Main Target Gene Research Articles

Role of miRNAs in the pathogenesis of T2DM, insulin secretion, insulin resistance, and β cell dysfunction: the story so far.

Diabetes, the most common endocrine disorder, also known as a silent killer disease, is characterized by uncontrolled hyperglycemia. According to the International Diabetes Federation, there were 451 million people with diabetes mellitus worldwide in 2017. It is a multifactorial syndrome caused by genetic as well as environmental factors. Noncoding RNAs, especially the miRNAs, play a significant role in the development as well as the progression of the disease. This is on account of insulin resistance or defects in β cell function. Various miRNAs including miR-7, miR-9, miR-16, miR-27, miR-24, miR-29, miR-124a, miR-135, miR-130a, miR-144, miR-181a, and miR-375 and many more have been associated with insulin resistance and other pathogenic conditions leading to the development of the disease. These miRNAs play significant roles in various pathways underlying insulin resistance such as PI3K, AKT/GSK, and mTOR. The main target genes of these miRNAs are FOXO1, FOXA2, STAT3, and PTEN. The miRNAs carry out important functions in insulin target tissues like the adipose tissue, liver, and muscle. MiRNAs miR-9, miR-375, and miR-124a, are also associated with the secretion of insulin from pancreatic cells. There is an interplay between the miRNAs and pancreatic cell growth, especially the miRNAs affecting development and proliferation of these cells. Most of the miRNAs target more than one gene which not only justifies their use as biomarkers but also their therapeutic potential. The current review has been compiled with an aim to discuss the role of various miRNAs involved in various pathogenic mechanisms including insulin resistance, insulin secretion, and the β cell dysfunction.

A Systematic Review and Bioinformatics Study on Genes and micro-RNAs Involving the Transformation of Endometriosis into Ovarian Cancer.

BackgroundAlong with the description of tumorigenesis processes in endometriosis-related ovarian cancer, identifying dysregulated miRNAs, the target genes of these miRNAs, and the processes abnormally affected by dysregulated miRNAs is essential, which was our goal.MethodsTwo reviewers individually evaluated the articles which collected relevant information including genes and miRNAs involved in the transformation of endometriosis into ovarian cancer. To assess the mature sequence of miRNAs and also their chromosomal positions, miRPathDB software was employed. To determine the main target gene predicted for each considered miRNAs, the TargetScanS Web server was applied. The interaction of each gene with other genes associated with endometrial-related ovarian cancer was determined by GeneMANIA software. Finally, to design integrated model of miRNAs-targeted genes interaction network, the Cytoscape software was used.ResultsThe final number of studies available for analysis was 6 manuscripts including 22 miRNAs described as involved in the transformation of endometriosis into different subtypes of ovarian cancers (14 miRNAs up-regulated and 8 miRNAs down-regulated). Three miRNAs of miR-141 (up-regulated), miR-205 (down-regulated), and miR-125b (down-regulated) were revealed as the originator for genetic interactions leading to carcinogenesis. We could show some common loops and pathways including uncontrolled cell proliferation and abnormal apoptosis (mediated by PTEN gene induced by miR-21 and miR-214), and disaggregation and epithelialization (mediated by ZEB1 and ZEB2 genes induced by miR-200).ConclusionAccording to our analysis, up-regulation of miR-141 and down-regulation of miR-205 and miR-125b have a central role in transforming endometriosis to ovarian cancer.

Tolloid-Like 1 Negatively Regulates Hepatic Differentiation of Human Induced Pluripotent Stem Cells Through Transforming Growth Factor Beta Signaling.

