Exploring causal correlations between circulating cytokines and atopic dermatitis: a bidirectional two-sample Mendelian randomization study.

Objectives: Numerous observational studies have reported associations between circulating cytokines and atopic dermatitis (AD); however, the causal relationships between them remain unclear. To explore the causal correlations and direction of causal effects between AD and levels of 91 circulating cytokines. Methods: Two-sample Mendelian randomization (MR) analyses were conducted to examine the causal relationships between 91 circulating cytokines and AD using summary statistics from genome-wide association studies (GWAS). Reverse MR analyses were performed to investigate reverse causation. Pleiotropy and heterogeneity tests were conducted to assess the robustness of the findings. Additional transcriptome database and clinical peripheral blood mononuclear cells (PBMCs) samples were utilized to validate the results of MR analyses. Results: Levels of interleukin (IL)-13, IL-18 Receptor 1, Tumor necrosis factor ligand superfamily member 14 (TNFSF14), TNF-related activation-induced cytokine (TRANCE), C-X-C motif chemokine (CXCL)11, IL-33, TNF-beta and CD5 were suggestively associated with the risk of AD (odds ratio, OR: 1.202, 95% CI: 1.018-1.422, p = 0.030; OR: 1.029, 95% CI: 1.029-1.157, p = 0.004; OR: 1.159, 95% CI: 1.018-1.320, p = 0.026; OR: 1.111, 95% CI: 1.016-1.214, p = 0.020; OR: 0.878, 95% CI: 0.783-0.984, p = 0.025; OR: 0.809, 95% CI: 0.661-0.991, p = 0.041; OR: 0.945, 95% CI: 0.896-0.997, p = 0.038; OR: 0.764, 95% CI: 0.652-0.895, p = 8.26e-04). In addition, levels of cytokines including Axin-1, CXCL5, CXCL10, Oncostatin-M (OSM), Sulfotransferase 1A1 (SULT1A1) and TNFSF14 were suggested to be consequences of AD (Beta: -0.080, p = 0.016; Beta: -0.062, p = 0.036; Beta: -0.066, p = 0.049; Beta: -0.073, p = 0.013; Beta: -0.089, p = 0.008; Beta: -0.079, p = 0.031). IL-13, IL-18R1, TNFSF14, and TRANCE were upregulated in both lesional skin biopsies and PBMCs from AD patients. Conclusion: The study indicates that several cytokines, including IL-13, IL-18R1, TNFSF14, TRANCE, CXCL11, IL-33, TNF-beta, and CD5, are upstream of AD development, whereas a few circulating cytokines are potentially downstream in the development of AD.

Generation of B7-H3 isoform regulated by ANXA2/NSUN2/YBX1 axis in human glioma.

In recent years, in the development of emerging immunotherapy, B7-H3 is also termed as CD276 and has become a novel chimeric antigen receptor (CAR)-T target against glioma and other tumours, and aroused extensive attention. However, B7-H3 has three isoforms (2, 3 and 4Ig) with the controversial expression and elusive function in tumour especially glioma. The current study mainly focuses on the regulatory factors and related mechanisms of generation of different B7-H3 isoforms. First, we have determined that 2Ig is dominant in glioma with high malignancy, and 4Ig is widely expressed, whereas 3Ig shows negative expression in all glioma. Next, we have further found that RNA binding protein annexin A2 (ANXA2) is essential for B7-H3 isoform maintenance, but fail to determine the choice of 4Ig or 2Ig. RNA methyltransferase NOP2/Sun RNA methyltransferase 2 (NSUN2) and 5-methylcytosine reader Y-box binding protein 1 (YBX1) facilitate the production of 2Ig. Our findings have uncovered a series of factors (ANXA2/NSUN2/YBX1) that can determine the alternative generation of different isoforms of B7-H3 in glioma. Our result aims to help peers gain a clearer understanding of the expression and regulatory mechanisms of B7H3 in tumour patients, and to provide better strategies for designing B7H3 as a target in immunotherapy.

Successful treatment of infantile refractory bullous pemphigoid with baricitinib.

