Key Regulatory Factors Research Articles

Protein Kinase C Theta Modulates PCMT1 through hnRNPL to Regulate FOXP3 Stability in Regulatory T Cells

T cell receptor signaling, together with cytokine-induced signals, can differentially regulate RNA processing to influence T helper versus regulatory Tcell fate. Protein kinase C family members have been shown to function in alternative splicing and RNA processing in various cell types. Tcell-specific protein kinase C theta, a molecular regulator of Tcell receptor downstream signaling, has been shown to phosphorylate splicing factors and affect post-transcriptional control of Tcell gene expression. In this study, we explored how using a synthetic cell-penetrating peptide mimic for intracellular anti-protein kinase C theta delivery fine-tunes differentiation of induced regulatory Tcells through its differential effects on RNA processing. We identified protein kinase C theta signaling as a critical modulator of two key RNA regulatory factors, heterogeneous nuclear ribonucleoprotein L (hnRNPL) and protein-l-isoaspartate O-methyltransferase-1 (PCMT1), and loss of protein kinase C theta function initiated a "switch" in post-transcriptional organization in induced regulatory Tcells. More interestingly, we discovered that protein-l-isoaspartate O- methyltransferase-1 acts as an instability factor in induced regulatory Tcells, by methylating the forkhead box P3 (FOXP3) promoter. Targeting protein-l-isoaspartate O-methyltransferase-1 using a cell-penetrating antibody revealed an efficient means of modulating RNA processing to confer a stable regulatory Tcell phenotype.

Pivotal role of the transcriptional co-activator YAP in trophoblast stemness of the developing human placenta.

Various pregnancy complications, such as severe forms of preeclampsia or intrauterine growth restriction, are thought to arise from failures in the differentiation of human placental trophoblasts. Progenitors of the latter either develop into invasive extravillous trophoblasts, remodeling the uterine vasculature, or fuse into multinuclear syncytiotrophoblasts transporting oxygen and nutrients to the growing fetus. However, key regulatory factors controlling trophoblast self-renewal and differentiation have been poorly elucidated. Using primary cells, three-dimensional organoids, and CRISPR-Cas9 genome-edited JEG-3 clones, we herein show that YAP, the transcriptional coactivator of the Hippo signaling pathway, promotes maintenance of cytotrophoblast progenitors by different genomic mechanisms. Genetic or chemical manipulation of YAP in these cellular models revealed that it stimulates proliferation and expression of cell cycle regulators and stemness-associated genes, but inhibits cell fusion and production of syncytiotrophoblast (STB)-specific proteins, such as hCG and GDF15. Genome-wide comparisons of primary villous cytotrophoblasts overexpressing constitutively active YAP-5SA with YAP KO cells and syncytializing trophoblasts revealed common target genes involved in trophoblast stemness and differentiation. ChIP-qPCR unraveled that YAP-5SA overexpression increased binding of YAP-TEAD4 complexes to promoters of proliferation-associated genes such as CCNA and CDK6 Moreover, repressive YAP-TEAD4 complexes containing the histone methyltransferase EZH2 were detected in the genomic regions of the STB-specific CGB5 and CGB7 genes. In summary, YAP plays a pivotal role in the maintenance of the human placental trophoblast epithelium. Besides activating stemness factors, it also directly represses genes promoting trophoblast cell fusion.

Substituting Angiotensin-(1-7) to Prevent Lung Damage in SARS-CoV-2 Infection?

Substituting Angiotensin-(1-7) to Prevent Lung Damage in SARS-CoV-2 Infection?