Single nucleotide polymorphisms in Tolloid‐like 1 (TLL1) and the expression of TLL1 are known to be closely related to hepatocarcinogenesis after hepatitis C virus elimination or liver fibrosis in patients with nonalcoholic fatty liver disease. TLL1 is a type of matrix metalloprotease and has two isoforms in humans, with the short isoform showing higher activity. However, the functional role of TLL1 in human liver development is unknown. Here, we attempted to elucidate the function of human TLL1 using hepatocyte‐like cells generated from human pluripotent stem cells. First, we generated TLL1‐knockout human induced pluripotent stem (iPS) cells and found that hepatic differentiation was promoted by TLL1 knockout. Next, we explored TLL1‐secreting cells using a model of liver development and identified that kinase insert domain receptor (FLK1)‐positive cells (mesodermal cells) highly express TLL1. Finally, to elucidate the mechanism by which TLL1 knockout promotes hepatic differentiation, the expression profiles of transforming growth factor beta (TGFβ), a main target gene of TLL1, and its related genes were analyzed in hepatic differentiation. Both the amount of active TGFβ and the expression of TGFβ target genes were decreased by TLL1 knockout. It is known that TGFβ negatively regulates hepatic differentiation. Conclusion: TLL1 appears to negatively regulate hepatic differentiation of human iPS cells by up‐regulating TGFβ signaling. Our findings will provide new insight into the function of TLL1 in human liver development.

Study on the Pharmacological Mechanism of Yinqiao Antipyreti Granules Against Influenza Based on Network Pharmacology

Objective: To explore the anti-influenza pharmacological mechanism of Yinqiao Antipyreti Granules by using network pharmacology. Method: Retrieve the main active ingredients, corresponding targets and target genes of Yinqiao Antipyreti Granules on the website of Chinese Medicine System Pharmacology Technology Platform (TCMSP), obtain the relevant target genes of influenza through the Human Gene Database (Gene Cards), and transfer the drugs The target of the active ingredient is mapped to the influenza target, and the intersection target obtained is the predicted target of Yinqiao Ture granules against influenza. Cytoscape 3.8.0 software was used to construct a drug active ingredient-intersection target network model, and key active ingredients were selected. Use the STRING website to construct an intersection target protein interaction network (PPI) and select key target genes. Enriched analysis of GO and KEGG pathways using Metascape on intersection targets. Results: There are 136 active ingredients in Yinqiao Antipyretic Granules and 131 anti-influenza-related targets. Conclusion: The main components of Yinqiao Antipyretic Granules are quercetin, luteolin, naringenin, formononetin, IL-6, STAT3, MAPK14, CASP3 and other gene targets, to suppress inflammatory response, regulate immunity, reduce the symptoms of influenza, and thus play an anti-influenza therapeutic role.

Pharmacological mechanism of Astragalus and Angelica in the treatment of idiopathic pulmonary fibrosis based on network pharmacology

IntroductionHerbal medicine is varied and complex, and research on multi-component, multi-target and multi-pathways of Astragalus and Angelica is lacking. This study aimed to study the pharmacological mechanism of Astragalus and Angelica in the treatment of idiopathic pulmonary fibrosis (IPF) using on network pharmacology. MethodsThe main active components, corresponding targets and target genes of Astragalus and Angelica were searched by TCMSP and UniProt database. The target genes of IPF were obtained by GeneCards database and the target genes of active components were intersected with IPF target genes to obtain predictive targets of Astragalus and Angelica acting on IPF. The medicine-compound-target-disease network was constructed by Cytoscape3.6.0 software. The protein protein interaction network was constructed by STRING database to select the key target genes. The DAVID database and KEGG PATHWAY Database were used to analysis the gene ontology (GO) enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment. ResultsSixty-nine overlapping genes were obtained by intersecting 100 compound target genes with 2231 IPF target genes, corresponding to 17 effective compounds, including 15 compounds from Astragalus and two compounds from Angelica. GO enrichment showed the main biological functions of potential genes of Astragalus and Angelica in the treatment of IPF. KEGG pathway enrichment showed the main pathways of Astragalus and Angelica in the treatment of IPF. ConclusionIn this study, the target and mechanism of the components of Astragalus and Angelica in the treatment of IPF have been systematically discussed, and have provided ideas for future clinical research.

Network Pharmacology Analysis of Traditional Chinese Medicine Formula Xiao Ke Yin Shui Treating Type 2 Diabetes Mellitus.

