Sortilin-induced lipid accumulation and atherogenesis are suppressed by HNF1b SUMOylation promoted by flavone of Polygonatum odoratum. / çŽ‰ç«¹é»„é ®é€šè¿‡ä¿ƒè¿›HNF1b蛋白的SUMO化修饰抑制sortilinä»‹å¯¼çš„è„‚è´¨ç§¯ç´¯åŠåŠ¨è„‰ç²¥æ ·ç¡¬åŒ–.

This study aims to investigate the impact of hepatocyte nuclear factor 1ß (HNF1b) on macrophage sortilin-mediated lipid metabolism and aortic atherosclerosis and explore the role of the flavone of Polygonatum odoratum (PAOA-flavone)-promoted small ubiquitin-related modifier (SUMO) modification in the atheroprotective efficacy of HNF1b. HNF1b was predicted to be a transcriptional regulator of sortilin expression via bioinformatics, dual-luciferase reporter gene assay, and chromatin immunoprecipitation. HNF1b overexpression decreased sortilin expression and cellular lipid contents in THP-1 macrophages, leading to a depression in atherosclerotic plaque formation in low-density lipoprotein (LDL) receptor-deficient (LDLR-/-) mice. Multiple SUMO1-modified sites were identified on the HNF1b protein and co-immunoprecipitation confirmed its SUMO1 modification. The SUMOylation of HNF1b protein enhanced the HNF1b-inhibited effect on sortilin expression and reduced lipid contents in macrophages. PAOA-flavone treatment promoted SUMO-activating enzyme subunit 1 (SAE1) expression and SAE1-catalyzed SUMOylation of the HNF1b protein, which prevented sortilin-mediated lipid accumulation in macrophages and the formation of atherosclerotic plaques in apolipoprotein E-deficient (ApoE-/-) mice. Interference with SAE1 abrogated the improvement in lipid metabolism in macrophage cells and atheroprotective efficacy in vivo upon PAOA-flavone administration. In summary, HNF1b transcriptionally suppressed sortilin expression and macrophage lipid accumulation to inhibit aortic lipid deposition and the development of atherosclerosis. This anti-atherosclerotic effect was enhanced by PAOA-flavone-facilitated, SAE1-catalyzed SUMOylation of the HNF1b protein.

HNF1B-driven three-dimensional chromatin structure for molecular classification in pancreatic cancers.

The molecular subtypes of pancreatic cancer (PC), either classical/progenitor-like or basal/squamous-like, are currently a major topic of research because of their direct association with clinical outcomes. Some transcription factors (TFs) have been reported to be associated with these subtypes. However, the mechanisms by which these molecular signatures of PCs are established remain unknown. Epigenetic regulatory processes, supported by dynamic changes in the chromatin structure, are essential for transcriptional profiles. Previously, we reported the importance of open chromatin profiles in the biological features and transcriptional status of PCs. Here, we aimed to analyze the relationships between three-dimensional (3D) genome structures and the molecular subtypes of human PCs using Hi-C analysis. We observed a correlation of the specific elements of 3D genome modules, including compartments, topologically associating domains, and enhancer-promoter loops, with the expression of related genes. We focused on HNF1B, a TF that is implicated in the progenitor subtype. Forced expression of HNF1B in squamous-type PC organoids induced the upregulation and downregulation of genes associated with progenitor and squamous subtypes, respectively. Long-range genomic interactions induced by HNF1B were accompanied by compartment modulation and H3K27ac redistribution. We also found that these HNF1B-induced changes in subtype-related gene expression required an intrinsically disordered region, suggesting a possible involvement of phase separation in compartment modulation. Thus, mapping of 3D structural changes induced by TFs, such as HNF1B, may become a useful resource for further understanding the molecular features of PCs.

Dysregulation of HNF1B/Clusterin axis enhances disease progression in a highly aggressive subset of pancreatic cancer patients.

