Insulin Transcription Research Articles

Structural impotence of PDX-1 protein in the epigenetic control of insulin expression

This study investigates the role of pancreatic and duodenal homeobox 1 (PDX-1) as an important transcription factor in the epigenetic regulation of insulin expression. The research focused on PDX-1 as a disordered protein, and how the flexibility of its intrinsically disordered regions (IDRs) allows PDX-1 to interact with various transcription factors and epigenetic enzymes, facilitating histone modifications and insulin transcription. Particularly, the role of PDX-1 in processes like acetylation, methylation, and demethylation was discussed by detailing its interactions with key enzymes such as p300, Set7, and JMJD3. For instance, PDX-1’s interaction with p300 leads to histone H4 hyperacetylation, enhancing insulin gene expression, while Set7-mediated methylation of H3K4 further promotes transcription. In addition, the demethylation of H3K27 by JMJD3 is also facilitated by the PDX-1 protein. Finally, structural predictions of PDX-1 using AlphaFold III and EMS Fold also highlight the importance of its IDRs in these interactions. The findings highlight the significance of PDX-1 in insulin regulation and offer insights into potential therapeutic targets for treating diabetes from an epigenetic approach.

Structural insights into i-motif DNA structures in sequences from the insulin-linked polymorphic region

The insulin-linked polymorphic region is a variable number of tandem repeats region of DNA in the promoter of the insulin gene that regulates transcription of insulin. This region is known to form the alternative DNA structures, i-motifs and G-quadruplexes. Individuals have different sequence variants of tandem repeats and although previous work investigated the effects of some variants on G-quadruplex formation, there is not a clear picture of the relationship between the sequence diversity, the DNA structures formed, and the functional effects on insulin gene expression. Here we show that different sequence variants of the insulin linked polymorphic region form different DNA structures in vitro. Additionally, reporter genes in cellulo indicate that insulin expression may change depending on which DNA structures form. We report the crystal structure and dynamics of an intramolecular i-motif, which reveal sequences within the loop regions forming additional stabilising interactions that are critical to formation of stable i-motif structures. The outcomes of this work reveal the detail in formation of stable i-motif DNA structures, with potential for rational based drug design for compounds to target i-motif DNA.

Among Other Tissues, Short-Term Garlic Oral Treatment Incrementally Improves Indicants of Only Pancreatic Islets of Langerhans Histology and Insulin mRNA Transcription and Synthesis in Diabetic Rats.

The source, mRNA transcription, and synthesis of insulin in the pancreas, in addition to the bile duct and liver, in streptozotocin (STZ)-induced diabetic rats (DR) in response to garlic oral treatment are not yet clear. This study investigated the accumulative effects of continued garlic oral treatment on changes in the pancreas, bile duct, and liver with regards to: 1-Insulin mRNA transcription, synthesis, and concentration in relation to changes in serum insulin (SI); 2-Insulinogenic cells insulin intensity and distribution, proliferation, and morphology. Fasting blood glucose (FBG) and insulin concentration in serum and pancreas (PI) and sources and mRNA transcription in the pancreas, bile duct, and liver in normal rats given normal saline (NR-NS) and DR given either NS (DR-NS) or garlic extract (DR-GE) before and after 1, 4, and 8 weeks of oral treatment were examined. Compared to NR-NS, DR-NS showed a significant increase in FBG and reductions in SI and PI and deterioration in islets histology, associated pancreatic insulin numerical intensities, and mRNA transcription. However, compared to DR-NS, the targeted biochemical, histological, and genetic variables of DR-GE were significantly and incrementally improved as garlic treatment continued. Insulin or its indicators were not detected either in the bile duct or the liver in DR-GE. 8 weeks of garlic oral treatment is enough to incrementally restore only pancreatic islets of Langerhans insulin intensity and insulinogenic cells proliferation, morphology, and distribution. These indices were associated with enhanced pancreatic insulin mRNA transcription and synthesis. Eight weeks of garlic treatment were not enough to stimulate insulinogenesis in either the bile duct or the liver.