Xiao Ke Yin Shui (XKYS) formula is a traditional Chinese medicine formula treating type 2 diabetes mellitus (T2DM). XKYS formula consists of four herbs, i.e., Coptidis rhizoma, Liriopes radix, bitter melon, and Cassiae semen. Herein, the chemical profiles of four herb extracts were investigated, and further analysis of the underlying mechanism of XKYS formula treating T2DM was performed using network pharmacology. The main components were selected for our network-based research. Targets of XKYS formula were mainly collected from two databases, SwissTargetPrediction and Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP), and the text-mining method was also implemented. T2DM relating genes and therapeutic targets were collected from five databases. Subsequently, STRING and Cytoscape were employed for the analysis of protein-protein interaction (PPI) networks. Functional annotation and pathway analysis were conducted to investigate the functions and relating pathways of target genes. The content of 12 compounds in the herb extracts was determined. With the analysis of PPI networks, a total of 76 genes were found to be important nodes and could be defined as the main target genes regulated by XKYS formula in the treatment of T2DM and its complications. Components in XKYS formula mainly regulate proteins including protein kinase B (Akt), phosphatidylinositol 3-kinase (PI3K), insulin receptor substrate (IRS), and tumor necrosis factor (TNF). XKYS formula exerts therapeutic effects in a synergetic manner and exhibits antidiabetic effect mainly via reducing insulin resistance. These findings could be guidelines in the further investigation of this formula.

Indoxyl Sulfate Stimulates Angiogenesis by Regulating Reactive Oxygen Species Production via CYP1B1.

Indoxyl sulfate (IS) is an accumulative protein-bound uremic toxin found in patients with kidney disease. It is reported that IS impairs the vascular endothelium, but a comprehensive overview of all mechanisms active in IS-injury currently remains lacking. Here we performed RNA sequencing in human umbilical vein endothelial cells (HUVECs) after IS or control medium treatment and identified 1293 genes that were affected in a IS-induced response. Gene enrichment analysis highlighted pathways involved in altered vascular formation and cell metabolism. We confirmed these transcriptome profiles at the functional level by demonstrating decreased viability and increased cell senescence in response to IS treatment. In line with the additional pathways highlighted by the transcriptome analysis, we further could demonstrate that IS exposure of HUVECs promoted tubule formation as shown by the increase in total tubule length in a 3D HUVECs/pericytes co-culture assay. Notably, the pro-angiogenic response of IS and increased ROS production were abolished when CYP1B1, one of the main target genes that was highly upregulated by IS, was silenced. This observation indicates IS-induced ROS in endothelial cells is CYP1B1-dependent. Taken together, our findings demonstrate that IS promotes angiogenesis and CYP1B1 is an important factor in IS-activated angiogenic response.

Abstract 2971: Ellagic acid and its metabolite Urolithin A induce prostate cancer cell death in p53 dependent and independent manner

Abstract Carcinoma of the prostate (CaP) is the most common cancer in men and the second leading cause of cancer-related death in men worldwide. Treatment of early stages of CaP often involves the surgical removal of all or part of the prostate gland; whereas malignant cases of advanced CaP generally requires a combination of therapies including removal of the entire prostate, radiotherapy and androgen deprivation. However, significant number of CaP patients relapse the CaP as the disease becomes hormonal independent. Therefore, it is important to target cell death pathways that function independently of androgen signaling. p53 is a common tumor suppressor that mediates apoptosis, cell cycle arrest and DNA repair. Inactivation of p53 is common in CaP cells and this repression of p53 function diminishes androgen receptor signaling. The most common negative regulator of p53 is MDM2, which is itself a target gene of p53 to form an autoregulatory negative feedback loop. MDM2 inhibits p53 transcriptional activity through the induction of p53 polyubiquitination and degradation in the proteasome. This loss of p53 may permit CaP cells to undergo uncontrolled cell growth and cancer progression. Previous studies have shown that certain polyphenols, derived from natural products, possess anticancer activity that may be attributed to their repression of MDM2 ligase activity and activation of p53. Pomegranates, berries, and walnuts contain several bioactive compounds, including the Ellagitannins (ETs). ETs are polyphenolic compounds that are hydrolyzed in the stomach to form Ellagic acid (EA) which is itself metabolized in the gut microbiota to Urolithin A (UA). The purpose of this study was to investigate the influence of EA and UA on the p53-MDM2 signaling pathway in CaP cells. Three models of CaP cell lines were used because each harbor different p53 genotypes: LNCaP (p53+/+), 22RV1(p53-/+) and PC3 (p53-/-). Here we found that, when 22RV1 and LNCaP were treated with EA and UA, the interaction between p53 and MDM2 was disrupted. As a result, both EA and UA caused an increase in p53 protein levels and increased the steady-state concentration of p21, a main downstream target gene of p53 that mediates cell cycle arrest. In addition, EA and UA increased the levels of PUMA and NOXA proteins, both target genes of p53 which confer p53’s pro-apoptotic function. Moreover, we confirmed UA inhibits MDM2-mediated polyubiquitination and degradation of p53. Finally, the data show that EA and UA induce apoptosis in PC3 cells (p53-/-), indicating p53 independent role of these compounds. These results suggest that EA and UA may have potential anti-neoplastic activity in CaP cells that may be at least partially attributed to the stabilization and activation of p53. Citation Format: Yasir I. Mohammed Saleem, Mustafa Selim. Ellagic acid and its metabolite Urolithin A induce prostate cancer cell death in p53 dependent and independent manner [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2971.