Pancreatic ductal adenocarcinoma (PDAC) is a lethal malignancy and is largely refractory to available treatments. Identifying key pathways associated with disease aggressiveness and therapeutic resistance may characterize candidate targets to improve patient outcomes. We used a strategy of examining the tumors from a subset of PDAC patient cohorts with the worst survival to understand the underlying mechanisms of aggressive disease progression and to identify candidate molecular targets with potential therapeutic significance. Non-negative matrix factorization (NMF) clustering, using gene expression profile, revealed three patient subsets. A 142-gene signature specific to the subset with the worst patient survival, predicted prognosis and stratified patients with significantly different survival in the test and validation cohorts. Gene-network and pathway analysis of the 142-gene signature revealed dysregulation of Clusterin (CLU) in the most aggressive patient subset in our patient cohort. Hepatocyte nuclear factor 1 b (HNF1B) positively regulated CLU, and a lower expression of HNF1B and CLU was associated with poor patient survival. Mechanistic and functional analyses revealed that CLU inhibits proliferation, 3D spheroid growth, invasiveness and epithelial-to-mesenchymal transition (EMT) in pancreatic cancer cell lines. Mechanistically, CLU enhanced proteasomal degradation of EMT-regulator, ZEB1. In addition, orthotopic transplant of CLU-expressing pancreatic cancer cells reduced tumor growth in mice. Furthermore, CLU enhanced sensitivity of pancreatic cancer cells representing aggressive patient subset, to the chemotherapeutic drug gemcitabine. Taken together, HNF1B/CLU axis negatively regulates pancreatic cancer progression and may potentially be useful in designing novel strategies to attenuate disease progression in PDAC patients.

Multiomics analysis reveals that hepatocyte nuclear factor 1ß regulates axon guidance genes in the developing mouse kidney.

The transcription factor hepatocyte nuclear factor 1ß (HNF-1ß) is essential for normal development of the kidney and other epithelial organs. In the developing mouse kidney, HNF-1ß is required for the differentiation and patterning of immature nephrons and branching morphogenesis of the ureteric bud (UB). Here, we used ChIP-sequencing (ChIP-seq) and RNA sequencing (RNA-seq) to identify genes that are regulated by HNF-1ß in embryonic mouse kidneys. ChIP-seq revealed that HNF-1ß binds to 8284 sites in chromatin from E14.5 mouse kidneys. Comparison with previous ATAC-seq and histone modification studies showed that HNF-1ß binding peaks colocalized with open chromatin and epigenetic marks of transcriptional activation (H3K27 acetylation, H3K4 trimethylation, H3K4 monomethylation), indicating that the binding sites were functional. To investigate the relationship between HNF-1ß binding and HNF-1ß-dependent gene regulation, RNA-seq was performed on UB cells purified from wild-type and HNF-1ß mutant embryonic kidneys. A total of 1632 genes showed reduced expression in HNF-1ß-deficient UB cells, and 485 genes contained nearby HNF-1ß binding sites indicating that they were directly activated by HNF-1ß. Conversely, HNF-1ß directly repressed the expression of 526 genes in the UB. Comparison with snATAC-seq analysis of UB-derived cells showed that both HNF-1ß-dependent activation and repression correlated with chromatin accessibility. Pathway analysis revealed that HNF-1ß binds near 68 axon guidance genes in the developing kidney. RNA-seq analysis showed that Nrp1, Sema3c, Sema3d, Sema6a, and Slit2 were activated by HNF-1ß, whereas Efna1, Epha3, Epha4, Epha7, Ntn4, Plxna2, Sema3a, Sema4b, Slit3, Srgap1, Unc5c and Unc5d were repressed by HNF-1ß. RNAscope in situ hybridization showed that Nrp1, Sema3c, Sema3d, Sema6a, and Slit2 were expressed in wild-type UB and were dysregulated in HNF-1ß mutant UB. These studies show that HNF-1ß directly regulates the expression of multiple axon guidance genes in the developing mouse kidney. Dysregulation of axon guidance genes may underlie kidney defects in HNF-1ß mutant mice.

The mechanism and effects of remdesivir-induced developmental toxicity in zebrafish: Blood flow dysfunction and behavioral alterations.