Curcumol attenuates liver sinusoidal endothelial cell angiogenesis via regulating Glis‐PROX1‐HIF‐1α in liver fibrosis

ObjectiveHepatic sinusoidal angiogenesis owing to dysfunctional liver sinusoidal endothelial cells (LSECs) accompanied by an abnormal angioarchitecture is a symbol related to liver fibrogenesis, which indicates a potential target for therapeutic interventions. However, there are few researches connecting angiogenesis with liver fibrosis, and the deeper mechanism remains to be explored.Materials and MethodsCell angiogenesis and angiogenic protein were examined in primary LSECs of rats, and multifarious cellular and molecular assays revealed the efficiency of curcumol intervention in fibrotic mice.ResultsWe found that curcumol inhibited angiogenic properties through regulating their upstream mediator hypoxia‐inducible factor‐1α (HIF‐1α). The transcription activation of HIF‐1α was regulated by hedgehog signalling on the one hand, and the protein stabilization of HIF‐1α was under the control of Prospero‐related homeobox 1 (PROX1) on the other. A deubiquitinase called USP19 could be recruited by PROX1 and involved in ubiquitin‐dependent degradation of HIF‐1α. Furthermore, our researches revealed that hedgehog signalling participated in the activation of PROX1 transcription probably in vitro. Besides, curcumol was found to ameliorate liver fibrosis and sinusoid angiogenesis via hedgehog pathway in carbon tetrachloride (CCl4) induced liver fibrotic mice. The protein expression of key regulatory factors, PROX1 and HIF‐1α, was consistent with the Smo, the marker protein of Hh signalling pathway.ConclusionsIn this article, we evidenced that curcumol controlling LSEC‐mediated angiogenesis could be a promising therapeutic approach for liver fibrosis.

Transdifferentiation of α-1,3-galactosyltransferase knockout pig bone marrow derived mesenchymal stem cells into pancreatic β-like cells by microenvironment modulation.

ObjectiveTo evaluate the pancreatic differentiation potential of α-1,3-galactosyltransferase knockout (GalTKO) pig-derived bone marrow-derived mesenchymal stem cells (BM-MSCs) using epigenetic modifiers with different pancreatic induction media.MethodsThe BM-MSCs have been differentiated into pancreatic β-like cells by inducing the overexpression of key transcription regulatory factors or by exposure to specific soluble inducers/small molecules. In this study, we evaluated the pancreatic differentiation of GalTKO pig-derived BM-MSCs using epigenetic modifiers, 5-azacytidine (5-Aza) and valproic acid (VPA), and two types of pancreatic induction media – advanced Dulbecco’s modified Eagle’s medium (ADMEM)-based and N2B27-based media. GalTKO BM-MSCs were treated with pancreatic induction media and the expression of pancreas-islets-specific markers was evaluated by real-time quantitative polymerase chain reaction, Western blotting, and immunofluorescence. Morphological changes and changes in the 5′-C-phosphate-G-3′ (CpG) island methylation patterns were also evaluated.ResultsThe expression of the pluripotent marker (POU class 5 homeobox 1 [OCT4]) was upregulated upon exposure to 5-Aza and/or VPA. GalTKO BM-MSCs showed increased expression of neurogenic differentiation 1 in the ADMEM-based (5-Aza) media, while the expression of NK6 homeobox 1 was elevated in cells induced with the N2B27-based (5-Aza) media. Moreover, the morphological transition and formation of islets-like cellular clusters were also prominent in the cells induced with the N2B27-based media with 5-Aza. The higher insulin expression revealed the augmented trans-differentiation ability of GalTKO BM-MSCs into pancreatic β-like cells in the N2B27-based media than in the ADMEM-based media.Conclusion5-Aza treated GalTKO BM-MSCs showed an enhanced demethylation pattern in the second CpG island of the OCT4 promoter region compared to that in the GalTKO BM-MSCs. The exposure of GalTKO pig-derived BM-MSCs to the N2B27-based microenvironment can significantly enhance their trans-differentiation ability into pancreatic β-like cells.

Chitin binding protein as a possible RNA binding protein in Leishmania parasites.