Urolithin A induces prostate cancer cell death in p53-dependent and in p53-independent manner.

Pomegranate and walnuts are widely consumed dietary sources and contain several bioactive compounds, including the ellagitannins (ETs). ETs are polyphenols that are metabolized in the gut microbiota to urolithin A (UA). p53 is a tumor suppressor that lost its activity through MDM2 activation in about half cancers. The purpose of this study was to investigate the influence of UA on the p53-MDM2 interaction pathway in prostate cancer cell lines. Three human prostate cancer cell lines were used that harbor different p53 genotypes; LNCaP (p53+/+), 22RV1(p53-/+) and PC3 (p53-/-). Cell viability was determined by CellTiter-Glo Luminescent assay. Apoptosis was confirmed by measuring annexin V by flow cytometry. The expression of p53, its target proteins, and apoptotic markers were measured by western blotting. Real-time qPCR was used to measure the gene expression of p21, a main target gene of p53. Co-immunoprecipitation-immunoblotting was used to assess the inhibition of interactions between p53 and MDM2 and to assess the effect of UA on MDM2-mediated p53 polyubiquitination. We found UA inhibited CaP cells' viability and induced apoptosis. For 22RV1 and LNCaP, we found UA increased p53 protein expression and its main target protein, p21, and MDM2, forming an autoregulatory feedback loop. In addition, UA increased the p53 proapoptotic proteins PUMA and NOXA. Moreover, UA inhibited the interaction between p53 and MDM2 and inhibited MDM2-mediated p53 polyubiquitination. UA downregulated MDM2 and XIAP protein expression in PC3 cells and upregulated p21 and p14ARF in a p53-independent manner. The influencing of UA on p53-MDM2 pathway may partly contribute to its anticancer effect.

Dynamic changes in peripheral blood-targeted miRNA expression profiles in patients with severe traumatic brain injury at high altitude

BackgroundThe aim of this work is to detect and compare the peripheral blood miRNA expression profiles in patients with severe traumatic brain injury (sTBI) 2, 12, 24, 48, and 72 h after injury at high altitude and to predict the target genes of differential expressed miRNAs.MethodsTwenty sTBI patients from high-altitude areas were randomly selected according to the inclusion and exclusion criteria and were divided into five groups: the 2-h group, 12-h group, 24-h group, 48-h group, and 72-h group. Peripheral blood miRNA expression profiles were detected using real-time quantitative PCR (qRT-PCR).ResultsThe expression levels of miR-18a, miR-203, miR-146a, miR-149, miR-23b, and miR-let-7b in peripheral blood showed significant differences between the 2-h group and the 12-h group. The expression levels of miR-203, miR-146a, miR-149, miR-23b, and miR-let-7f in peripheral blood were up-regulated in the 24-h group. In the 48-h group, the expression levels of miR-181d, miR-29a, and miR-18b were upregulated. In the 72-h group, the expression levels of miR-203, miR-146a, miR-149, miR-23b, and miR-let-7f changed. The main target genes of the differentiation expressed miRNAs were genes that regulate inflammatory responses, apoptosis, and DNA damage/repair.ConclusionsmiRNAs may be involved in the pathogenesis of sTBI by dynamically regulating the target genes that regulate inflammatory responses, apoptosis, and DNA damage/repair pathways.