Downregulation of Glis3 in INS1 cells exposed to chronically elevated glucose contributes to glucotoxicity-associated β cell dysfunction

ABSTRACT Chronically elevated levels of glucose are deleterious to pancreatic β cells and contribute to β cell dysfunction, which is characterized by decreased insulin production and a loss of β cell identity. The Krüppel-like transcription factor, Glis3 has previously been shown to positively regulate insulin transcription and mutations within the Glis3 locus have been associated with the development of several pathologies including type 2 diabetes mellitus. In this report, we show that Glis3 is significantly downregulated at the transcriptional level in INS1 832/13 cells within hours of being subjected to high glucose concentrations and that diminished expression of Glis3 is at least partly attributable to increased oxidative stress. CRISPR/Cas9-mediated knockdown of Glis3 indicated that the transcription factor was required to maintain normal levels of both insulin and MafA expression and reduced Glis3 expression was concomitant with an upregulation of β cell disallowed genes. We provide evidence that Glis3 acts similarly to a pioneer factor at the insulin promoter where it permissively remodels the chromatin to allow access to a transcriptional regulatory complex including Pdx1 and MafA. Finally, evidence is presented that Glis3 can positively regulate MafA transcription through its pancreas-specific promoter and that MafA reciprocally regulates Glis3 expression. Collectively, these results suggest that decreased Glis3 expression in β cells exposed to chronic hyperglycemia may contribute significantly to reduced insulin transcription and a loss of β cell identity.

FXR controls insulin content by regulating Foxa2-mediated insulin transcription

Farnesoid X receptor (FXR) is a nuclear ligand-activated receptor of bile acids that plays a role in the modulation of insulin content. However, the underlying molecular mechanisms remain unclear. Forkhead box a2 (Foxa2) is an important nuclear transcription factor in pancreatic β-cells and is involved in β-cell function. We aimed to explore the signaling mechanism downstream of FXR to regulate insulin content and underscore its association with Foxa2 and insulin gene (Ins) transcription. All experiments were conducted on FXR transgenic mice, INS-1 823/13 cells, and diabetic Goto-Kakizaki (GK) rats undergoing sham or Roux-en-Y gastric bypass (RYGB) surgery. Islets from FXR knockout mice and INS-1823/13 cells with FXR knockdown exhibited substantially lower insulin levels than that of controls. This was accompanied by decreased Foxa2 expression and Ins transcription. Conversely, FXR overexpression increased insulin content, concomitant with enhanced Foxa2 expression and Ins transcription in INS-1 823/13 cells. Moreover, FXR knockdown reduced FXR recruitment and H3K27 trimethylation in the Foxa2 promoter. Importantly, Foxa2 overexpression abrogated the adverse effects of FXR knockdown on Ins transcription and insulin content in INS-1 823/13 cells. Notably, RYGB surgery led to improved insulin content in diabetic GK rats, which was accompanied by upregulated FXR and Foxa2 expression and Ins transcription. Collectively, these data suggest that Foxa2 serves as the target gene of FXR in β-cells and mediates FXR-enhanced Ins transcription. Additionally, the upregulated FXR/Foxa2 signaling cascade could contribute to the enhanced insulin content in diabetic GK rats after RYGB.

Taurine promotes insulin synthesis by enhancing Isl-1 expression through miR-7a/RAF1/ERK1/2 pathway.

It has been well established that the circulating taurine affects the insulin synthesis in pancreatic islet β-cells, whereas miR-7a and LIM-homeodomain transcription factor Isl-1 are important intracellular factors regulating insulin transcription and synthesis. However, it still remains unknown whether taurine regulates insulin synthesis by affecting miR-7a and/or Isl-1 expressions in mouse pancreatic islet β-cells. The present study was thus proposed to identify the effects of taurine on the expressions of miR-7a and/or Isl-1 and their relations to insulin synthesis in mouse pancreatic islet β-cells by using miR-7a2 knockout (KO) and taurine transporter (TauT) KO mouse models and the related in vitro experiments. The results demonstrated that taurine supplement significantly decreased the pancreas miR-7a expression, but sharply upregulated the pancreas Isl-1 and insulin expressions, and serum insulin levels. However, the enhanced effects of taurine on Isl-1 expression and insulin synthesis were mitigated in the TauT KO and miR-7a2 KO mice. In addition, our results confirmed that taurine markedly increased pancreas RAF1 and ERK1/2 expressions. Collectively, the present study firstly demonstrates that taurine regulates insulin synthesis through TauT/miR-7a/RAF1/ERK1/2/Isl-1 signaling pathway, which are crucial for our understanding the mechanisms of taurine affecting insulin synthesis, and also potential for establishing the therapeutic strategies for diabetes and the diseases related to metabolism.