Leishmaniasis includes a broad spectrum of pathological outcomes in humans caused by protozoan parasites from the genus Leishmania. In recent years, proteomic techniques have introduced novel proteins with critical functions in Leishmania parasites. Based on our report of a Chitin binding protein (CBP) in our previous immunoproteomic study, this article suggests that CBP might be an RNA binding protein (RBP) in Leishmania parasites. RBPs, as key regulatory factors, have a role in post-transcriptional gene regulation. The presence of RBPs in Leishmania parasites has not been considered so far; however, this study aims to open a new venue regarding RBPs in Leishmania parasites. Confirming CBP as an RBP in Leishmania parasites, exploring other RBPs and their functions might lead to interesting issues in leishmaniasis. In fact, due to the regulatory role of RBPs in different diseases including cancers and their further classification as therapeutic targets, the emerging evaluation of CBP and RBPs from Leishmania parasites may allow the discovery of novel and effective drugs against leishmaniasis.

Inhibition of FGFR Signaling Partially Rescues Osteoarthritis in Mice Overexpressing High Molecular Weight FGF2 Isoforms.

Fibroblast growth factor 2 (FGF2) and fibroblast growth factor receptors (FGFRs) are key regulatory factors in osteoarthritis (OA). HMWTg mice overexpress the high molecular weight FGF2 isoforms (HMWFGF2) in osteoblast lineage and phenocopy both Hyp mice (which overexpress the HMWFGF2 isoforms in osteoblasts and osteocytes) and humans with X-linked hypophosphatemia (XLH). We previously reported that, similar to Hyp mice and XLH subjects who develop OA, HMWTg mice also develop an OA phenotype associated with increased degradative enzymes and increased FGFR1 compared with VectorTg mice. Therefore, in this study, we examined whether in vivo treatment with the FGFR tyrosine kinase inhibitor NVP-BGJ398 (BGJ) would modulate development of the OA phenotype in knee joints of HMWTg mice. VectorTg and HMWTg mice (21 days of age) were treated with vehicle or BGJ for 13 weeks. Micro-computed tomography images revealed irregular shape and thinning of the subchondral bone with decreased trabecular number and thickness within the epiphyses of vehicle-treated HMWTg knees, which was partially rescued following BGJ treatment. Articular cartilage thickness was decreased in vehicle-treated HMWTg mice, and was restored to the cartilage thickness of VectorTg mice in the BGJ-treated HMWTg group. Increased OA degradative enzymes present in HMWTg vehicle-treated joints decreased after BGJ treatment. OA in HMWTg mice was associated with increased Wnt signaling that was rescued by BGJ treatment. This study demonstrates that overexpression of the HMWFGF2 isoforms in preosteoblasts results in osteoarthropathy that can be partially rescued by FGFR inhibitor via reduction in activated Wnt signaling.

Prohibitin (PHB) interacts with AKT in mitochondria to coordinately modulate sperm motility.

Prohibitin (PHB), an evolutionarily conserved mitochondrial inner membrane protein, is highly expressed in cells that require strong mitochondrial function. Recently, we demonstrated that the deletion of Phb in spermatocytes results in impaired mitochondrial function. In addition, PHB expression in the mitochondrial sheath of human sperm has a significantly negative correlation with mitochondrial reactive oxygen species levels, but a positive one with mitochondrial membrane potential and sperm motility. These results suggest that mitochondrial PHB expression plays a role in sperm motility. However, the mechanism of PHB-mediated regulation of sperm motility remains unknown. Here, we demonstrate for the first time that PHB interacts with protein kinase B (AKT) and exists in a complex with phospho-PHB (pT258) and phospho-AKT in the mitochondrial sheath of murine sperm, as determined using colocalization and coimmunoprecipitation assays. After blocking AKT activity using wortmannin (a phosphatidylinositol 3-kinase [PI3K] inhibitor), murine sperm have significantly (P < 0.05) decreased levels of phospho-PHB (pT258) and the total and progressive motility. Furthermore, significantly (P < 0.05) lower levels of phospho-PI3K P85 subunit α+γ (pY199 and pY467) and phospho-AKT (pS473; pT308) are found in sperm from infertile asthenospermic and oligoasthenospermic men compared with normospermic subjects, which suggest a reduced activity of the PI3K/AKT pathway in these infertile subjects. Importantly, these sperm from infertile subjects also have a significantly (P < 0.05) lower level of phospho-PHB (pT258). Collectively, our findings suggest that the interaction of PHB with AKT in the mitochondrial sheath is critical for sperm motility, where PHB phosphorylation (pT258) level and PI3K/AKT activity are key regulatory factors.