The antiviral drug remdesivir has been used to treat the growing number of coronavirus disease 2019 (COVID-19) patients. However, the drug is mainly excreted through urine and feces and introduced into the environment to affect non-target organisms, including fish, which has raised concerns about potential ecotoxicological effects on aquatic organisms. Moreover, studies on the ecological impacts of remdesivir on aquatic environments have not been reported. Here, we aimed to explore the toxicological impacts of microinjection of remdesivir on zebrafish early embryonic development and larvae and the associated mechanism. We found that 100 µM remdesivir delayed epiboly and impaired convergent movement of embryos during gastrulation, and dose-dependent increases in mortality and malformation were observed in remdesivir-treated embryos. Moreover, 10-100 µM remdesivir decreased blood flow and swimming velocity and altered the behavior of larvae. In terms of molecular mechanisms, 80 differentially expressed genes (DEGs) were identified by transcriptome analysis in the remdesivir-treated group. Some of these DEGs, such as manf, kif3a, hnf1ba, rgn, prkcz, egr1, fosab, nr4a1, and ptgs2b, were mainly involved in early embryonic development, neuronal developmental disorders, vascular disease and the blood flow pathway. These data reveal that remdesivir can impair early embryonic development, blood flow and behavior of zebrafish embryos/larvae, probably due to alterations at the transcriptome level. This study suggests that it is important to avoid the discharge of remdesivir to aquatic ecosystems and provides a theoretical foundation to hinder remdesivir-induced ecotoxicity to aquatic environments.

Mechanisms of ion transport regulation by HNF1ß in the kidney: beyond transcriptional regulation of channels and transporters.

Hepatocyte nuclear factor 1ß (HNF1ß) is a transcription factor essential for the development and function of the kidney. Mutations in and deletions of HNF1ß cause autosomal dominant tubule interstitial kidney disease (ADTKD) subtype HNF1ß, which is characterized by renal cysts, diabetes, genital tract malformations, and neurodevelopmental disorders. Electrolyte disturbances including hypomagnesemia, hyperuricemia, and hypocalciuria are common in patients with ADTKD-HNF1ß. Traditionally, these electrolyte disturbances have been attributed to HNF1ß-mediated transcriptional regulation of gene networks involved in ion transport in the distal part of the nephron including FXYD2, CASR, KCNJ16, and FXR. In this review, we propose additional mechanisms that may contribute to the electrolyte disturbances observed in ADTKD-HNF1ß patients. Firstly, kidney development is severely affected in Hnf1b-deficient mice. HNF1ß is required for nephron segmentation, and the absence of the transcription factor results in rudimentary nephrons lacking mature proximal tubule, loop of Henle, and distal convoluted tubule cluster. In addition, HNF1ß is proposed to be important for apical-basolateral polarity and tight junction integrity in the kidney. Interestingly, cilia formation is unaffected by Hnf1b defects in several models, despite the HNF1ß-mediated transcriptional regulation of many ciliary genes. To what extent impaired nephron segmentation, apical-basolateral polarity, and cilia function contribute to electrolyte disturbances in HNF1ß patients remains elusive. Systematic phenotyping of Hnf1b mouse models and the development of patient-specific kidney organoid models will be essential to advance future HNF1ß research.

LncRNA4474 inhibits renal fibrosis by regulating hepatocyte nuclear factor-1ß through miR-615 modulation.

Long noncoding RNAs (lncRNAs) and microRNAs (miRNAs) are involved in the development and progression of renal fibrosis. lncRNAs can regulate target messenger RNAs (mRNAs) by competitively binding to miRNAs. However, research on lncRNA-miRNA-mRNA interactions remains inadequate. Therefore, the aim of the present study was to investigate the possible function of lncRNA-miRNA-mRNA interactions in chronic renal fibrosis. The relationships among the expression levels of lncRNA4474, miR-615, and hepatocyte nuclear factor-1ß (HNF-1ß) mRNAs were determined through RNA sequencing. The biological roles of lncRNA4474, miR-615, and HNF-1ß in renal fibrosis were investigated with gain-of-function and loss-of-function experiments. Results showed that miR-615 expression increased in unilateral ureteral obstruction rats, accompanied by decreased lncRNA4474 and HNF-1ß mRNA expression. The overexpression of HNF-1ß attenuated the development of chronic renal fibrosis, whereas HNF-1ß knockdown promoted the development. Increase in HNF-1ß expression downregulated and upregulated the expression levels of miR-615 and lncRNA4474, respectively, thereby attenuating renal fibrosis progression. Furthermore, lncRNA4474 promoted the expression of HNF-1ß by inhibiting miR-615 expression, whereas miR-615 regulated the expression of HNF-1ß and thus activated the Wnt signaling pathway. This study demonstrated that the overexpression of lncRNA4474 may attenuate fibrosis progression, accompanied by the downregulation of miR-615 and upregulation of HNF-1ß. Hence, this study provides novel information that can be useful in the early diagnosis and treatment of renal fibrosis.