The Impact of Resistance and Aerobic Training on the Insulin Synthesis Genes Expression in Diabetic Rats

Objective: In the last two decades, genetic studies have strongly supported the effective role of genetic factors on the synthesis of insulin from the pancreas. The purpose of this research is to determine the effect of aerobic and resistance exercises on the expression of NeruoD1 and PDX1 in pancreatic tissue, as well as serum insulin and glucose levels in type 2 diabetic rats.
 Materials and Methods: 21 male wistar rats (220±10 g) were included. Then diabetic rats were randomly assigned into 3 groups: 1) control (no-training), 2) resistance training and 3) aerobic training. Exercise training lasted 10 weeks and 5 times weekly for training groups. After intervention, NeruoD1 and PDX1 expression in pancreas, insulin and glucose were compared between groups. Data compared by one way ANOVA and Tukey's post hoc test between groups (P < 0.05).
 Results: Compared to control rats, resistance and aerobic training led to significant increase in serum insulin (P: 0.001, P: 0.013 respectively), PDX1 expression (P: 0.001, P: 0.001 respectively) and decrease glucose (P: 0.001, P: 0.001 respectively). Significant difference was not observed between control and aerobic groups with regard to NeruoD1 expression (P: 0.077). In addition, NeruoD1 expression in resistance group was significantly higher than aerobic and control groups (P: 0.018, P: 0.001, respectively).
 Conclusion: Despite the improvement of insulin and glucose in response to both aerobic and resistance training, it seems that resistance training affects genes affecting insulin synthesis and transcription more than aerobic training

EDEM1 regulates the insulin mRNA level by inhibiting the endoplasmic reticulum stress-induced IRE1/JNK/c-Jun pathway

Pancreatic beta cells produce and secrete insulin as a response to rises in blood glucose. Despite the advances in understanding glucose-regulated insulin transcription and translation the mechanisms triggering the synthesis of new insulin molecules are still incompletely described. In this report, we identify EDEM1 as a new modulator of insulin synthesis and secretion. In the presence of EDEM1, INS-1E cells secrete significantly more insulin upon glucose stimulation compared to control cells. We found that overexpression of EDEM1 inhibited the IRE1/JNK/c-Jun pathway, leading to an increase in the insulin mRNA level. Similarly, EDEM1 transduced human islets secreted significantly more insulin upon stimulation. Furthermore, EDEM1 improved insulin secretion restoring normoglycemia and glucose tolerance in diabetic rats. We propose EDEM1 as a regulator of the UPR via IRE1/XBP1s and IRE1/JNK/c-Jun signaling cascades and insulin transcription in pancreatic β-cells, supporting EDEM1 as a potential target for the treatment of diabetes.

TFEB and TFE3 control glucose homeostasis by regulating insulin gene expression.

To fulfill their function, pancreatic beta cells require precise nutrient-sensing mechanisms that control insulin production. Transcription factor EB (TFEB) and its homolog TFE3 have emerged as crucial regulators of the adaptive response of cell metabolism to environmental cues. Here, we show that TFEB and TFE3 regulate beta-cell function and insulin gene expression in response to variations in nutrient availability. We found that nutrient deprivation in beta cells promoted TFEB/TFE3 activation, which resulted in suppression of insulin gene expression. TFEB overexpression was sufficient to inhibit insulin transcription, whereas beta cells depleted of both TFEB and TFE3 failed to suppress insulin gene expression in response to amino acid deprivation. Interestingly, ChIP-seq analysis showed binding of TFEB to super-enhancer regions that regulate insulin transcription. Conditional, beta-cell-specific, Tfeb-overexpressing, and Tfeb/Tfe3 double-KO mice showed severe alteration of insulin transcription, secretion, and glucose tolerance, indicating that TFEB and TFE3 are important physiological mediators of pancreatic function. Our findings reveal a nutrient-controlled transcriptional mechanism that regulates insulin production, thus playing a key role in glucose homeostasis at both cellular and organismal levels.