Landscape inferred from gene expression data governs pluripotency in embryonic stem cells

Embryonic stem cells (ESCs) can differentiate into diverse cell types and have the ability of self-renewal. Therefore, the study of cell fate decisions on embryonic stem cells has far-reaching significance for regenerative medicine and other biomedical fields. Mathematical models have been used to study emryonic stem cell differentiation. However, the underlying mechanisms of cell differentiation and lineage reprogramming remain to be elucidated. Especially, how to integrate the computational models with quantitative experimental data is still challenging. In this work, we developed a data-constrained modelling approach, and established a model of mouse embryonic stem cells. We used the truncated moment equations (TME) method to quantify the potential landscape of the ESC network. We identified two attractors on the landscape, which represent the embryonic stem cell (ESC) state and differentiated cell (DC) state, respectively, and quantified high dimensional biological paths for differentiation and reprogramming process. Through identifying the optimal combinations of gene targets based on a landscape control strategy, we offered some predictions about the key regulatory factors that govern the differentiation and reprogramming in ESCs.

Establishment of porcine nuclear transfer-derived embryonic stem cells using induced pluripotent stem cells as donor nuclei.

We investigated whether sequential reprogramming via porcine induced pluripotent stem cells (piPSCs) or exposure to oocyte cytoplasm following nuclear transfer could generate nuclear transfer-derived ESCs (piPSCs-ntESCs). Nuclear transfer embryos were reconstructed with piPSCs possessing a ZsGreen fluorescent marker for expression of exogenous Nanog and Lin28. Reconstructed oocytes developed to morphologically normal 8-cell/morulae (35/93, 37.6%) and blastocysts (12/93, 12.9%). Although most green fluorescent protein-positive blastocysts showed efficient outgrowth (8/10, 80%), none formed primary colonies and all cultures degenerated. Conversely, 15% of fluorescent positive 8-cell/morula stage embryos showed outgrowth (6/40), with three forming primary colonies (7.5%). All three were expanded and maintained as piPSC-ntESC lines. These cell lines expressed stem cell marker genes and proteins. Despite inactivation of one X chromosome, all piPSC-ntESC lines formed teratomas comprising derivatives from all three embryonic germ layers. Strong SSEA1, 3, and 4 expression was detected at the 8-cell/morula stage in embryos reconstructed from both piPSCs and porcine embryonic fibroblasts (PEFs). SSEA3 was notably absent from IVF controls at pre-implantation embryo stages. Finally, we attempted to establish ntESCs from 8-cell/morulae reconstructed with PEFs using the same culture conditions as those for piPSC-ntESC derivation. Although eight primary colonies arose from 107 embryos (7.5%), they all degenerated after the first passage culture. Early and sustained expression of key reprogramming regulatory factors may be critical for pluripotent stem cell derivation to derive piPSC-ntESCs from 8-cell/morula stages, while the expression of SSEAs may be involved in the initial stem cell colony formation phases.