Comprehensive quantitative analysis of alternative splicing variants reveals the HNF1B mRNA splicing pattern in various tumour and non-tumour tissues.

Hepatocyte nuclear factor-1-beta (HNF1B) is a transcription factor and putative biomarker of solid tumours. Recently, we have revealed a variety of HNF1B mRNA alternative splicing variants (ASVs) with unknown, but potentially regulatory, functions. The aim of our work was to quantify the most common variants and compare their expression in tumour and non-tumour tissues of the large intestine, prostate, and kidney. The HNF1B mRNA variants 3p, Δ7, Δ7-8, and Δ8 were expressed across all the analysed tissues in 28.2-33.5%, 1.5-2%, 0.8-1.7%, and 2.3-6.9% of overall HNF1B mRNA expression, respectively, and occurred individually or in combination. The quantitative changes of ASVs between tumour and non-tumour tissue were observed for the large intestine (3p, Δ7-8), prostate (3p), and kidney samples (Δ7). Decreased expression of the overall HNF1B mRNA in the large intestine and prostate cancer samples compared with the corresponding non-tumour samples was observed (p = 0.019 and p = 0.047, respectively). The decreased mRNA expression correlated with decreased protein expression in large intestine carcinomas (p < 0.001). The qualitative and quantitative pattern of the ASVs studied by droplet digital PCR was confirmed by next-generation sequencing, which suggests the significance of the NGS approach for further massive evaluation of the splicing patterns in a variety of genes.

Identification of downstream effectors of retinoic acid specifying the zebrafish pancreas by integrative genomics.

Retinoic acid (RA) is a key signal for the specification of the pancreas. Still, the gene regulatory cascade triggered by RA in the endoderm remains poorly characterized. In this study, we investigated this regulatory network in zebrafish by combining RNA-seq, RAR ChIP-seq and ATAC-seq assays. By analysing the effect of RA and of the RA receptor (RAR) inverse-agonist BMS493 on the transcriptome and on the chromatin accessibility of endodermal cells, we identified a large set of genes and regulatory regions regulated by RA signalling. RAR ChIP-seq further defined the direct RAR target genes in zebrafish, including hox genes as well as several pancreatic regulators like mnx1, insm1b, hnf1ba and gata6. Comparison of zebrafish and murine RAR ChIP-seq data highlighted the conserved direct target genes and revealed that some RAR sites are under strong evolutionary constraints. Among them, a novel highly conserved RAR-induced enhancer was identified downstream of the HoxB locus and driving expression in the nervous system and in the gut in a RA-dependent manner. Finally, ATAC-seq data unveiled the role of the RAR-direct targets Hnf1ba and Gata6 in opening chromatin at many regulatory loci upon RA treatment.

Hepatocyte nuclear factor-1ß shapes the energetic homeostasis of kidney tubule cells.