Infantile bullous pemphigoid (BP) is a rare autoantibody-mediated skin disorder. We report the effective treatment of a 6-month-old infant with BP using baricitinib, a Janus kinase (JAK) inhibitor, after failure with steroids and intravenous immunoglobulin. The patient achieved full remission and discontinued all medications without any relapses. To our knowledge, this is the first case of baricitinib used in an infant with BP.

Revisiting the complexity of and algorithms for the graph traversal edit distance and its variants.

The graph traversal edit distance (GTED), introduced by Ebrahimpour Boroojeny et al. (2018), is an elegant distance measure defined as the minimum edit distance between strings reconstructed from Eulerian trails in two edge-labeled graphs. GTED can be used to infer evolutionary relationships between species by comparing de Bruijn graphs directly without the computationally costly and error-prone process of genome assembly. Ebrahimpour Boroojeny et al. (2018) propose two ILP formulations for GTED and claim that GTED is polynomially solvable because the linear programming relaxation of one of the ILPs always yields optimal integer solutions. The claim that GTED is polynomially solvable is contradictory to the complexity results of existing string-to-graph matching problems. We resolve this conflict in complexity results by proving that GTED is NP-complete and showing that the ILPs proposed by Ebrahimpour Boroojeny et al. do not solve GTED but instead solve for a lower bound of GTED and are not solvable in polynomial time. In addition, we provide the first two, correct ILP formulations of GTED and evaluate their empirical efficiency. These results provide solid algorithmic foundations for comparing genome graphs and point to the direction of heuristics. The source code to reproduce experimental results is available at https://github.com/Kingsford-Group/gtednewilp/ .

In situ tissue profile of rat trigeminal nerve in trigeminal neuralgia using spatial transcriptome sequencing.

BACKGROUND: Trigeminal neuralgia (TN) is the most common neuropathic disorder in the maxillofacial region. The etiology and pathogenesis of TN have not been clearly determined to date, although there are many hypotheses. OBJECTIVE: The goal of this study was to investigate the interactions between different types of cells in TN, particularly the impact and intrinsic mechanism of demyelination on the trigeminal ganglion, and to identify new important target genes and regulatory pathways in TN. METHODS: TN rat models were prepared by trigeminal root compression, and trigeminal nerve tissues were isolated for spatial transcriptome sequencing. The gene expression matrix was reduced dimensionally by PCA and presented by UMAP. Gene function annotation was analyzed by Metascape. The progression of certain clusters and the developmental pseudotime were analyzed using the Monocle package. Modules of the gene coexpression network between different groups were analyzed based on weighted gene coexpression network analysis and assigned AddModuleScore values. The intercellular communication of genes in these networks via ligand-receptor interactions was analyzed using CellPhoneDB analysis. RESULTS: The results suggested that the trigeminal ganglion could affect Schwann cell demyelination and remyelination responses through many ligand-receptor interactions, while the effect of Schwann cells on the trigeminal ganglion was much weaker. Additionally, ferroptosis may be involved in the demyelination of Schwann cells. CONCLUSIONS: This study provides spatial transcriptomics sequencing data on TN, reveals new markers, and redefines the relationship between the ganglion and myelin sheath, providing a theoretical basis and supporting data for future mechanistic research and drug development.

Improving the productivity of gibberellic acid by combining small-molecule compounds-based targeting technology and transcriptomics analysis in Fusarium fujikuroi.

Gibberellic acid (GA3), produced industrially by Fusarium fujikuroi, stands as a crucial plant growth regulator extensively employed in the agriculture filed while limited understanding of the global metabolic network hinders researchers from conducting rapid targeted modifications. In this study, a small-molecule compounds-based targeting technology was developed to increase GA3 production. Firstly, various small molecules were used to target key nodes of different pathways and the result displayed that supplement of terbinafine improved significantly GA3 accumulation, which reached to 1.08 g/L. Subsequently, lipid and squalene biosynthesis pathway were identified as the key pathways influencing GA3 biosynthesis by transcriptomic analysis. Thus, the strategies including in vivo metabolic engineering modification and in vitro supplementation of lipid substrates were adopted, both contributed to an enhanced GA3 yield. Finally, the engineered strain demonstrated the ability to achieve a GA3 yield of 3.24 g/L in 5 L bioreactor when utilizing WCO as carbon source and feed.

