Aldo-ketoreductase 1c19 ablation does not affect insulin secretion in murine islets

Beta cell failure is a critical feature of diabetes. It includes defects of insulin production, secretion, and altered numbers of hormone-producing cells. In previous work, we have shown that beta cell failure is mechanistically linked to loss of Foxo1 function. This loss of function likely results from increased Foxo1 protein degradation, due to hyperacetylation of Foxo1 from increased nutrient turnover. To understand the mechanisms of Foxo1-related beta cell failure, we performed genome-wide analyses of its target genes, and identified putative mediators of sub-phenotypes of cellular dysfunction. Chromatin immunoprecipitation analyses demonstrated a striking pattern of Foxo1 binding to the promoters of a cluster of aldo-ketoreductases on chromosome 13: Akr1c12, Akr1c13, Akr1c19. Of these, Akr1c19 has been reported as a marker of Pdx1-positive endodermal progenitor cells. Here we show that Akr1c19 expression is dramatically decreased in db/db islets. Thus, we investigated whether Akr1c19 is involved in beta cell function. We performed gain- and loss-of-function experiments in cultured beta cells and generated Akr1c19 knockout mice. We show that Foxo1 and HNF1a cooperatively regulate Akr1c19 expression. Nonetheless, functional characterization of Akr1c19 both using islets and knockout mice did not reveal abnormalities on glucose homeostasis. We conclude that reduced expression of Akr1c19 is not sufficient to affect islet function

Nascent RNA interaction keeps PRC2 activity poised and in check

Polycomb-repressive complex 2 (PRC2) facilitates the maintenance and inheritance of chromatin domains repressive to transcription through catalysis of methylation of histone H3 at Lys27 (H3K27me2/3). However, through its EZH2 subunit, PRC2 also binds to nascent transcripts from active genes that are devoid of H3K27me2/3 in embryonic stem cells. Here, biochemical analyses indicated that RNA interaction inhibits SET domain-containing proteins, such as PRC2, nonspecifically in vitro. However, CRISPR-mediated truncation of a PRC2-interacting nascent RNA rescued PRC2-mediated deposition of H3K27me2/3. That PRC2 activity is inhibited by interactions with nascent transcripts supports a model in which PRC2 can only mark for repression those genes silenced by transcriptional repressors

Phosphorylation of the PRC2 component Ezh2 is cell cycle-regulated and up-regulates its binding to ncRNA

Ezh2 functions as a histone H3 Lys 27 (H3K27) methyltransferase when comprising the Polycomb-Repressive Complex 2 (PRC2). Trimethylation of H3K27 (H3K27me3) correlates with transcriptionally repressed chromatin. The means by which PRC2 targets specific chromatin regions is currently unclear, but noncoding RNAs (ncRNAs) have been shown to interact with PRC2 and may facilitate its recruitment to some target genes. Here we show that Ezh2 interacts with HOTAIR and Xist. Ezh2 is phosphorylated by cyclin-dependent kinase 1 (CDK1) at threonine residues 345 and 487 in a cell cycle-dependent manner. A phospho-mimic at residue 345 increased HOTAIR ncRNA binding to Ezh2, while the phospho-mimic at residue 487 was ineffectual. An Ezh2 domain comprising T345 was found to be important for binding to HOTAIR and the 5′ end of Xist

Notch-mediated Ephrin signaling disrupts islet architecture and ß cell function

Altered islet architecture is associated with β cell dysfunction and type 2 diabetes (T2D) progression, but molecular effectors of islet spatial organization remain mostly unknown. Although Notch signaling is known to regulate pancreatic development, we observed “reactivated” β cell Notch activity in obese mouse models. To test the repercussions and reversibility of Notch effects, we generated doxycycline-dependent, β cell–specific Notch gain-of-function mice. As predicted, we found that Notch activation in postnatal β cells impaired glucose-stimulated insulin secretion and glucose intolerance, but we observed a surprising remnant glucose intolerance after doxycycline withdrawal and cessation of Notch activity, associated with a marked disruption of normal islet architecture. Transcriptomic screening of Notch-active islets revealed increased Ephrin signaling. Commensurately, exposure to Ephrin ligands increased β cell repulsion and impaired murine and human pseudoislet formation. Consistent with our mouse data, Notch and Ephrin signaling were increased in metabolically inflexible β cells in patients with T2D. These studies suggest that β cell Notch/Ephrin signaling can permanently alter islet architecture during a morphogenetic window in early life.This work was supported by NIH DK103818 (to UBP), an NCI Outstanding Investigator Award R35 CA197745 (to AC), a Russell Berrie Foundation Award (to AB), and American Diabetes Association Grant 1-17-PMF-025 (to AB)