Energetic metabolism controls key steps of kidney development, homeostasis, and epithelial repair following acute kidney injury (AKI). Hepatocyte nuclear factor-1ß (HNF-1ß) is a master transcription factor that controls mitochondrial function in proximal tubule (PT) cells. Patients with HNF1B pathogenic variant display a wide range of kidney developmental abnormalities and progressive kidney fibrosis. Characterizing the metabolic changes in PT cells with HNF-1ß deficiency may help to identify new targetable molecular hubs involved in HNF1B-related kidney phenotypes and AKI. Here, we combined 1 H-NMR-based metabolomic analysis in a murine PT cell line with CrispR/Cas9-induced Hnf1b invalidation (Hnf1b-/- ), clustering analysis, targeted metabolic assays, and datamining of published RNA-seq and ChIP-seq dataset to identify the role of HNF-1ß in metabolism. Hnf1b-/- cells grown in normoxic conditions display intracellular ATP depletion, increased cytosolic lactate concentration, increased lipid droplet content, failure to use pyruvate for energetic purposes, increased levels of tricarboxylic acid (TCA) cycle intermediates and oxidized glutathione, and a reduction of TCA cycle byproducts, all features consistent with mitochondrial dysfunction and an irreversible switch toward glycolysis. Unsupervised clustering analysis showed that Hnf1b-/- cells mimic a hypoxic signature and that they cannot furthermore increase glycolysis-dependent energetic supply during hypoxic challenge. Metabolome analysis also showed alteration of phospholipid biosynthesis in Hnf1b-/- cells leading to the identification of Chka, the gene coding for choline kinase α, as a new putative target of HNF-1ß. HNF-1ß shapes the energetic metabolism of PT cells and HNF1B deficiency in patients could lead to a hypoxia-like metabolic state precluding further adaptation to ATP depletion following AKI.

Modeling HNF1B-associated monogenic diabetes using human iPSCs reveals an early stage impairment of the pancreatic developmental program.

Heterozygous mutations in HNF1B in humans result in a multisystem disorder, including pancreatic hypoplasia and diabetes mellitus. Here we used a well-controlled human induced pluripotent stem cell pancreatic differentiation model to elucidate the molecular mechanisms underlying HNF1B-associated diabetes. Our results show that lack of HNF1B blocks specification of pancreatic fate from the foregut progenitor (FP) stage, but HNF1B haploinsufficiency allows differentiation of multipotent pancreatic progenitor cells (MPCs) and insulin-secreting ß-like cells. We show that HNF1B haploinsufficiency impairs cell proliferation in FPs and MPCs. This could be attributed to impaired induction of key pancreatic developmental genes, including SOX11, ROBO2, and additional TEAD1 target genes whose function is associated with MPC self-renewal. In this work we uncover an exhaustive list of potential HNF1B gene targets during human pancreas organogenesis whose downregulation might underlie HNF1B-associated diabetes onset in humans, thus providing an important resource to understand the pathogenesis of this disease.

MafA, NeuroD1, and HNF1ß synergistically activate the Slc2a2 (Glut2) gene in ß-cells.

Glucose transporter type 2 (GLUT2), encoded by the SLC2A2 gene, is an essential component of glucose-stimulated insulin secretion in pancreatic islet ß-cells. Like that of the gene encoding insulin, expression of the SLC2A2 gene expression is closely linked to ß-cell functionality in rodents, but the mechanism by which ß-cell-specific expression of SLC2A2 is controlled remains unclear. In this report, to identify putative enhancer elements of the mouse Slc2a2 gene, we examined evolutional conservation of the nucleotide sequence of its genomic locus, together with ChIP-seq data of histone modifications and various transcription factors published in previous studies. Using luciferase reporter assays, we found that an evolutionarily conserved region (ECR) located approximately 40 kbp downstream of the transcription start site of Slc2a2 functions as an active enhancer in the MIN6 ß-cell line. We also found that three ß-cell-enriched transcription factors, MafA, NeuroD1, and HNF1ß, synergistically activate transcription through this 3' downstream distal enhancer (ECR3') and the proximal promoter region of the gene. Our data also indicate that the simultaneous binding of HNF1ß to its target sites within the promoter and ECR3' of Slc2a2 is indispensable for transcriptional activation, and that binding of MafA and NeuroD1 to their respective target sites within the ECR3' enhances transcription. Co-immunoprecipitation experiments suggested that MafA, NeuroD1, and HNF1ß interact with each other. Overall, these results suggest that promoter-enhancer communication through MafA, NeuroD1, and HNF1ß is critical for Slc2a2 gene expression. These findings provide clues to help elucidate the mechanism of regulation of Slc2a2 gene expression in ß-cells.