Improving Hi-C contact matrices using genome graphs.

Three-dimensional chromosome structure plays an important role in fundamental genomic functions. Hi-C, a high-throughput, sequencing-based technique, has drastically expanded our comprehension of 3D chromosome structures. The first step of Hi-C analysis pipeline involves mapping sequencing reads from Hi-C to linear reference genomes. However, the linear reference genome does not incorporate genetic variation information, which can lead to incorrect read alignments, especially when analyzing samples with substantial genomic differences from the reference such as cancer samples. Using genome graphs as the reference facilitates more accurate mapping of reads, however, new algorithms are required for inferring linear genomes from Hi-C reads mapped on genome graphs and constructing corresponding Hi-C contact matrices, which is a prerequisite for the subsequent steps of the Hi-C analysis such as identifying topologically associated domains and calling chromatin loops. We introduce the problem of genome sequence inference from Hi-C data mediated by genome graphs. We formalize this problem, show the hardness of solving this problem, and introduce a novel heuristic algorithm specifically tailored to this problem. We provide a theoretical analysis to evaluate the efficacy of our algorithm. Finally, our empirical experiments indicate that the linear genomes inferred from our method lead to the creation of improved Hi-C contact matrices. These enhanced matrices show a reduction in erroneous patterns caused by structural variations and are more effective in accurately capturing the structures of topologically associated domains.

The role of NPY2R/NFATc1/DYRK1A regulatory axis in sebaceous glands for sebum synthesis.

BACKGROUND: Sebaceous glands (SGs) synthesize and secret sebum to protect and moisturize the dermal system via the complicated endocrine modulation. Dysfunction of SG are usually implicated in a number of dermal and inflammatory diseases. However, the molecular mechanism behind the differentiation, development and proliferation of SGs is far away to fully understand. METHODS: Herein, the rat volar and mammary tissues with abundant SGs from female SD rats with (post-natal day (PND)-35) and without puberty onset (PND-25) were arrested, and conducted RNA sequencing. The protein complex of Neuropeptide Y receptor Y2 (NPY2R)/NPY5R/Nuclear factor of activated T cells 1 (NFATc1) was performed by immunoprecipitation, mass spectrum and gel filtration. Genome-wide occupancy of NFATc1 was measured by chromatin immunoprecipitation sequencing. Target proteins' expression and localization was detected by western blot and immunofluorescence. RESULTS: NPY2R gene was significantly up-regulated in volar and mammary SGs of PND-25. A special protein complex of NPY2R/NPY5R/NFATc1 in PND-25. NFATc1 was dephosphorylated and activated, then localized into nucleus to exert as a transcription factor in volar SGs of PND-35. NFATc1 was especially binding at enhancer regions to facilitate the distal SG and sebum related genes' transcription. Dual specificity tyrosine phosphorylation regulated kinase 1A (DYRK1A) contributed to NFATc1 phosphorylation in PND-25, and inactivated of DYRK1A resulted in NFATc1 dephosphorylation and nuclear localization in PND-35. CONCLUSIONS: Our findings unmask the new role of NPY2R/NFATc1/DYRK1A in pubertal SG, and are of benefit to advanced understanding the molecular mechanism of SGs' function after puberty, and provide some theoretical basis for the treatment of acne vulgaris from the perspective of hormone regulation.

Revisiting the Complexity of and Algorithms for the Graph Traversal Edit Distance and Its Variants.

The graph traversal edit distance (GTED), introduced by Ebrahimpour Boroojeny et al. (2018), is an elegant distance measure defined as the minimum edit distance between strings reconstructed from Eulerian trails in two edge-labeled graphs. GTED can be used to infer evolutionary relationships between species by comparing de Bruijn graphs directly without the computationally costly and error-prone process of genome assembly. Ebrahimpour Boroojeny et al. (2018) propose two ILP formulations for GTED and claim that GTED is polynomially solvable because the linear programming relaxation of one of the ILPs always yields optimal integer solutions. The claim that GTED is polynomially solvable is contradictory to the complexity results of existing string-to-graph matching problems. We resolve this conflict in complexity results by proving that GTED is NP-complete and showing that the ILPs proposed by Ebrahimpour Boroojeny et al. do not solve GTED but instead solve for a lower bound of GTED and are not solvable in polynomial time. In addition, we provide the first two, correct ILP formulations of GTED and evaluate their empirical efficiency. These results provide solid algorithmic foundations for comparing genome graphs and point to the direction of heuristics.