Expression, Localization, and Activity of the Aryl Hydrocarbon Receptor in the Human Placenta.

The human placenta is an organ between the blood of the mother and the fetus, which is essential for fetal development. It also plays a role as a selective barrier against environmental pollutants that may bypass epithelial barriers and reach the placenta, with implications for the outcome of pregnancy. The aryl hydrocarbon receptor (AhR) is one of the most important environmental-sensor transcription factors and mediates the metabolism of a wide variety of xenobiotics. Nevertheless, the identification of dietary and endogenous ligands of AhR suggest that it may also fulfil physiological functions with which pollutants may interfere. Placental AhR expression and activity is largely unknown. We established the cartography of AhR expression at transcript and protein levels, its cellular distribution, and its transcriptional activity toward the expression of its main target genes. We studied the profile of AhR expression and activity during different pregnancy periods, during trophoblasts differentiation in vitro, and in a trophoblast cell line. Using diverse methods, such as cell fractionation and immunofluorescence microscopy, we found a constitutive nuclear localization of AhR in every placental model, in the absence of any voluntarily-added exogenous activator. Our data suggest an intrinsic activation of AhR due to the presence of endogenous placental ligands.

MicroRNA-150 down Regulation in Acute Myeloid Leukaemia Patients and Its Prognostic Implication.

BACKGROUND:MicroRNAs (miRNAs) are small, non-coding RNAs that are important for post-transcriptional gene regulation in both healthy and morbid conditions. Numerous miRNAs promote tumorigenesis, while others have a tumour suppressive effects. Acute myeloid leukaemia (AML) is a heterogeneous group of genetically diverse hematopoietic malignancies with variable response to treatment.AIM:Our study aimed to investigate the possible role of miR-150 in de novo adult AML and the impact of its level on survival, and we used in the silicon analysis to predict the main target genes involved in miR-150 mediated cancer pathway.MATERIAL AND METHODS:We evaluated miR-150 expression profiling assay using TaqMan primer probes RT-PCR in the plasma of 50 adult AML patients, before the start of treatment and at day 28 of treatment, along with 20 normal adult control samples. miR-16 was used as an endogenous reference for standardisation. Follow-up of patients during treatment at day 28 of induction chemotherapy and after one year was done.RESULTS:In this study, we found a significantly lower level of miR-150 in AML patients when compared to controls (p = 0.005) with 0.62 fold change than in healthy controls. Patients were divided into two groups: the low miR-150 group (miR-150 < 1) and the high miR-150 group (miR-150 > 1). A statistically significant difference was found between the two groups regarding initial total leukocytic count and initial PB blast count while for the TLC, HB and PLT count at follow up. No difference in the overall survival between the low and the high miR-150 groups could be demonstrated.CONCLUSION:Our results suggest that miR-150 functions as a tumour suppressor and gatekeeper in inhibiting cell transformation and that its downregulation is required for leukemogenesis.

Key elements involved in Epstein\u2013Barr virus-associated gastric cancer and their network regulation