A Review of Functional Characterization of Single Amino Acid Change Mutations in HNF Transcription Factors in MODY Pathogenesis.

Mutations in HNF transcription factor genes cause the most common subtypes of maturity-onset of diabetes of youth (MODY), a monogenic form of diabetes mellitus. Mutations in the HNF1-α, HNF4-α, and HNF1-ß genes are primarily considered as the cause of MODY3, MODY1, and MODY5 subtypes, respectively. Although patients with different subtypes display similar symptoms, they may develop distinct diabetes-related complications and require different treatments depending on the type of the mutation. Genetic analysis of MODY patients revealed more than 400 missense/nonsense mutations in HNF1-α, HNF4-α, and HNF1-ß genes, however only a small portion of them are functionally characterized. Evaluation of nonsense mutations are more direct as they lead to premature stop codons and mostly in mRNA decay or nonfunctional truncated proteins. However, interpretation of the single amino acid change (missense) mutation is not such definite, as effect of the variant may vary depending on the location and also the substituted amino acid. Mutations with benign effect on the protein function may not be the pathologic variant and further genetic testing may be required. Here, we discuss the functional characterization analysis of single amino acid change mutations identified in HNF1-α, HNF4-α, and HNF1-ß genes and evaluate their roles in MODY pathogenesis. This review will contribute to comprehend HNF nuclear family-related molecular mechanisms and to develop more accurate diagnosis and treatment based on correct evaluation of pathologic effects of the variants.

Bifunctional protein PCBD2 operates as a co-factor for hepatocyte nuclear factor 1ß and modulates gene transcription.

Hepatocyte nuclear factor 1ß (HNF1ß) is an essential transcription factor in development of the kidney, liver, and pancreas. HNF1ß-mediated transcription of target genes is dependent on the cell type and the development stage. Nevertheless, the regulation of HNF1ß function by enhancers and co-factors that allow this cell-specific transcription is largely unknown. To map the HNF1ß interactome we performed mass spectrometry in a mouse kidney inner medullary collecting duct cell line. Pterin-4a-carbinolamine dehydratase 2 (PCBD2) was identified as a novel interaction partner of HNF1ß. PCBD2 and its close homolog PCBD1 shuttle between the cytoplasm and nucleus to exert their enzymatic and transcriptional activities. Although both PCBD proteins share high sequence identity (48% and 88% in HNF1 recognition helix), their tissue expression patterns are unique. PCBD1 is most abundant in kidney and liver while PCBD2 is also abundant in lung, spleen, and adipose tissue. Using immunolocalization studies and biochemical analysis we show that in presence of HNF1ß the nuclear localization of PCBD1 and PCBD2 increases significantly. Promoter luciferase assays demonstrate that co-factors PCBD1 and PCBD2 differentially regulate the ability of HNF1ß to activate the promoters of transcriptional targets important in renal electrolyte homeostasis. Deleting the N-terminal sequence of PCBD2, not found in PCBD1, diminished the differential effects of the co-factors on HNF1ß activity. All together these results indicate that PCBD1 and PCBD2 can exert different effects on HNF1ß-mediated transcription. Future studies should confirm whether these unique co-factor activities also apply to HNF1ß-target genes involved in additional processes besides ion transport in the kidney.

Inhibition of hepatocyte nuclear factor 1ß contributes to cisplatin nephrotoxicity via regulation of nf-κb pathway.

Cisplatin nephrotoxicity has been considered as serious side effect caused by cisplatin-based chemotherapy. Recent evidence indicates that renal tubular cell apoptosis and inflammation contribute to the progression of cisplatin-induced acute kidney injury (AKI). Hepatocyte nuclear factor 1ß (HNF1ß) has been reported to regulate the development of kidney cystogenesis, diabetic nephrotoxicity, etc However, the regulatory mechanism of HNF1ß in cisplatin nephrotoxicity is largely unknown. In the present study, we examined the effects of HNF1ß deficiency on the development of cisplatin-induced AKI in vitro and in vivo. HNF1ß down-regulation exacerbated cisplatin-induced RPTC apoptosis by indirectly inducing NF-κB p65 phosphorylation and nuclear translocation. HNF1ß knockdown C57BL/6 mice were constructed by injecting intravenously with HNF1ß-interfering shRNA and PEI. The HNF1ß scramble and knockdown mice were treated with 30 mg/kg cisplatin for 3 days to induce acute kidney injury. Cisplatin treatment caused increased caspase 3 cleavage and p65 phosphorylation, elevated serum urea nitrogen and creatinine, and obvious histological damage of kidney such as fractured tubules in control mice, which were enhanced in HNF1ß knockdown mice. These results suggest that HNF1ß may ameliorate cisplatin nephrotoxicity in vitro and in vivo, probably through regulating NF-κB signalling pathway.