Targeted suppression of metastasis regulatory transcription factor SOX2 in various cancer cell lines using a sequence-specific designer pyrrole–imidazole polyamide

Metastasis, a deadly feature of cancer, compromises the prognosis and accounts for mortality in the majority of cancer patients. SOX2, a well-known pluripotency transcription factor, plays a central role in cell fate determination and has an overlapping role as a regulatory factor in tumorigenesis and metastasis. The demand is increasing for clinically useful strategies for artificial control of SOX2 expression and its complex transcription machinery in cancer cells. N-Methylpyrrole (Py) and N-methylimidazole (Im) polyamides are small programmable designer ligands that can be pre-programmed to selectively recognize DNA sequence and control endogenous gene expression. Herein, we evaluated the anticancer activity of a designer ligand (SOX2i). SOX2i remarkably altered the expression of SOX2 at the mRNA and protein level in human cancer cell lines such as SW620 (colorectal adenocarcinoma), MKN45 (gastric adenocarcinoma), MCF7 (breast carcinoma), U2OS (osteosarcoma) and other cancer cell lines of different origin and type. Genome-wide transcriptome analysis and cell-based assays showed SOX2 to be a downregulated upstream regulator that alters cell proliferation, cell cycle progression, metabolism and apoptotic pathway. Studies in the mouse model confirmed the anti-metastatic property of SOX2i. SOX2i inhibited the expression of genes associated with EMT and stemness. Moreover, Wnt-canonical signaling was found to be downregulated in the SOX2i-treated group. Our proof-of-concept study supports the potential of DNA-based programmable small molecules for controlling the key regulatory factors associated with tumorigenesis and metastasis.

PSVII-36 Examining the effect of a physiological dose of the polyamine; spermine, on myogenic regulatory transcription factor expression

Abstract Examining the effect of a physiological dose of the polyamine; spermine, on myogenic regulatory transcription factor expression. Spermine is a micronutrient derived from amino acids. It affects cell growth, proliferation, differentiation and gene regulation in many cell types. Polyamines have long been overlooked with respect to their biological effects on muscle growth. Myogenic regulatory factors (myoD, myf5) initiate, promote and regulate myogenesis. Supporting myogenic transcription factors transcription and by such enhancing muscle production in livestock, through nutrition may be possible with dietary spermine supplementation. We examined the effect of spermine (0.5mM) in undifferentiated c2c12 muscle cells at two time points (8hr and 16hr). Using TaqMan-MGB qRT-PCR we quantified mRNA for key myogenic regulatory factors in a minimum of three experiments each containing 3 technical replicates. We report a significant increase in myoD (P = 0.02) and myf5 (P = 0.05) mRNA at 8hr following spermine treatment as compared to controls (no spermine). After 16 hr exposure to spermine (0.5mM) treatment myoD (P = 0.01) remained significantly different from controls. Our results indicate that spermine supports myogenesis through expression of increased myogenic regulatory factors at early stages of myogenesis. These findings support the need to further test the hypothesis that spermine promotes increased muscle growth and ultimately may represent a dietary means to maximize muscle growth in livestock species.

Abnormal expression of miR-135b-5p in bone tissue of patients with osteoporosis and its role and mechanism in osteoporosis progression.