BackgroundThe molecular mechanism of Epstein–Barr virus (EBV)-associated gastric cancer (EBVaGC) remains elusive. A collection of molecular regulators including transcription factor and noncoding RNA (ncRNAs) may affect the carcinogenesis of EBVaGC by regulating the expression and function of key genes. In this study, integration of multi-level expression data and bioinformatics approach was used to identify key elements and their interactions involved in mechanism of EBVaGC and their network regulation.MethodsData of the gene expression profiling data sets (GSE51575) was downloaded from GEO database. Differentially expressed genes between EBVaGC and normal samples were identified by GEO2R. Gene ontology and pathway enrichment analyses were performed using R packages Cluster profiler. STRING database was used to find interacting proteins between different genes. Transcription factors in differentially expressed genes were obtained from TF Checkpoint database. Using Cytoscape, we built transcription factor regulation network. miRNAs involved in the gene-interacting proteins and the miRNA-targeted lncRNA were predicted through miRWalk. Using ViRBase, EBV related miRNA regulation network was built. Overlapping genes and regulators of the above three networks were further identified, and the cross network was constructed using Cytoscape software. Moreover, the differential expressions of the target genes and transcription factors in the cross network were explored in different molecular subtypes of GC using cBioPortal. By histological verification, the expression of two main target genes in the cross network were further analyzed.ResultsA total of 104 genes showed differential expressions between EBVaGC and normal tissues, which were associated with digestion, G-protein coupled receptor binding, gastric acid secretion, etc. Pathway analysis showed that the differentially expressed genes were mainly enriched in gastric acid secretion and protein digestion and absorption. Using STRING dataset, a total of 54 proteins interacted with each other. Based on the transcription factor network, the hub transcription factors IRX3, NKX6-2, PTGER3 and SMAD5 were identified to regulate their target genes SST and GDF5, etc. After screening and matching in miRwalk datasets, a ceRNA network was established, in which the top five miRNAs were hsa-miR-4446-3p, hsa-miR-5787, hsa-miR-1915-3p, hsa-miR-335-3p and hsa-miR-6877-3p, and the top two lncRNAs were RP5-1039K5.19 and TP73-AS1. According to the EBV related miRNA regulation network, CXCL10 and SMAD5 were found to be regulated by EBV-miR-BART1-3p and EBV-mir-BART22, respectively. By overlapping the three networks, CXCL10, GDF5, PTGER3, SMAD5, miR-6877-3p, RP5-1039K5.19, TP73-AS1, EBV-miR-BART1-3p and EBV-mir-BART22 were found to be key elements of regulation mechanism of EBVaGC. CXCL10, GDF5, PTGER3 and SMAD5 were also differentially expressed among the four molecular subtypes of GC. The histological verification experiment showed differential expressions of the two main target genes GDF5 and CXCL10 between EBVaGC and non-tumor tissues as well as EBVnGC.ConclusionIn the current study, our results revealed key elements and their interactions involved in EBVaGC. Some hub transcription factors, miRNAs, lncRNAs and EBV related miRNAs were observed to regulate their target genes. Overlapping genes and regulators were observed in diverse regulation networks, such as CXCL10, GDF5, PTGER3, SMAD5, miR-6877-3p, RP5-1039K5.19, TP73-AS1, EBV-miR-BART1-3p and EBV-mir-BART22. Moreover, CXCL10, GDF5, PTGER3 and SMAD5 were also differentially expressed among the four molecular subtypes of GC. The histological verification experiment showed differential expressions of the two main target genes GDF5 and CXCL10 between EBVaGC and non-tumor tissues as well as EBVnGC. Therefore, the identified key elements and their network regulation may be specifically involved in EBVaGC mechanisms.

Inhibition of HIF1α-Dependent Upregulation of Phospho-l-Plastin Resensitizes Multiple Myeloma Cells to Frontline Therapy.

The introduction of novel frontline agents in multiple myeloma (MM), like immunomodulatory drugs and proteasome inhibitors, has improved the overall survival of patients. Yet, MM is still not curable, and drug resistance (DR) remains the main challenge. To improve the understanding of DR in MM, we established a resistant cell line (MOLP8/R). The exploration of DR mechanisms yielded an overexpression of HIF1α, due to impaired proteasome activity of MOLP8/R. We show that MOLP8/R, like other tumor cells, overexpressing HIF1α, have an increased resistance to the immune system. By exploring the main target genes regulated by HIF1α, we could not show an overexpression of these targets in MOLP8/R. We, however, show that MOLP8/R cells display a very high overexpression of LCP1 gene (l-Plastin) controlled by HIF1α, and that this overexpression also exists in MM patient samples. The l-Plastin activity is controlled by its phosphorylation in Ser5. We further show that the inhibition of l-Plastin phosphorylation restores the sensitivity of MOLP8/R to immunomodulatory drugs (IMiDs) and proteasome inhibitors (PIs). Our results reveal a new target gene of DR, controlled by HIF1α.

Multivalency regulates activity in an intrinsically disordered transcription factor.