Gestational bisphenol A exposure induces fatty liver development in male offspring mice through the inhibition of HNF1b and upregulation of PPARγ.

Bisphenol A (BPA) is an endocrine-disrupting chemical (EDC) associated with non-alcoholic fatty liver disease (NAFLD). The effects of gestational BPA exposure on hepatic lipid accumulation in offspring are not fully understood. Here, we investigate the sex-dependent effects of gestational BPA exposure on hepatic lipid and glucose metabolism in the offspring of mice to reveal the mechanisms underlying gestational BPA exposure-associated NAFLD. Pregnant mice were administered gavage with or without 1 µg kg-1 day-1 BPA at embryonic day 7.5 (E7.5)-E16.5. Hepatic glucose and lipid metabolism were evaluated in these models. Both male and female offspring mice exhibited hepatic fatty liver after BPA treatment. Lipid accumulation and dysfunction of glucose metabolism were observed in male offspring. We revealed abnormal expression of lipid regulators in the liver and that inhibition of peroxisome proliferator-activated receptor γ (PPARγ) repressed hepatic lipid accumulation induced by gestational BPA exposure. We also found a sex-dependent decrease of hepatocyte nuclear factor 1b (HNF1b) expression in male offspring. The transcriptional repression of PPARγ by HNF1b was confirmed in L02 cells. Downregulation of HNF1b, upregulation of PPARγ, and subsequent upregulation of hepatic lipid accumulation were essential for NAFLD development in male offspring gestationally exposed to BPA as well as BPA-exposed adult male mice. Dysregulation of the HNF1b/PPARγ pathway may be involved in gestational BPA exposure-induced NAFLD in male offspring. These data provide new insights into the mechanism of gestational BPA exposure-associated sex-dependent glucose and lipid metabolic dysfunction. Graphical abstract Schematic of the mechanism of gestational BPA exposure-induced glucose and lipid metabolic dysfunction.

Hypoxia inducible factor-2α importance for migration, proliferation, and self-renewal of trunk neural crest cells.

BACKGROUND: The neural crest is a transient embryonic stem cell population. Hypoxia inducible factor (HIF)-2α is associated with neural crest stem cell appearance and aggressiveness in tumors. However, little is known about its role in normal neural crest development. RESULTS: Here, we show that HIF-2α is expressed in trunk neural crest cells of human, murine, and avian embryos. Knockdown as well as overexpression of HIF-2α in vivo causes developmental delays, induces proliferation, and self-renewal capacity of neural crest cells while decreasing the proportion of neural crest cells that migrate ventrally to sympathoadrenal sites. Reflecting the in vivo phenotype, transcriptome changes after loss of HIF-2α reveal enrichment of genes associated with cancer, invasion, epithelial-to-mesenchymal transition, and growth arrest. CONCLUSIONS: Taken together, these results suggest that expression levels of HIF-2α must be strictly controlled during normal trunk neural crest development and that dysregulated levels affects several important features connected to stemness, migration, and development.

HNF1B inhibits cell proliferation via repression of SMAD6 expression in prostate cancer.

