Disruptions in energy homeostasis can lead to diseases like obesity and diabetes, affecting millions of people each year. Tanycytes, the adult stem cells in the hypothalamus, play crucial roles in assisting hypothalamic neurons in maintaining energy balance. Although tanycytes have been extensively studied in rodents, our understanding of human tanycytes remains limited. In this study, we utilized single-cell transcriptomics data to explore the heterogeneity of human embryonic tanycytes, investigate their gene regulatory networks, analyze their intercellular communication, and examine their developmental trajectory. Our analysis revealed the presence of two clusters of β tanycytes and three clusters of α tanycytes in our dataset. Surprisingly, human embryonic tanycytes displayed significant similarities to mouse tanycytes in terms of marker gene expression and transcription factor activities. Trajectory analysis indicated that α tanycytes were the first to be generated, giving rise to β tanycytes in a dorsal-ventral direction along the third ventricle. Furthermore, our CellChat analyses demonstrated that tanycytes generated earlier along the developmental lineages exhibited increased intercellular communication compared to those generated later. In summary, we have thoroughly characterized the heterogeneity of human embryonic tanycytes from various angles. We are confident that our findings will serve as a foundation for future research on human tanycytes.

Transcription factors play an important role in regulating the expression of detoxification genes (e.g. P450s) that confer insecticide resistance. Our previous study identified a series of candidate transcription factors (CYP6B7-fenvalerate association proteins, CAPs) that may be related to fenvalerate-induced expression of CYP6B7 in a field HDTJ strain of H. armigera. Whether these CAPs can mediate the transcript of CYP6B7 induced by fenvalerate in a susceptible HDS strain of H. armigera remains unknown. Further study showed that the expression levels of multiple CAPs were significantly induced by fenvalerate in HDS strain. Knockdown of CAP19 [fatty acid synthase-like (FAS)], CAP22 [polysaccharide biosynthesis domain-containing protein 1 (PBDC1)], CAP24 [5-formyltetrahydrofolate cycloligase (5-FCL)], CAP30 [peptidoglycan recognition protein LB-like (PGRP)] and CAP33 [NADH dehydrogenase [ubiquinone] 1 alpha subcomplex subunit 11 (NDUFA11)] resulted in significant inhibition of CYP6B7 and some other P450 genes expression; meanwhile, the sensitivity of HDS strain larvae to fenvalerate was significantly increased. In addition, PBDC1, PGRP and NDUFA11, either alone or in combination, could significantly enhance the activity of CYP6B7 promoter in HDS strain, as well as the expression level of CYP6B7 gene in Sf9 cells line. These results suggested that PBDC1, PGRP and NDUFA11 may be involved in the transcript regulation of key detoxifying genes in response to fenvalerate in HDS strain of H. armigera.

Introduction: Kinesin family member 5A (KIF5A) is a motor neuron protein expressed in neurons and involved in anterograde transportation of organelles, proteins, and RNA. Variations in the KIF5A gene that interfere with axonal transport have emerged as a distinguishing feature in several neurodegenerative disorders, including hereditary spastic paraplegia (HSP10), Charcot-Marie-Tooth disease type 2 (CMT2), and Amyotrophic Lateral Sclerosis (ALS). Methods: In this study, we implemented a computational structural and systems biology approach to uncover the role of KIF5A in ALS. Using the computational structural biology method, we explored the role of non-synonymous Single Nucleotide Polymorphism (nsSNPs) in KIF5A. Further, to identify the potential inhibitory molecule against the highly destabilizing structure variant, we docked 24 plant-derived phytochemicals involved in ALS. Results: We found KIF5AS291F variant showed the most structure destabilizing behavior and the phytocompound \"epigallocatechin gallate\" showed the highest binding affinity (-9.0 Kcal/mol) as compared to wild KIF5A (-8.4 Kcal/mol). Further, with the systems biology approach, we constructed the KIF5A protein-protein interaction (PPI) network to identify the associated Kinesin Families (KIFs) proteins, modules, and their function. We also constructed a transcriptional and post-transcriptional regulatory network of KIF5A. With the network topological parameters of PPIN (Degree, Bottleneck, Closeness, and MNC) using CytoHubba and computational knock-out experiment using Network Analyzer, we found KIF1A, 5B, and 5C were the significant proteins. The functional modules were highly enriched with microtubule motor activity, chemical synaptic transmission in neurons, GTP binding, and GABA receptor activity. In regulatory network analysis, we found KIF5A post-transcriptionally down-regulated by miR-107 which is further transcriptionally up-regulated by four TFs (HIF1A, PPARA, SREBF1, and TP53) and down-regulated by three TFs (ZEB1, ZEB2, and LIN28A). Discussion: We concluded our study by finding a crucial variant of KIF5A and its potential therapeutic target (epigallocatechin gallate) and KIF5A associated significant genes with important regulators which could decrypt the novel therapeutics in ALS and other neurodegenerative diseases.