The transcription factor ASCIZ (ATMIN, ZNF822) has an unusually high number of recognition motifs for the product of its main target gene, the hub protein LC8 (DYNLL1). Using a combination of biophysical methods, structural analysis by NMR and electron microscopy, and cellular transcription assays, we developed a model that proposes a concerted role of intrinsic disorder and multiple LC8 binding events in regulating LC8 transcription. We demonstrate that the long intrinsically disordered C-terminal domain of ASCIZ binds LC8 to form a dynamic ensemble of complexes with a gradient of transcriptional activity that is inversely proportional to LC8 occupancy. The preference for low occupancy complexes at saturating LC8 concentrations with both human and Drosophila ASCIZ indicates that negative cooperativity is an important feature of ASCIZ-LC8 interactions. The prevalence of intrinsic disorder and multivalency among transcription factors suggests that formation of heterogeneous, dynamic complexes is a widespread mechanism for tuning transcriptional regulation.

Statins regulates inflammatory macrophage phenotype through the activation of AhR

Abstract Statins are 3-hydroxy-3-methylglutaryl coenzyme-A (HMG-CoA) reductase competitive inhibitors, that lower blood cholesterol, but also exert potent anti-inflammatory effects. The xenobiotic receptor, aryl hydrocarbon receptor (AhR), plays crucial role in the control of inflammatory responses. Evidences suggest that selected statins promote the transcription of the main AhR target gene, cytochrome P450 family 1 (Cyp1a1). Macrophages are essential components of innate and adaptive immunity, characterized by AhR expression and activity. Based on these evidences, we investigated potential AhR-mediated immune regulatory properties of statins in Bone Marrow Derived Macrophages (BMDM). We found that BMDM activation with the pro-inflammatory cytokine TNFa induced the inflammatory marker, inducible nitric oxide synthase (iNOS), which was completely prevented in BMDMs treated with TNFa in combination with specific statins, namely, atorvastatin and rosuvastatin. We confirmed high AhR expression in BMDM that was increased upon treatment with TNFa. We investigated the ability of different statins to activate AhR using a luciferase reporter assay cell lines and in BMDCs. We found that selected statins, such as atorvastatin and rosuvastatin, activated AhR in a dose dependent manner, an activation prevented in cells expressing selected AhR mutants. Notably, AhR deficient BMDMs, expressed undetectable levels of the main statin target protein HMG-CoA, which was instead highly expressed in wild type BMDMs. These data demonstrate that natural or synthetic statins can play regulatory roles in macrophages. Moreover, our results suggest a potential involvement of AhR in statin-mediated anti-inflammatory effects in macrophages.

The role of the Hedgehog signaling pathway in cancer: A comprehensive review.

The Hedgehog (Hh) signaling pathway was first identified in the common fruit fly. It is a highly conserved evolutionary pathway of signal transmission from the cell membrane to the nucleus. The Hh signaling pathway plays an important role in the embryonic development. It exerts its biological effects through a signaling cascade that culminates in a change of balance between activator and repressor forms of glioma-associated oncogene (Gli) transcription factors. The components of the Hh signaling pathway involved in the signaling transfer to the Gli transcription factors include Hedgehog ligands (Sonic Hh [SHh], Indian Hh [IHh], and Desert Hh [DHh]), Patched receptor (Ptch1, Ptch2), Smoothened receptor (Smo), Suppressor of fused homolog (Sufu), kinesin protein Kif7, protein kinase A (PKA), and cyclic adenosine monophosphate (cAMP). The activator form of Gli travels to the nucleus and stimulates the transcription of the target genes by binding to their promoters. The main target genes of the Hh signaling pathway are PTCH1, PTCH2, and GLI1. Deregulation of the Hh signaling pathway is associated with developmental anomalies and cancer, including Gorlin syndrome, and sporadic cancers, such as basal cell carcinoma, medulloblastoma, pancreatic, breast, colon, ovarian, and small-cell lung carcinomas. The aberrant activation of the Hh signaling pathway is caused by mutations in the related genes (ligand-independent signaling) or by the excessive expression of the Hh signaling molecules (ligand-dependent signaling - autocrine or paracrine). Several Hh signaling pathway inhibitors, such as vismodegib and sonidegib, have been developed for cancer treatment. These drugs are regarded as promising cancer therapies, especially for patients with refractory/advanced cancers.