85-OR: Swi/Snf Controls Transcriptional Programs Required for ß-Cell Development and Postnatal Islet Function

A key feature of diabetes is insufficient functional β-cell mass. Strategies to augment functional β-cell mass include directed differentiation of stem cells towards a β-cell fate, which requires extensive knowledge of transcriptional programs governing differentiation in vivo. Pdx1, an essential transcription factor involved in β-cell differentiation, dynamically recruits coregulators to drive different gene expression programs. A recent study identified the ATP-dependent Swi/Snf chromatin remodeling complex as an important Pdx1-interacting partner. In the developing pancreas, conditional removal of the Brg Swi/Snf ATPase subunit compromised final pancreas mass, and loss of both ATPase subunits, Brg1 and Brm, led to glucose dyshomeostasis. While Swi/Snf is essential during early pancreas development and mature β-cell function, the role of Swi/Snf during endocrine cell development has yet to be explored. Here, we generated a conditional knockout of either Brg1 (Brg1ΔendoBrm+/-), Brm (Brg1Δendo/+Brm-/-), or both subunits (DKOΔendo) during endocrine development using Neurogenin 3 Cre. Four-week-old Brg1ΔendoBrm+/- mice are glucose intolerant, hyperglycemic, and hypoinsulinemic, with no phenotype observed in Brg1Δendo/+Brm-/-mice. Impaired glucose homeostasis of Brg1ΔendoBrm+/- mice was due in part to reduced islet number, impaired insulin secretion, and altered gene expression programs. DKOΔendo mice have not been found at weaning; however, postnatal day 6 DKOΔendo mice are severely hyperglycemic with reduced plasma insulin levels. Single-cell RNA-Seq of embryonic day 15.5 lineage-labeled cells revealed reduced insulin and glucagon transcript in the β-cell and α-cell clusters, respectively, in both Brg1ΔendoBrm+/- and DKOΔendo mice and reduced Neurogenin 3­ transcript in DKOΔendo endocrine progenitor clusters. These data suggest Swi/Snf plays a critical role in governing gene-expression programs essential for endocrine cell development. Disclosure R.K.Davidson: Employee; Eli Lilly and Company. S.Kanojia: None. M.E.Osmulski: None. J.Spaeth: None. Funding National Institutes of Health (R01DK129287, R03DK127129)

Type I Diabetes in Zebrafish Reduces Sperm Quality and Increases Insulin and Glucose Transporter Transcripts.

Type I diabetes is a prominent human pathology with increasing incidence in the population; however, its cause is still unknown. This disease promotes detrimental effects on reproduction, such as lower sperm motility and DNA integrity. Hence, the investigation of the underlying mechanisms of this metabolic disturbance in reproduction and its transgenerational consequences is of the utmost importance. The zebrafish is a useful model for this research considering its high homology with human genes as well as its fast generation and regeneration abilities. Therefore, we aimed to investigate sperm quality and genes relevant to diabetes in the spermatozoa of Tg(ins:nfsb-mCherry) zebrafish, a model for type I diabetes. Diabetic Tg(ins:nfsb-mCherry) males showed significantly higher expression of transcripts for insulin a (insa) and glucose transporter (slc2a2) compared to controls. Sperm obtained from the same treatment group showed significantly lower sperm motility, plasma membrane viability, and DNA integrity compared to that from the control group. Upon sperm cryopreservation, sperm freezability was reduced, which could be a consequence of poor initial sperm quality. Altogether, the data showed similar detrimental effects related to type I diabetes in zebrafish spermatozoa at the cellular and molecular levels. Therefore, our study validates the zebrafish model for type I diabetes research in germ cells.

Ethyl-acetate fraction from a cinnamon-cortex extract protects pancreatic β-cells from oxidative stress damage.