De novo design of modular peptide-binding proteins by superhelical matching.

General approaches for designing sequence-specific peptide-binding proteins would have wide utility in proteomics and synthetic biology. However, designing peptide-binding proteins is challenging, as most peptides do not have defined structures in isolation, and hydrogen bonds must be made to the buried polar groups in the peptide backbone1-3. Here, inspired by natural and re-engineered protein-peptide systems4-11, we set out to design proteins made out of repeating units that bind peptides with repeating sequences, with a one-to-one correspondence between the repeat units of the protein and those of the peptide. We use geometric hashing to identify protein backbones and peptide-docking arrangements that are compatible with bidentate hydrogen bonds between the side chains of the protein and the peptide backbone12. The remainder of the protein sequence is then optimized for folding and peptide binding. We design repeat proteins to bind to six different tripeptide-repeat sequences in polyproline II conformations. The proteins are hyperstable and bind to four to six tandem repeats of their tripeptide targets with nanomolar to picomolar affinities in vitro and in living cells. Crystal structures reveal repeating interactions between protein and peptide interactions as designed, including ladders of hydrogen bonds from protein side chains to peptide backbones. By redesigning the binding interfaces of individual repeat units, specificity can be achieved for non-repeating peptide sequences and for disordered regions of native proteins.

Transcriptome characterization of human gingival mesenchymal and periodontal ligament stem cells in response to electronic-cigarettes.

The potential toxicities and threats of electronic cigarettes (E-cigs) on periodontal health remain elusive. Gingival mesenchymal stem cells (GMSCs) and periodontal ligament stem cells (PDLSCs) contribute to cell differentiation and regeneration for periodontium as well as inflammatory modulation. However, the effects of E-cig exposure on periodontal tissues, particularly GMSCs and PDLSCs, and the underlying epigenetic mechanisms remain largely unknown. In this study, we conducted RNA-seq analysis to examine the transcriptome of human GMSCs and PDLSCs exposed to four types of E-cigs (aerosol and liquid with tobacco and menthol flavor) and conventional tobacco smoke in vitro. Our results showed that E-cig exposure primarily impacted the immunoregulation and inflammatory responses to pathogenic microorganisms in GMSCs, and the microenvironment, differentiation and response to corticosteroid in PDLSCs, which were significantly different from the damage effects caused by tobacco smoke. Additionally, we discovered a large number of differentially expressed non-coding RNAs among the different E-cig exposure methods and flavors. We also noticed that in GMSCs, CXCL2 was especially down-regulated by E-cig aerosol exposure whereas up-regulated by E-liquid exposure compared to control. Of note, the enhancer elements near CXCL2 and other genes located at Chromosome 4 contributed to the transcription activity of these genes, and KDM6B was remarkably elevated in response to E-liquid exposure. Lastly, we conducted ChIP-seq analysis to confirm that the elevated gene transcription by E-liquids was due to the weakened H3K27me3 at genome-wide enhancer elements in GMSCs, but not at promoter regions. Taken together, our results characterized the diverse gene expression profiles of GMSCs and PDLSCs in response to E-cigs with different exposure methods and flavors in vitro, and indicated a novel mechanism of KDM6B-mediated H3K27me3 on enhancers for gene transcription regulation. Our data could be served as a resource for emphasizing the understanding of E-cigs in periodontal health.

Validation of diagnostic criteria for atopic dermatitis and proposal of novel diagnostic criteria for adult and elderly Chinese populations: a multicentre, prospective, clinical setting-based study.