Osteoporosis (OP) is an age-related bone disease occurring worldwide. Osteoporotic fracture is one of the leading causes of disability and death in elderly patients. MicroRNAs (miRNAs/miRs) are key molecular regulatory factors in bone remodeling processes. The present study investigated the expression and mechanism of miR-135b-5p in patients with osteoporosis. The present results suggested that miR-135b-5p was upregulated in bone tissue fragments of patients with osteoporosis compared with the control patients. MC3T3-E1 cells were used to perform osteogenic differentiation induction. Reverse transcription-quantitative PCR and western blot assay were used to detect the mRNA and protein expression levels of the osteogenic markers osteocalcin (OC), Osterix and alkaline phosphatase (ALP). A specific kit was used for detecting ALP activity. The present results indicated that the mRNA expression levels of OC, Osterix and ALP significantly increased on the 7 and 14th day after osteogenic differentiation induction compared with the control group. Protein expression levels of OC, Osterix and ALP also increased on the 7 and 14th day after induction. ALP assay showed that ALP activity was significantly increased on the 7 and 14th day after induction. In addition, the present study found that miR-135b-5p was downregulated in MC3T3-E1 cells 7 and 14 days after osteogenic differentiation induction. The results of TargetScan analysis and dual luciferase reporter gene assay indicated that runt-related transcription factor 2 (RUNX2) was a direct target gene of miR-135b-5p. RUNX2 was upregulated in MC3T3-E1 cells on the 7 and 14th day after induction. Moreover, the present study found that compared with the osteogenic differentiation induction group, miR-135b-5p mimic significantly decreased OC, Osterix and ALP expression, and reduced ALP activity in MC3T3-E1 cells. However, these reductions were reversed following overexpression of RUNX2. The present results showed that miR-135b-5p mimic significantly reduced cell viability in MC3T3-E1 cells and induced cell apoptosis, and these effects were significantly reversed following RUNX2 overexpression. In summary, the present results suggested that miR-135-5p participated in the occurrence and development of osteoporosis via inhibition of osteogenic differentiation and osteoblast growth by targeting RUNX2. The present study suggested a novel potential target that may faciliate the treatment of osteoporosis, and further study is required to examine this possibility.

STAT3 speeds up progression of osteoarthritis through NF‑κB signaling pathway

Osteoarthritis (OA) is the most common motor system disease in the elderly, with a high incidence and a huge social and economic burden. Therefore, it is urgent to study its potential pathogenesis to improve the therapeutic effect of the disease. In this study, we constructed a number of regulator-mediated OA dysfunction modules, and carried out in-depth analysis in order to examine the disease development process. Differential expression analysis, co-expression analysis and enrichment analysis were combined to screen genes related to disease progression. Subsequently, key regulatory factors in the process of OA were identified based on the pivotal regulators that may manipulate important parts of the module subnetwork. A total of 16 OA dysfunction modules were obtained, involving the aggregation of 3,239 module genes. Then, enrichment analysis showed that module genes were significantly involved in apoptosis, inflammation-related functions and signaling pathways. Finally, we revealed a series of regulators, including 842 ncRNA (miR-132-3p, miR-130a-3p and miR-590-3p), 59 transcription factors (NFKB1, RELA and STAT3). We consider that STAT3 is the core transcription factor and promotes the development of OA through the signal of NF-κB. Overall, our results provide biologists and pharmacists with a new way of thinking to reveal the disease process of OA, and provide a wider range of candidate targets for follow-up research.

Integration of multi-omics data to mine cancer-related gene modules.

The identification of cancer-related genes is a major research goal, with implications for determining the pathogenesis of cancer and identifying biomarkers for early diagnosis and treatment. In this study, by integrating multi-omics data, including gene expression, DNA copy number variation, DNA methylation, transcription factors, miRNA, and lncRNA data, we propose a method for mining cancer-related genes based on network models. First, using random forest-based feature selection method multi-omics data are integrated to identify key regulatory factors that affect gene expression, and then genome-wide regulatory networks are constructed. Next, by comparing the regulatory networks of key candidate genes in variant samples and non-variant samples, a differential expression regulatory network is generated. The differential network contains a collection of abnormal regulatory genes of key candidate genes. Then, by introducing the functional similarity as a distance metric for gene sets, a density-based clustering method is used to mine gene modules related to cancer. We applied this method to LUSC (lung squamous cell carcinoma) and mined cancer-related gene modules composed of 20 genes. GO function and KEGG pathway analyses indicated that the modules were closely related to cancer. A survival analysis was used to verify that the excavated gene modules can effectively distinguish between high- and low-risk groups. Overall, these results suggest that the proposed method can be used to identify cancer-related gene modules, providing a basis for the development of biomarkers for diagnosis and treatment.

LncRNA Expression Reveals the Potential Regulatory Roles in Hepatocyte Proliferation during Rat Liver Regeneration.