Background: The pathogenesis of diabetes mellitus is mediated mainly by oxidative stress produced by damaged pancreatic β-cells. We identified that an ethyl-acetate fraction (EA) from a cinnamon-cortex extract (CCE) is rich in flavonoid, and showed no toxicity to β cells. Objective: In this study, we evaluated the pharmacologic activities of EA on pancreatic β cells using a model of oxidative stress induced by H2O2 or alloxan. Results: The results showed that EA could significantly reduce reactive oxygen (ROS) accumulation to improve the survival of cells. Western blot showed that EA treatment upregulated expression of nuclear factor erythroid 2 related factor 2, heme oxygenase-1, and gamma glutamylcysteine synthetase. The same model study found that EA also can protect β cells against the apoptosis induced by oxidative stress. Furthermore, EA can enhance insulin secretion in rat and mouse β cell lines treated or not with alloxan or H2O2. The expression of the insulin transcription factor PDX-1 increased in an EA concentration-dependent manner. At last, the major functional compounds of EA analysis showed that three compounds, cinnamyl alcohol, coumarin, and cinnamic acid, had similar effects as EA. Conclusions: In sum, our data suggested that EA fraction from CCE can protect β cells from oxidative stress, and increase insulin secretion to improve the function of β cells. This function might be due to these three compounds found in EA. Our findings provide a theoretical basis and functional molecules for the use of CCE against diabetes mellitus.

Differential methylation of microRNA encoding genes may contribute to high myopia.

Introduction: High myopia (HM), an eye disorder with a refractive error ≤-6.0 diopters, has multifactorial etiology with environmental and genetic factors involved. Recent studies confirm the impact of alterations in DNA methylation and microRNAs (miRNAs) on myopia. Here, we studied the combined aspects evaluating to the role of methylation of miRNA encoding genes in HM. Materials and Methods: From the genome-wide DNA methylation data of 18 Polish children with HM and 18 matched controls, we retrieved differentially methylated CG dinucleotides localized in miRNA encoding genes. Putative target genes of the highest-ranked miRNAs were obtained from the miRDB and included in overrepresentation analyses in the ConsensusPathDB. Expression of target genes was assessed using the RNA sequencing data of retinal ARPE-19 cell line. Results: We identified differential methylation of CG dinucleotides in promoter regions of MIR3621, MIR34C, MIR423 (increased methylation level), and MIR1178, MIRLET7A2, MIR885, MIR548I3, MIR6854, MIR675, MIRLET7C, MIR99A (decreased methylation level) genes. Several targets of these miRNAs, e.g. GNAS, TRAM1, CTNNB1, EIF4B, TENM3 and RUNX were previously associated with myopia/HM/refractive error in Europeans in genome-wide association studies. Overrepresentation analyses of miRNAs' targets revealed enrichment in pathways/processes related to eye structure/function, such as axon guidance, transcription, focal adhesion, and signaling pathways of TGF-β, insulin, MAPK and EGF-EGFR. Conclusion: Differential methylation of indicated miRNA encoding genes might influence their expression and contribute to HM pathogenesis via disrupted regulation of transcription of miRNAs' target genes. Methylation of genes encoding miRNAs may be a new direction in research on both the mechanisms determining HM and non-invasive indicators in diagnostics.

Effect of hydroalcoholic seed extract of Nigella sativa on hepatic and pancreatic factors of Nrf2 and FGF21 in the regulation of insulin transcription factors of MafA and PDX-1 in streptozotocin-treated diabetic rats

IntroductionNigella sativa (N. sativa), one of the most commonly used medicinal herbs with antioxidant properties, increases blood insulin levels and lowers fasting blood sugar. Nuclear Erythroid Factor-Related Factor 2 (Nrf2) and Fibroblast Growth Factor 21 (FGF21) are two antioxidant factors that are increased by oxidative stress and hyperglycemia. The present study investigated how hydroalcoholic extract of N. sativa seed (HENS) increases blood insulin levels, taking into account changes in antioxidant factors and expression of insulin transcription factors.Materials and methodsTwo groups of male diabetic wistar rats were treated orally with HESN at doses of 200 and 400 mg/kg-body weight for one month. Fasting blood sugar (FBS) and insulin were measured using standard kits by photometric and ELISA methods, respectively. The expression levels of the Nrf2, FGF21 and β-Klotho genes as well as the insulin gene-stimulating transcription factors of MafA and PDX-1 were evaluated using real-time PCR. Oxidative stress was assessed by assessing serum total oxidation status (TOS), malondialdehyde (MDA), and total antioxidant capacity (TAC).ResultsHSEN showed a significant reducing effect on FBS and oxidative biomarkers and an increasing effect on serum insulin levels in treated diabetic rats compared to untreated diabetics (P < 0.05). The elevated levels of NRF2 and FGF21 in the liver and pancreas of the diabetic control group were significantly reduced after treatment with both HESN doses (P < 0.05). Following the ameliorative effects of HENS on pancreatic tissue and the reduction of oxidative stress, the expression level of MafA and PDX1 genes approached the level of these factors in healthy rats (P < 0.05).ConclusionThis study showed the therapeutic effects of HENS on diabetic pancreas by reducing oxidative stress and tissue damage, modifying the expression levels of PDX-1 and MafA genes, and regulating insulin secretion and blood glucose levels.