BACKGROUND: A previous validation study showed a very low sensitivity and higher specificity associated with Hanifin and Rajka criteria (H&R) and the UK Working Party criteria (UKWP) in diagnosing AD vs. the Chinese criteria of atopic dermatitis (AD) for children (CCAD). However, their diagnostic efficacy in adult and elderly Chinese populations remains unknown. OBJECTIVES: To validate the diagnostic efficacy of three sets of AD criteria in adult and elderly Chinese populations in a hospital setting. METHODS: A total of 1034 patients (aged 19-95 years) from five university hospital dermatological clinics were recruited. Medical history, dermatological examination, AD diagnosis and evaluation of AD severity were done by dermatologists. Each patient was investigated by two dermatologist panels, one to establish a clinical diagnosis, and the other to identify and record the major or minor signs of H&R criteria, UKWP criteria and CCAD. Taking clinical diagnosis as the reference, the diagnostic efficacy of three sets of diagnostic criteria was evaluated. The χ2 test or rank sum test were used for between-groups comparisons. RESULTS: CCAD had a higher sensitivity (84.0%), especially among mild and moderate cases of AD (72.7% and 90.3%, respectively), than the H&R (58.0%; P < 0.001) and UKWP criteria (56.0%; P < 0.001) in diagnosing AD. The specificity of CCAD (92.7%) was slightly lower than the H&R (97.3%; P < 0.001) or UKWP criteria (97.4%; P < 0.001). The CCAD had the highest Youden index (0.77), accuracy rate (0.90) and Kappa value (0.76) of the three sets of diagnostic criteria. CONCLUSIONS: Consistent with results in a population of Chinese children, although the H&R and UKWP criteria had a high specificity for diagnosing AD, their low sensitivity limited their use in adult and elderly Chinese patients. Based on the high sensitivity and favourable diagnostic efficacy, the CCAD is proposed for AD diagnosis in adult and elderly Chinese populations, especially for cases of mild and moderate AD.

Enhancement of T cell infiltration via tumor-targeted Th9 cell delivery improves the efficacy of antitumor immunotherapy of solid tumors.

Insufficient infiltration of T cells severely compromises the antitumor efficacy of adoptive cell therapy (ACT) against solid tumors. Here, we present a facile immune cell surface engineering strategy aiming to substantially enhance the anti-tumor efficacy of Th9-mediated ACT by rapidly identifying tumor-specific binding ligands and improving the infiltration of infused cells into solid tumors. Non-genetic decoration of Th9 cells with tumor-targeting peptide screened from phage display not only allowed precise targeted ACT against highly heterogeneous solid tumors but also substantially enhanced infiltration of CD8+ T cells, which led to improved antitumor outcomes. Mechanistically, infusion of Th9 cells modified with tumor-specific binding ligands facilitated the enhanced distribution of tumor-killing cells and remodeled the immunosuppressive microenvironment of solid tumors via IL-9 mediated immunomodulation. Overall, we presented a simple, cost-effective, and cell-friendly strategy to enhance the efficacy of ACT against solid tumors with the potential to complement the current ACT.

Lidocaine alleviates inflammation and pruritus in atopic dermatitis by blocking different population of sensory neurons.

BACKGROUND AND PURPOSE: Atopic dermatitis is a common chronic pruritic inflammatory disease of the skin involving neuro-immune communication. Neuronal mechanism-based therapeutic treatments remain lacking. We investigated the efficacy of intravenous lidocaine therapy on atopic dermatitis and the underlying neuro-immune mechanism. EXPERIMENTAL APPROACH: Pharmacological intervention, immunofluorescence, RNA-sequencing, genetic modification and immunoassay were performed to dissect the neuro-immune basis of itch and inflammation in atopic dermatitis-like mouse model and in patients. KEY RESULTS: Lidocaine alleviated skin lesions and itch in both atopic dermatitis patients and calcipotriol (MC903)-induced atopic dermatitis model by blocking subpopulation of sensory neurons. QX-314, a charged NaV blocker that enters through pathologically activated large-pore ion channels and selectivity inhibits a subpopulation of sensory neurons, has the same effects as lidocaine in atopic dermatitis model. Genetic silencing NaV 1.8-expressing sensory neurons was sufficient to restrict cutaneous inflammation and itch in the atopic dermatitis model. However, pharmacological blockade of TRPV1-positive nociceptors only abolished persistent itch but did not affect skin inflammation in the atopic dermatitis model, indicating a difference between sensory neuronal modulation of skin inflammation and itch. Inhibition of activity-dependent release of calcitonin gene-related peptide (CGRP) from sensory neurons by lidocaine largely accounts for the therapeutic effect of lidocaine in the atopic dermatitis model. CONCLUSION AND IMPLICATIONS: NaV 1.8+ sensory neurons play a critical role in pathogenesis of atopic dermatitis and lidocaine is a potential anti-inflammatory and anti-pruritic agent for atopic dermatitis. A dissociable difference for sensory neuronal modulation of skin inflammation and itch contributes to further understanding of pathogenesis in atopic dermatitis.