Prostate cancer is the most common malignancy in men in developed countries. In previous study, we identified HNF1B (Hepatocyte Nuclear Factor 1ß) as a downstream effector of Enhancer of zeste homolog 2 (EZH2). HNF1B suppresses EZH2-mediated migration of two prostate cancer cell lines via represses the EMT process by inhibiting SLUG expression. Besides, HNF1B expression inhibits cell proliferation through unknown mechanisms. Here, we demonstrated that HNF1B inhibited the proliferation rate of prostate cancer cells. Overexpression of HNF1B in prostate cancer cells led to the arrest of G1 cell cycle and decreased Cyclin D1 expression. In addition, we re-explored data from ChIP-sequencing (ChIP-seq) and RNA-sequencing (RNA-seq), and demonstrated that HNF1B repressed Cyclin D1 via direct suppression of SMAD6 expression. We also identified CDKN2A as a HNF1B-interacting protein that would contribute to HNF1B-mediated repression of SMAD6 expression. In summary, we provide the novel mechanisms and evidence in support HNF1B as a tumour suppressor gene for prostate cancer.

HNF1B, EZH2 and ECI2 in prostate carcinoma. Molecular, immunohistochemical and clinico-pathological study.

Hepatocyte nuclear factor 1 beta (HNF1B) is a tissue specific transcription factor, which seems to play an important role in the carcinogenesis of several tumors. In our study we focused on analyzing HNF1B in prostate carcinoma (PC) and adenomyomatous hyperplasia (AH), as well as its possible relation to the upstream gene EZH2 and downstream gene ECI2. The results of our study showed that on an immunohistochemical level, the expression of HNF1B was low in PC, did not differ between PC and AH, and did not correlate with any clinical outcomes. In PC, mutations of HNF1B gene were rare, but the methylation of its promotor was a common finding and was positively correlated with Gleason score and stage. The relationship between HNF1B and EZH2/ECI2 was equivocal, but EZH2 and ECI2 were positively correlated on both mRNA and protein level. The expression of EZH2 was associated with poor prognosis. ECI2 did not correlate with any clinical outcomes. Our results support the oncosuppressive role of HNF1B in PC, which may be silenced by promotor methylation and other mechanisms, but not by gene mutation. The high expression of EZH2 (especially) and ECI2 in PC seems to be a potential therapeutic target.

Upregulation of fibronectin following loss of p53 function is a poor prognostic factor in ovarian carcinoma with a unique immunophenotype.

BACKGROUND: We previously demonstrated that ovarian high grade serous carcinomas (OHGSeCa) and ovarian clear cell carcinomas (OCCCa) with an HNF-1ß+/p53+/ARID1A+ immunophenotype were associated with the worst unfavorable prognosis. To clarify the molecular mechanisms underlying this finding, we focused on alterations in the p53 signaling pathway in these tumors. METHODS: Changes in cell phenotype and function following knockdown of wild-type p53 (p53-KD) were assessed using OCCCa cells expressing endogenous HNF-1ß and ARID1A. The prognostic significance of molecules that were deregulated following p53-KD was also examined using 129 OCCCa/OHGSeCa cases. RESULTS: p53-KD cells had increased expression of Snail, phospho-Akt (pAkt), and pGSK3ß, and decreased E-cadherin expression, leading to epithelial-mesenchymal transition (EMT)/cancer stem cell (CSC) features. The cells also exhibited acceleration of cell motility and inhibition of cell proliferation and apoptosis. Next generation sequencing revealed that fibronectin (FN) expression was significantly increased in the p53 KD-cells, in line with our observation that wild-type p53 (but not mutant p53) repressed FN1 promoter activity. In addition, treatment of OCCCa cells with FN significantly increased cell migration capacity and decreased cell proliferation rate, independent of induction of EMT features. In clinical samples, FN/p53 scores were significantly higher in OCCCa/OHGSeCa with the HNF-1ß+/p53+/ARID1A+ immunophenotype when compared to others. Moreover, high FN/high p53 expression was associated with the worst overall survival and progression-free survival in OCCCa/OHGSeCa patients. CONCLUSION: These findings suggest that upregulation of FN following loss of p53 function may impact the biological behavior of OCCCa/OHGSeCa, particularly in tumors with an HNF-1ß+/p53+/ARID1A+ immunophenotype, through alterations in cell mobility and cell proliferation. The accompanying induction of EMT/CSC properties and inhibition of apoptosis due to p53 abnormalities also contribute to the establishment and maintenance of tumor phenotypic characteristics. Video Abstract.