Liver regeneration is a tissue growth process after loss or injury of liver tissue, which is a compensatory hyperplasia rather than true regeneration, mainly depending on hepatocyte proliferation. Currently, a large number of studies on hepatocyte proliferation have been conducted. However, studies on the regulation of long noncoding RNA (lncRNA) on hepatocyte proliferation are still limited. To identify specially expressed lncRNA during rat liver regeneration, high-throughput sequencing technology was performed, and a total of 2446 lncRNAs and 4091 mRNAs were identified as significantly differentially expressed. Gene ontology (GO) enrichment analysis was performed to analyze the role of differentially expressed mRNAs, and 695 mRNAs were identified to be related to cell proliferation. Then, an lncRNA-mRNA coexpression network based on the differentially expressed lncRNAs and proliferation-related genes was constructed to analyze the potential function of lncRNAs on hepatocyte proliferation, and ten lncRNAs, NONRATT003557.2, NONRATT005357.2, NONRATT003292.2, NONRATT001466.2, NONRATT003289.2, NONRATT001047.2, NONRATT005180.2, NONRATT004419.2, NONRATT005336.2, and NONRATT005335.2, were selected as key regulatory factors, which may play crucial roles in hepatocyte proliferation during rat liver regeneration. Finally, a protein-protein interaction (PPI) network was established to illuminate the interaction between proliferation-related genes, and ten hub genes (Aurkb, Cdk1, Cdc20, Bub1b, Mad2l1, Kif11, Prc1, Ccna2, Top2a, and Ccnb1) were screened with the MCC method in the PPI network, which may be important biomarkers involved in the hepatocyte proliferation during rat liver regeneration. These results may provide clues for a more comprehensive understanding of the molecular mechanism of hepatocyte proliferation during rat liver regeneration.

Reprogramming of Fibroblasts to Oligodendrocyte Progenitor-like Cells Using CRISPR/Cas9-Based Synthetic Transcription Factors.

SummaryCell lineage reprogramming via transgene overexpression of key master regulatory transcription factors has been well documented. However, the poor efficiency and lack of fidelity of this approach is problematic. Synthetic transcription factors (sTFs)—built from the repurposed CRISPR/Cas9 system—can activate endogenous target genes to direct differentiation or trigger lineage reprogramming. Here we explored whether sTFs could be used to steer mouse neural stem cells and mouse embryonic fibroblasts toward the oligodendrocyte lineage. We developed a non-viral modular expression system to enable stable multiplex delivery of pools of sTFs capable of transcriptional activation of three key oligodendrocyte lineage master regulatory genes (Sox10, Olig2, and Nkx6-2). Delivery of these sTFs could enhance neural stem cell differentiation and initiated mouse embryonic fibroblast direct reprograming toward oligodendrocyte progenitor-like cells. Our findings demonstrate the value of sTFs as tools for activating endogenous genes and directing mammalian cell-type identity.

SigHotSpotter: scRNA-seq-based computational tool to control cell subpopulation phenotypes for cellular rejuvenation strategies.

SummarySingle-cell RNA-sequencing is increasingly employed to characterize disease or ageing cell subpopulation phenotypes. Despite exponential increase in data generation, systematic identification of key regulatory factors for controlling cellular phenotype to enable cell rejuvenation in disease or ageing remains a challenge. Here, we present SigHotSpotter, a computational tool to predict hotspots of signaling pathways responsible for the stable maintenance of cell subpopulation phenotypes, by integrating signaling and transcriptional networks. Targeted perturbation of these signaling hotspots can enable precise control of cell subpopulation phenotypes. SigHotSpotter correctly predicts the signaling hotspots with known experimental validations in different cellular systems. The tool is simple, user-friendly and is available as web-server or as stand-alone software. We believe SigHotSpotter will serve as a general purpose tool for the systematic prediction of signaling hotspots based on single-cell RNA-seq data, and potentiate novel cell rejuvenation strategies in the context of disease and ageing.Availability and implementationSigHotSpotter is at https://SigHotSpotter.lcsb.uni.lu as a web tool. Source code, example datasets and other information are available at https://gitlab.com/srikanth.ravichandran/sighotspotter.Supplementary information Supplementary data are available at Bioinformatics online.