Genomic distribution of signal transducer and activator of transcription (STAT) family in colorectal cancer.

JAK/STAT pathway has been widely acknowledged in the development of human cancers. However, the role of different phosphorylated STAT proteins translocating into nucleus in transcription activation of target genes is not fully understood. In present research, ChIP-seq was carried on to investigate the genome-wide distribution of the activated STAT1, STAT2, STAT3, STAT5 and STAT6 in colorectal cancer HCT-116 cells. Our observations indicated that the homodimers rather than heterodimers of STAT protein predominantly occupied on genomic DNA. STAT3 accounted for the largest proportion among all STAT proteins HCT-116 cells. Furthermore, the biased binding motif targeted by different STAT homodimers suggested the distinct biological functions. Here, we noticed that NR5A2 was a specific co-activator of STAT3 by DNA motif analysis. Co-IP assay determined that NR5A2 indeed interacted with STAT3 homodimer rather than other homodimers or heterodimers. NR5A2 knockdown resulted in a reduced binding affinity of STAT3 homodimer in the original regions. Taken together, we characterize the genome-wide landscape of activated STAT proteins, and reveal the differences of binding patterns as well as the target genes and associated functions between homodimer and heterodimer of STAT proteins in HCT-116 cells. We also present some new findings and possible mechanisms regarding the role of NR5A2 on STAT3 in CRC. Our findings may provide new insights into the design of STAT inhibitors to treat CRC and other diseases.

Predictive value of tumor-infiltrating lymphocytes detected by flow cytometry in colorectal cancer.

The high heterogeneity of tumor cells and the surrounding immune microenvironment affects the response to treatment in colorectal cancer (CRC) patients. Therefore, there is a need to identify new immune biomarkers to predict the treatment efficacy of CRC. This study aimed to explore the predictive value of tumor-infiltrating lymphocytes (TIL) for survival in CRC patients. Flow cytometry and gated analysis were performed to measure the TILs in tissue samples obtained from 536 CRC patients. The COX regression analysis showed that the CD8 + CD279+ cells had the highest impact of all evaluated TILs on postoperative disease-free survival (DFS) (P < 0.05). The optimal CD8 + CD279+ cutoff point for the prediction of survival was 12.2%. The Kaplan-Meier analysis showed significantly higher DFS in the high CD8 + CD279+ group compared with the low CD8 + CD279+ group (P < 0.05). CD8 + CD279+ cells were associated with DFS in CRC patients with the KARS mutation, MSI/MMR, perineural invasion, and those treated with neoadjuvant chemotherapy and other chemotherapeutic treatments (P < 0.05). After the multivariate adjustment, the expression of CD8 + CD279+ remained an independent risk factor for DFS. Overall, the CD8 + CD279+ cells were identified as an independent prognostic factor in CRC patients and could be used as a potential marker for postoperative DFS.

Transcriptome-scale spatial gene expression in rat arcuate nucleus during puberty.