Inhibition of pancreatic EZH2 restores progenitor insulin in T1D donor

Type 1 diabetes (T1D) is an autoimmune disease that selectively destroys insulin-producing β-cells in the pancreas. An unmet need in diabetes management, current therapy is focussed on transplantation. While the reprogramming of progenitor cells into functional insulin-producing β-cells has also been proposed this remains controversial and poorly understood. The challenge is determining why default transcriptional suppression is refractory to exocrine reactivation. After the death of a 13-year-old girl with established insulin-dependent T1D, pancreatic cells were harvested in an effort to restore and understand exocrine competence. The pancreas showed classic silencing of β-cell progenitor genes with barely detectable insulin (Ins) transcript. GSK126, a highly selective inhibitor of EZH2 methyltransferase activity influenced H3K27me3 chromatin content and transcriptional control resulting in the expression of core β-cell markers and ductal progenitor genes. GSK126 also reinstated Ins gene expression despite absolute β-cell destruction. These studies show the refractory nature of chromatin characterises exocrine suppression influencing β-cell plasticity. Additional regeneration studies are warranted to determine if the approach of this n-of-1 study generalises to a broader T1D population.

Mitchell-Riley Syndrome: Improving Clinical Outcomes and Searching for Functional Impact of RFX-6 Mutations.

Aims/HypothesisCaused by biallelic mutations of the gene encoding the transcription factor RFX6, the rare Mitchell–Riley syndrome (MRS) comprises neonatal diabetes, pancreatic hypoplasia, gallbladder agenesis or hypoplasia, duodenal atresia, and severe chronic diarrhea. So far, sixteen cases have been reported, all with a poor prognosis. This study discusses the multidisciplinary intensive clinical management of 4 new cases of MRS that survived over the first 2 years of life. Moreover, it demonstrates how the mutations impair the RFX6 function.MethodsClinical records were analyzed and described in detail. The functional impact of two RFX6R181W and RFX6V506G variants was assessed by measuring their ability to transactivate insulin transcription and genes that encode the L-type calcium channels required for normal pancreatic beta-cell function.ResultsAll four patients were small for gestational age (SGA) and prenatally diagnosed with duodenal atresia. They presented with neonatal diabetes early in life and were treated with intravenous insulin therapy before switching to subcutaneous insulin pump therapy. All patients faced recurrent hypoglycemic episodes, exacerbated when parenteral nutrition (PN) was disconnected. A sensor-augmented insulin pump therapy with a predictive low-glucose suspension system was installed with good results. One patient had a homozygous c.1517T>G (p.Val506Gly) mutation, two patients had a homozygous p.Arg181Trp mutation, and one patient presented with new compound heterozygosity. The RFX6V506G and RFX6R181W mutations failed to transactivate the expression of insulin and genes that encode L-type calcium channel subunits required for normal pancreatic beta-cell function.Conclusions/InterpretationMultidisciplinary and intensive disease management improved the clinical outcomes in four patients with MRS, including adjustment of parenteral/oral nutrition progression and advanced diabetes technologies. A better understanding of RFX6 function, in both intestine and pancreas cells, may break ground in new therapies, particularly regarding the use of drugs that modulate the enteroendocrine system.

The Genetics of Type 2 Diabetes in Youth: Where We Are and the Road Ahead

The Genetics of Type 2 Diabetes in Youth: Where We Are and the Road Ahead

Valproic Acid Initiates Transdifferentiation of the Human Ductal Adenocarcinoma Cell-line Panc-1 Into α-Like Cells.