Though originally characterized as an inactive or transcriptionally repressive factor, the NF-κB p50 homodimer has become appreciated as a physiologically relevant driver of specific target gene expression. By virtue of its low affinity for cytoplasmic IκB protein inhibitors, p50 accumulates in the nucleus of resting cells, where it is a binding target for the transcriptional co-activator IκBζ. In this study, we employed X-ray crystallography to analyze the structure of the p50 homodimer on κB DNA from the promoters of human interleukin-6 (IL-6) and neutrophil-gelatinase-associated lipocalin (NGAL) genes, both of which respond to IκBζ. The NF-κB p50 homodimer binds 11-bp on IL-6 κB DNA, while, on NGAL κB DNA, the spacing is 12-bp. This begs the question: what DNA binding mode is preferred by NF-κB p50 homodimer? To address this, we engineered a \"Test\" κB-like DNA containing the core sequence 5\'-GGGGAATTCCCC-3\' and determined its X-ray crystal structure in complex with p50. This revealed that, when presented with multiple options, NF-κB p50 homodimer prefers to bind 11-bp, which necessarily imposes asymmetry on the complex despite the symmetry inherent in both the protein and its target DNA, and that the p50 dimerization domain can contact DNA via distinct modes.

Pinus massoniana is a major fast-growing timber tree species planted in arid areas of south China, which has a certain drought-resistant ability. However, severe drought and long-term water shortage limit its normal growth and development. Therefore, in this study, physiological indices, and the transcriptome sequencing and cloning of AP2/ERF transcription factor of P. massonsiana were determined to clarify its molecular mechanism of drought stress. The results showed that stomatal conductance (Gs) content was significantly decreased, and superoxide dismutase (SOD) activity, and malondialdehyde (MDA) and abscisic acid (ABA) content were significantly increased under drought stress. Transcriptomic analysis revealed that compared to the control, 9, 3550, and 4142 unigenes with differential expression were identified by comparing plants subjected to light, moderate or severe drought. AP2/ERF with high expression was screened out for cloning. To investigate the biological functions of ERF1, it was over-expressed in wild-type Populus davdianaand × P. bolleana via the leaf disc method. Under drought stress, compared to wild-type plants, ERF1 over-expressing poplar lines (OE) maintained a higher photosynthetic rate and growth, while the transpiration rate and stomatal conductance significantly decreased and water use efficiency was improved, indicating that drought tolerance was enhanced. This study provides an insight into the molecular mechanism of drought stress adaptation in P. massoniana.

Although intrauterine adhesion (IUA) has been well recognized as a critical factor in infertility, little information is available regarding the molecular mechanisms. We performed a high-throughput RNA sequencing in the endometrium of three IUA patients and three normal controls. And another two gene expression profiles (PMID34968168 and GSE160365) were analyzed together. A total of 252 DEGs were identified. Cell cycle, E2F target, G2M checkpoint, integrin3 pathway and H1F1 signaling were aberrantly regulated in the IUA endometrium. 10 hub genes (CCL2, TFRC, THY1, IGF1, CTGF, SELL, SERPINE1, HBB, HBA1, LYZ) were exhibited in PPI analysis. FOXM1, IKBKB and MYC were three common transcription factors of DEGs. Five chemicals (MK-1775, PAC-1, TW-37, BIX-01294, 3-matida) were identified as putative therapeutic agents for IUA. Collectively, a series of DEGs associated with IUA were disclosed. Five chemicals and ten hub genes may be further explored as potential drugs and targets for IUA treatment.