S1 Data -

Diabetes mellitus is characterized by insulin resistance and β-cell failure. The latter involves impaired insulin secretion and β-cell dedifferentiation. Sulfonylurea (SU) is used to improve insulin secretion in diabetes, but it suffers from secondary failure. The relationship between SU secondary failure and β-cell dedifferentiation has not been examined. Using a model of SU secondary failure, we have previously shown that functional loss of oxidoreductase Cyb5r3 mediates effects of SU failure through interactions with glucokinase. Here we demonstrate that SU failure is associated with partial β-cell dedifferentiation. Cyb5r3 knockout mice show more pronounced β-cell dedifferentiation and glucose intolerance after chronic SU administration, high-fat diet feeding, and during aging. A Cyb5r3 activator improves impaired insulin secretion caused by chronic SU treatment, but not β-cell dedifferentiation. We conclude that chronic SU administration affects progression of β-cell dedifferentiation and that Cyb5r3 activation reverses secondary failure to SU without restoring β-cell dedifferentiation.</div

Reflections on the state of diabetes research and prospects for treatment.

Research on the etiology and treatment of diabetes has made substantial progress. As a result, several new classes of anti-diabetic drugs have been introduced in clinical practice. Nonetheless, the number of patients achieving glycemic control targets has not increased for the past 20 years. Two areas of unmet medical need are the restoration of insulin sensitivity and the reversal of pancreatic beta cell failure. In this review, we integrate research advances in transcriptional regulation of insulin action and pathophysiology of beta cell dedifferentiation with their potential impact on prospects of a durable cure for patients suffering from type 2 diabetes

Chronic GLB treatment increases marker of β-cell dedifferentiation.

a: Representative Aldh1a3 staining (green) in islets from pancreatic sections of mice treated with GLB for 6 weeks (n = 4). Scale bars: 50 μm. b-d: Quantification of Aldh1a3 positive cells (B), Aldh1a3 mRNA expression (c) and Aldh1a3 protein levels (d) in islets from treated with GLB (n = 4). e: Representative images from pancreatic sections of mice treated with GLB stained with Aldh1a3 (green), Pdx1 (red), and DAPI (blue). Scale bars: 50 μm. f: Relative mRNA expressions of FoxO1, Cyb5r3, MafA, Pdx1, Ngn3, Ins1, Ins2, Kir6.2 and Sur1 in islets from mice treated with GLB (n = 4). g and h: Plasma insulin (g) and blood glucose (h) levels in mice treated with GLB under free feeding condition (n = 4–6). i and j: Plasma insulin (i) and blood glucose (j) levels in mice treated with GLB during glucose tolerance test (n = 4). *P < 0.05 by unpaired Student’s t-test. Data represent means ± SEM.</p

Chronic GLB treatment further impairs glucose-induced insulin secretion in Cyb5r3 βKO.

a and b: Plasma insulin (a) blood glucose (b) levels in WT and Cyb5r3 βKO treated with GLB under free feeding condition (n = 4). c and d: Plasma insulin level and its area under the curve (c) and blood glucose levels and its area under the curve (d) in WT and Cyb5r3 βKO treated with GLB during glucose tolerance test (n = 4). *P < 0.05 by two-way ANOVA followed by Tukey’s test. Data represent means ± SEM.</p

THII does not reduce β-cell dedifferentiation by chronic GLB treatment.

a: Representative Aldh1a3 staining (green) in islets from pancreatic sections of mice treated with GLB and THII for 6 weeks (n = 4). Scale bars: 50 μm. b and c: Quantification of Aldh1a3 positive cells (b) and Aldh1a3 mRNA expression (c) in islets from mice treated with GLB and THII (n = 4). d: Relative mRNA expressions of FoxO1, Cyb5r3, MafA, Pdx1, Ngn3, Ins1, Ins2, Kir6.2 and Sur1 in islets from mice treated with GLB and THII (n = 4). *P < 0.05 by two-way ANOVA followed by Tukey’s test. Data represent means ± SEM.</p

THII improves impaired glucose-induced insulin secretion in GLB treated mice.

a and b: Plasma insulin (a) blood glucose (b) levels in mice treated with GLB and THII under free feeding condition (n = 4). c and d: Plasma insulin level and its area under the curve (a) and blood glucose levels and its area under the curve (d) in mice treated with GLB and THII during glucose tolerance test (n = 4). *P < 0.05 by two-way ANOVA followed by Tukey’s test. Data represent means ± SEM.</p