Non-mesenchymal pancreatic cells are a potential source for cell replacement. Their transdifferentiation can be achieved by triggering epigenetic remodeling through e. g. post-translational modification of histones. Valproic acid, a branched-chain saturated fatty acid with histone deacetylase inhibitor activity, was linked to the expression of key transcription factors of pancreatic lineage in epithelial cells and insulin transcription. However, the potential of valproic acid to cause cellular reprogramming is not fully understood. To shed further light on it we employed next-generation RNA sequencing, real-time PCR, and protein analyses by ELISA and western blot, to assess the impact of valproic acid on transcriptome and function of Panc-1-cells. Our results indicate that valproic acid has a significant impact on the cell cycle, cell adhesion, histone H3 acetylation, and metabolic pathways as well as the initiation of epithelial-mesenchymal transition through acetylation of histone H3 resulting in α-cell-like characteristics. We conclude that human epithelial pancreatic cells can be transdifferentiated into cells with endocrine properties through epigenetic regulation by valproic acid favoring an α-cell-like phenotype.

Analysis of Half a Billion Datapoints Across Ten Machine-Learning Algorithms Identifies Key Elements Associated With Insulin Transcription in Human Pancreatic Islet Cells.

Machine learning (ML)-workflows enable unprejudiced/robust evaluation of complex datasets. Here, we analyzed over 490,000,000 data points to compare 10 different ML-workflows in a large (N=11,652) training dataset of human pancreatic single-cell (sc-)transcriptomes to identify genes associated with the presence or absence of insulin transcript(s). Prediction accuracy/sensitivity of each ML-workflow was tested in a separate validation dataset (N=2,913). Ensemble ML-workflows, in particular Random Forest ML-algorithm delivered high predictive power (AUC=0.83) and sensitivity (0.98), compared to other algorithms. The transcripts identified through these analyses also demonstrated significant correlation with insulin in bulk RNA-seq data from human islets. The top-10 features, (including IAPP, ADCYAP1, LDHA and SST) common to the three Ensemble ML-workflows were significantly dysregulated in scRNA-seq datasets from Ire-1αβ-/- mice that demonstrate dedifferentiation of pancreatic β-cells in a model of type 1 diabetes (T1D) and in pancreatic single cells from individuals with type 2 Diabetes (T2D). Our findings provide direct comparison of ML-workflows in big data analyses, identify key elements associated with insulin transcription and provide workflows for future analyses.

TAZ promotes PDX1-mediated insulinogenesis.

Transcriptional co-activator with PDZ-binding motif (TAZ) is a key mediator of the Hippo signaling pathway and regulates structural and functional homeostasis in various tissues. TAZ activation is associated with the development of pancreatic cancer in humans, but it is unclear whether TAZ directly affects the structure and function of the pancreas. So we sought to identify the TAZ function in the normal pancreas. TAZ defect caused structural changes in the pancreas, particularly islet cell shrinkage and decreased insulin production and β-cell markers expression, leading to hyperglycemia. Interestingly, TAZ physically interacted with the pancreatic and duodenal homeobox 1 (PDX1), a key insulin transcription factor, through the N-terminal domain of TAZ and the homeodomain of PDX1. TAZ deficiency decreased the DNA-binding and transcriptional activity of PDX1, whereas TAZ overexpression promoted PDX1 activity and increased insulin production even in a low glucose environment. Indeed, high glucose increased insulin production by turning off the Hippo pathway and inducing TAZ activation in pancreatic β-cells. Ectopic TAZ overexpression along with PDX1 activation was sufficient to produce insulin in non-β-cells. TAZ deficiency impaired the mesenchymal stem cell differentiation into insulin-producing cells (IPCs), whereas TAZ recovery restored normal IPCs differentiation. Compared to WT control, body weight increased in TAZ-deficient mice with age and even more with a high-fat diet (HFD). TAZ deficiency significantly exacerbated HFD-induced glucose intolerance and insulin resistance. Therefore, TAZ deficiency impaired pancreatic insulin production, causing hyperglycemia and exacerbating HFD-induced insulin resistance, indicating that TAZ may have a beneficial effect in treating insulin deficiency in diabetes.