The Drosophila transcription factor СР190 is one of the key proteins that determine the activity of housekeeping gene promoters and insulators. CP190 has an N-terminal BTB domain that allows for dimerization. Many of known Drosophila architectural proteins interact with the hydrophobic peptide-binding groove in the BTB domain, which is presumably a mechanisms for recruiting CP190 to regulatory elements. To study the role of the BTB domain in the interaction with architectural proteins, we obtained transgenic flies expressing CP190 variants with mutations in the peptide-binding groove, which disrupts their interaction with architectural proteins. As a result of the studies, it was found that mutations in the BTB domain do not affect binding of the CP190 protein to polytene chromosomes. Thus, our studies confirm the previously obtained data that CP190 is recruited to regulatory elements by several transcription factors interacting, in addition to BTB, with other CP190 domains.

In plants, gene expression is orchestrated by thousands of transcription factors (TFs). For instance, a large set of bHLH TFs are involved in the regulation of iron homeostasis in Arabidopsis thaliana. The identification of the direct target genes of TFs through uncovering the interaction between the TFs and cis-regulatory elements has become an essential step toward a comprehensive understanding of the iron homeostasis transcriptional regulatory network in Arabidopsis. Chromatin immunoprecipitation (ChIP) followed by qRT-PCR (ChIP-qPCR), sequencing (ChIP-seq), or chip hybridization (ChIP-chip) is a robust tool to investigate protein-DNA interactions in plants in a physiological context. The procedure generally includes six steps: DNA-protein crosslink, isolation of nuclei, shearing of chromatin, immunoprecipitation, DNA purification, and qRT-PCR analyses. In this protocol, we describe guidelines, experimental setup, and conditions for ChIP experiment in Arabidopsis. This protocol focuses on seedlings grown in control and iron deficiency conditions, but can readily be adapted for use with other Arabidopsis tissues or samples. In addition, the protocol could also be applied to perform ChIP-chip or ChIP-seq experiments.

ReViTA (Reverse in VitroTranscription Assay) is a novel in vitro transcription-based method to study gene expression under the regulation of specific transcription factors. The ReViTA system uses a plasmid with a control sequence, the promoter region of the studied gene, the transcription factor of interest, and an RNA polymerase saturated with σ70. The main objective of this study was to evaluate the method; thus, as a proof of concept, two different transcription factors were used, a transcriptional inducer, AlgR, and a repressor, LexA, from Pseudomonas aeruginosa. After the promoters were incubated with the transcription factors, the plasmid was transcribed into RNA and reverse transcribed to cDNA. Gene expression was measured using qRTPCR. Using the ReViTA plasmid, transcription induction of 55% was observed when AlgR protein was added and a 27% transcription reduction with the repressor LexA, compared with the samples without transcription factors. The results demonstrated the correct functioning of ReViTA as a novel method to study transcription factors and gene expression. Thus, ReViTA could be a rapid and accessible in vitro method to evaluate genes and regulators of various species.

The specificity in gene regulation is controlled by interactions between transcription factors (TFs) and genomic DNA regions such as promoters and enhancers. Enhanced yeast one-hybrid (eY1H) assays are among the methods used for high-throughput detection of transcription factor-DNA interactions. Here, we describe the procedure for screening interactions between DNA regions of interest (\"DNA-baits\") and an array of transcription factors (\"TF-preys\"), after DNA-bait and TF-prey yeast strains have been generated. Using a high-density array robotic platform, this method can be used to screen interactions between multiple DNA regions and >1000 TFs within a single experiment.