BACKGROUND: A variety of neurons in hypothalamus undergo a complicated regulation on transcription activity of multiple genes for hypothalamic-pituitary-gonadal axis activation during pubertal development. Identification of puberty-associated cell composition and characterization of the unique transcriptional signatures across different cells are beneficial to isolation of specific neurons and advanced understanding of their functions. METHODS: The hypothalamus of female Sprague-Dawley rats in postnatal day-25, 35 and 45 were used to define the dynamic spatial atlas of gene expression in the arcuate nucleus (ARC) by 10× Genomics Visium platform. A surface protein expressed selectively by kisspeptin neurons was used to sort neurons by flow cytometric assay in vitro. The transcriptome of the isolated cells was examined using Smart sequencing. RESULTS: Four subclusters of neurons with similar gene expression signatures in ARC were identified. Only one subcluster showed the robust expression of Kiss1, which could be isolated by a unique membrane surface biomarker Solute carrier family 18 member A3 (SLC18A3). Moreover, genes in different subclusters presenting three expression modules distinctly functioned in each pubertal stage. Different types of cells representing distinct functions on glial or neuron differentiation, hormone secretion as well as estradiol response precisely affect and coordinate with each other, resulting in a complicated regulatory network for hypothalamic-pituitary-gonadal axis initiation and modulation. CONCLUSION: Our data revealed a comprehensive transcriptomic overview of ARC within different pubertal stages, which could serve as a valuable resource for the study of puberty and sexual development disorders.

The roles of DNA methylation and hydroxymethylation at short interspersed nuclear elements in the hypothalamic arcuate nucleus during puberty.

Puberty is the gateway to adult reproductive competence, encompassing a suite of complex, integrative, and coordinated changes in neuroendocrine functions. However, the regulatory mechanisms of transcriptional reprogramming in the arcuate nucleus (ARC) during onset of puberty are still not fully understood. To understand the role of epigenetics in regulating gene expression, mouse hypothalamic ARCs were isolated at 4 and 8 weeks, and the transcriptome, DNA hydroxymethylation, DNA methylation, and chromatin accessibility were assessed via RNA sequencing (RNA-seq), reduced representation bisulfite sequencing (RRBS-seq), reduced representation hydroxymethylation profiling (RRHP)-seq, and assay for transposase-accessible chromatin (ATAC-seq), respectively. The overall DNA hydroxymethylation and DNA methylation changes in retroelements (REs) were associated with gene expression modeling for puberty in the ARC. We focused on analyzing DNA hydroxymethylation and DNA methylation at two short interspersed nuclear elements (SINEs) located on the promoter of the 5-hydroxytryptamine receptor 6 (Htr6) gene and the enhancer of the KISS-1 metastasis suppressor (Kiss1) gene and investigated their regulatory roles in gene expression. Our data uncovered a novel epigenetic mechanism by which SINEs regulate gene expression during puberty.

The epigenetic role of HTR1A antagonist in facilitaing GnRH expression for pubertal initiation control.

Serotonin (5-hydroxytryptamine [5-HT]), a metabolite of tryptophan, acts on the components of the hypothalamus-hypophysis-gonad axis and induces puberty delay in mammals via 5-HT receptor 1A (HTR1A). However, the roles of HTR1A in the hypothalamus in pubertal regulation of gene expression are not fully understood. In the current study, the upregulated gonadotropin-releasing hormone (GnRH) expression in GT1-7 GnRH neuronal cells induced by the HTR1A antagonist WAY-100635 maleate was observed in vitro. Furthermore, RNA sequencing (RNA-seq) showed decreased expression of chromobox 4 (CBX4), a member of the polycomb-repressive complex 1 (PRC1), and the loss of RING2 and YY1 interaction with CBX4, suggesting the degradation of the PRC1 in GT1-7 cells treated with maleate. Chromatin immunoprecipitation sequencing (ChIP-seq) showed that the genome-wide occupancy of CBX4 and histone H2A lysine-119 ubiquitination (H2AK119ub) was compromised, especially on the promoter of GnRH. Finally, we determined that inactivation of phosphatidylinositol 3-kinase (PI3K)/Akt and mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) contributed to CBX4 downregulation. Taken together, we concluded that HTR1A antagonists could enhance GnRH transcription via PRC1 degradation and H2AK119ub loss driven by reduced CBX4 expression through PI3K/Akt and MAPK/ERK pathway suppression in GT1-7 cells and provided a potential epigenetic mechanism of action of HTR1A on GnRH gene expression for mammalian puberty onset.