multi-modal data to uncover similar and dissimilar molecular trajectories in DCIS/IBC pairs. DAVID, KEGG and Reactome databases were utilized to analyze differential pathway activity between DCIS and IBC lesions. Results: IBC samples significantly overexpressed pathways associated with cycling cells and proliferation. For DCIS, many of the enriched pathways were related to CYP enzymes and xenobiotic metabolism. We also assessed enrichment of specific transcription factor binding sites (TFBS) with the associated differentially expressed genes. The top site enriched in IBC was a promoter regulatory element of unknown function (M120 motif KRCTCNNNNMANAGC) as well as sites regulated by heat shock transcription factor 4 (HSF4). Top TFBS enriched in DCIS included MEF2, HMEF2, AR and CEBPE. The top pathways associated with IBC using DAVID were related to extracellular matrix (ECM) organization and regulation. Interestingly, the top pathways associated with DCIS also included several pathways associated with ECM organization. On Reactome pathways diagrams for ECM formation distinct components of ECM formation and organization were enriched in IBC vs DCIS lesions, with invadopodia formation components and proteoglycans specifically enriched in IBC and integrin/laminin signaling enriched in DCIS. Among the 39 cases profiled, we observed distinct patterns of DCIS/IBC pairs with highly similar mutational profiles indicative of derivation from a shared progenitor as well as pairs with little-to-no molecular concordance that likely arose independently. Even within mutationally similar DCIS/IBC pairs there were different degrees of gene expression concordance. We subdivided mutationally similar pairs into two groups based on gene expression similarity (Euclidean Distance) and identified 115 genes that differed between these groups (q &amp;lt; 0.05), with the dissimilar DCIS lesions hallmarked by upregulation of a variety of pathways related to B-cell function, complement cascade and cell cycle progression. Specific mutations also were related to conserved transcriptional programs in these lesions, with MCL-1 amplified tumors showing a common gene expression signature of 357 altered genes (q &amp;lt; 0.05) with key differences in extracellular matrix organization and adaptive immune response pathways. Dissimilarly, tumors with CKS1B copy number alterations associated with 627 differentially expressed genes (q &amp;lt; 0.05) with the top targets related to endothelial function and angiogenesis. Conclusions: Integrative multi-modal analysis helps us better understand tumor progression at multiple levels of biomolecular dysregulation and shows the complex interplay among genome alterations, gene expression reprogramming and tissue morphology. Such approaches may yield future biomarker signatures that better encapsulate the diverse mechanisms by which DCIS lesions evolve. Citation Format: Henry Kaplan, Alexa Dowdell, Racheli Ben Shimoli, Angela Crabtree, Ann Berry, Fred Robinson, Christopher Carney, Carlo Bifulco, Brian Piening. Mapping inter- and intra-tumor heterogeneity in Ductal Carcinoma in situ and invasive breast cancer using integrative multi-omic profiling [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO3-24-01.

Computation-guided transcription factor biosensor specificity engineering for adipic acid detection

Transcription factor (TF)-based biosensors that connect small-molecule sensing with readouts such as fluorescence have proven to be useful synthetic biology tools for applications in biotechnology. However, the development of specific TF-based biosensors is hindered by the limited repertoire of TFs specific for molecules of interest since current construction methods rely on a limited set of characterized TFs. In this study, we present an approach for engineering the specificity of TFs through a computation-based workflow using molecular docking that enables targeted alteration of TF ligand specificity. Using this method, we engineer the LysR family BenM TF to alter its specificity from its cognate ligand cis,cis-muconic acid to adipic acid through a single amino acid substitution identified by our computational workflow. When implemented in a cell-free system, the engineered biosensor shows higher ligand sensitivity, expanding the potential applications of this circuit. We further investigate ligand binding through molecular dynamics to analyze the substitution, elucidating the impact of modulating a single amino acid position on the mechanism of BenM ligand binding. This study represents the first application of biomolecular modeling methods for altering BenM specificity and for gaining insights into how mutations influence the structural dynamics of BenM. Such methods can potentially be applied to other TFs to alter specificity and analyze the dynamics responsible for these changes, highlighting the applicability of computational tools for informing experiments. In addition, our developed adipic acid biosensor can be applied for the identification and engineering of enzymes to produce adipic acid.

Anti-obesity effects of fucoidan from Sargassum thunbergii in adipocytes and high fat diet induced obese mice through inhibiting adipogenic specific transcription factor

The prevalence of obesity has increased and is a health concern worldwide. Due to the concerns regarding synthetic anti-obesity treatments, nowadays natural products become a trend. Previous studies proved that there is a potential to use marine algae as anti-obesity agents. Therefore, in this study, the lipid inhibitory effect of crude polysaccharide of amyloglucosidase-assisted hydrolysate from Sargassum thunbergii (STAC) and its fucoidan fractions (STAFs) on 3T3-L1 cells and high-fat diet (HFD)-induced obese mice were investigated. According to the results, the STAF3, showed the highest xylose content and exhibited significant inhibitory effects on lipid accumulation by downregulating adipogenic and lipogenic proteins in 3T3-L1 cells. Furthermore, oral supplementation with STAC significantly declined gain in body weight and fat weight, and serum lipid contents in an HFD-induced obesity mouse model. Structural and chemical characterizations demonstrated that purified STAF3 has consistent surface morphology and small particle size, with similar structural characteristics as commercial fucoidan. Together, these results indicate that STAC and purified STAF3 from Sargassum thunbergia is a potent source to develop as ananti-obesity agents or functional food products to counter obesity.

Hkb is required for DIP-α expression and target recognition in the Drosophila neuromuscular circuit

Our nervous system contains billions of neurons that form precise connections with each other through interactions between cell surface proteins. In Drosophila, the Dpr and DIP immunoglobulin protein subfamilies form homophilic or heterophilic interactions to instruct synaptic connectivity, synaptic growth, and cell survival. However, the upstream regulatory mechanisms of Dprs and DIPs are not clear. On the other hand, while transcription factors have been implicated in target recognition, their downstream cell surface proteins remain mostly unknown. We conduct an F1 dominant modifier genetic screen to identify regulators of Dprs and DIPs. We identify huckebein (hkb), a transcription factor previously implicated in target recognition of the dorsal Is motor neuron. We show that hkb genetically interacts with DIP-α and loss of hkb leads to complete removal of DIP-α expression specifically in dorsal Is motor neurons. We then confirm that this specificity is through the dorsal Is motor neuron specific transcription factor, even-skipped (eve), which acts downstream of hkb. Analysis of the genetic interaction between hkb and eve reveals that they act in the same pathway to regulate dorsal Is motor neuron connectivity. Our study provides insight into the transcriptional regulation of DIP-α and suggests that distinct regulatory mechanisms exist for the same CSP in different neurons.