The FXR Agonist, Obeticholic Acid, Suppresses HCC Proliferation & Metastasis: Role of IL-6/STAT3 Signalling Pathway

The nuclear receptor, farnesoid X receptor (FXR), has been recently considered as a tumor suppressor in HCC. IL-6/Janus kinase 2 (Jak-2)/signal transducer and activator of transcription 3 (STAT3) pathway has been implicated as a key player in many cancer types. This study aimed at investigating the potential effect of the FXR agonist, obeticholic acid (OCA), on HCC and the involvement of IL-6/STAT3 pathway. The potential regulation of STAT3 by its main feedback inhibitor target gene, the suppressor of cytokine signaling 3 (SOCS3), triggered by OCA was also explored. Cytotoxicity studies were performed on HepG2, Huh7, and SNU-449 cell lines using OCA alone and combined with the FXR antagonist guggulsterone (Gugg). OCA cytotoxic effect was significantly hampered in presence of Gugg. OCA also caused cell cycle arrest and inhibited invasion and migration of HCC cells. Decrease in STAT3 phosphorylation and SOCS3 upregulation were also observed. Moreover, Jak-2, IL-1β, and IL-6 levels were decreased. These results were correlated with an upregulation of FXR and small heterodimer partner (SHP) levels. Effects of OCA on IL-6/STAT3 main key players were reversed in presence of Gugg. Overall, these findings suggest a potential effect of OCA in HCC via interfering with IL-6/STAT3 signalling pathway in vitro.

Downregulation of the Sonic Hedgehog/Gli pathway transcriptional target Neogenin-1 is associated with basal cell carcinoma aggressiveness.

Basal Cell Carcinoma (BCC) is one of the most diagnosed cancers worldwide. It develops due to an unrestrained Sonic Hedgehog (SHH) signaling activity in basal cells of the skin. Certain subtypes of BCC are more aggressive than others, although the molecular basis of this phenomenon remains unknown. We have previously reported that Neogenin-1 (NEO1) is a downstream target gene of the SHH/GLI pathway in neural tissue. Given that SHH participates in epidermal homeostasis, here we analyzed the epidermal expression of NEO1 in order to identify whether it plays a role in adult epidermis or BCC. We describe the mRNA and protein expression profile of NEO1 and its ligands (Netrin-1 and RGMA) in human and mouse control epidermis and in a broad range of human BCCs. We identify in human BCC a significant positive correlation in the levels of NEO1 receptor, NTN-1 and RGMA ligands with respect to GLI1, the main target gene of the canonical SHH pathway. Moreover, we show via cyclopamine inhibition of the SHH/GLI pathway of ex vivo cultures that NEO1 likely functions as a downstream target of SHH/GLI signaling in the skin. We also show how Neo1 expression decreases throughout BCC progression in the K14-Cre:Ptch1lox/lox mouse model and that aggressive subtypes of human BCC exhibit lower levels of NEO1 than non-aggressive BCC samples. Taken together, these data suggest that NEO1 is a SHH/GLI target in epidermis. We propose that NEO1 may be important in tumor onset and is then down-regulated in advanced BCC or aggressive subtypes.

Lactate stimulates CA IX expression in normoxic cancer cells.

Besides hypoxia, other factors and molecules such as lactate, succinate, and reactive oxygen species activate transcription factor hypoxia-inducible factor-1 (HIF-1) even in normoxia. One of the main target gene products of HIF-1 is carbonic anhydrase IX (CA IX). CA IX is overexpressed in many tumors and serves as prognostic factor for hypoxic, aggressive and malignant cancers. CA IX is also induced in normoxia in high cell density. In this study, we observed that lactate induces CA IX expression in normoxic cancer cells in vitro and in vivo. We further evidenced that participation of both HIF-1 and specificity protein 1 (SP1) transcription factors is crucial for lactate-driven normoxic induction of the CA9 gene. By inducing CA IX, lactate can facilitate the maintenance of cancer cell aggressive behavior in normoxia.