Identification of key genes, regulatory factors, and drug target genes of recurrent implantation failure (RIF)

Objective: Recurrent implantation failure (RIF) exacerbates the physical trauma of infertile women that undergone in vitro fertilization-embryo transfer (IVF-ET). We aimed to identify the key genes, regulatory factors, and drug target genes involved in the RIF.Methods: The dataset GSE58144 that obtained from the Gene Expression Omnibus mainly contained 43 RIF and 72 control endometrial samples. Differently expressed genes (DEGs) between RIF and control groups were firstly analyzed, followed by the pathway and Gene Ontology (GO) enrichment analysis. Then, protein-protein interaction (PPI) network and miRNA-transcript factor (TF)-DEGs network were established. Finally, a drug-target interaction network was constructed.Results: A total of 399 DEGs were identified between the RIF and controls. In the PPI and key module network, UBE2I, PLK4, XPO1, AURKB, and NUP107 were identified as the hub genes, which mainly enriched in RNA transport and cell division cycle-related pathways and GO items. In the miRNA-TF-DEGs network, E2F4, SIN3A, miRNA489, miRNA199A, miRNA369-3P, miRNA422, and miRNA522 were considered as the key regulatory factors during RIF. In addition, HTR1A, NR3C1, and GABRA3 were the main targets of the drugs annotated in DrugBank.Conclusion: The effects of PLK4, XPO1, AURKB, and NUP107 on the RIF may be via affecting the proliferation and differentiation of endometrial stromal cells. Besides, SIN3A and miRNA199A may be crucial for embryo implantation. In addition, NR3C1 may be used as a possible target for the clinical therapy of RIF.

Identification and Temporal Expression Analysis of Conserved and Novel MicroRNAs in the Leaves of Winter Wheat Grown in the Field.

Cold acclimation and vegetative/reproductive transition are two important evolutionary adaptive mechanisms for winter wheat surviving the freezing temperature in winter and successful seeds setting in the next year. MicroRNA (miRNA) is a class of regulatory small RNAs (sRNAs), which plays critical roles in the growth and development of plants. However, the regulation mechanism of miRNAs during cold acclimation and vegetative/reproductive transition of winter wheat is not much understood. In this study, four sRNA libraries from leaves of winter wheat grown in the field at the three-leaf stage, winter dormancy stage, spring green-up stage, and jointing stage were analyzed to identify known and novel miRNAs and to understand their potential roles in the growth and development of winter wheat. We examined miRNA expression using a high-throughput sequencing technique. A total of 373 known, 55 novel, and 27 putative novel miRNAs were identified. Ninety-one miRNAs were found to be differentially expressed at the four stages. Among them, the expression of six known and eight novel miRNAs was significantly suppressed at the winter dormancy stage, whereas the expression levels of seven known and eight novel miRNAs were induced at this stage; three known miRNAs and three novel miRNAs were significantly induced at the spring green-up stage; six known miRNAs were induced at the spring green-up stage and reached the highest expression level at the jointing stage; and 20 known miRNAs and 10 novel miRNAs were significantly induced at the jointing stage. Expression of a number of representative differentially expressed miRNAs was verified using quantitative real-time polymerase chain reaction (qRT-PCR). Potential target genes for known and novel miRNAs were predicted. Moreover, six novel target genes for four Pooideae species-specific miRNAs and two novel miRNAs were verified using the RNA ligase-mediated 5′-rapid amplification of cDNA ends (RLM-5’RACE) technique. These results indicate that miRNAs are key non-coding regulatory factors modulating the growth and development of wheat. Our study provides valuable information for in-depth understanding of the regulatory mechanism of miRNAs in cold acclimation and vegetative/reproductive transition of winter wheat grown in the field.