Human Krüppel-like factor 11 inhibits human proinsulin promoter activity in pancreatic beta cells

Niu, Xinjie; Perakakis, Nikolaos; Laubner, Katharina; Limbert, Catarina; Stahl, T.; Brendel, Matthias; Bretzel, R. G.; Seufert, Jochen; Päth, G

doi:10.1007/s00125-007-0770-5

Cited by 3 publications

(13 citation statements)

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“…Our own research, however, could not confirm these findings. We observed that hKLF11 inhibits hInsP in rat INS-1E as well as mouse b-TC3 b-cells (Niu et al, 2007). A negative regulation of hInsP by cotransfected hKLF11 was also observed by others in mouse MIN6-m9 b-cells (Kuroda et al, 2009).…”

Section: Introductionsupporting

confidence: 78%

“…Construction of human KLF11 expression plasmid (hKLF11-pcDNA 3.1) and human insulin promoter-driven secreted alkaline phosphatase (SEAP) reporter plasmids (5 0 -deletion series À881 + 54 to À101 + 54hInsP-pSEAP) has been described previously (Niu et al, 2007). The human p300 coding sequence was subcloned from CMVß-p300 (kindly provided by Timothy J. Kieffer, University of British Columbia Vancouver, BC Canada) into the CMV promoter-driven pcDNA3.1 (Invitrogen/Life Technologies GmbH, Darmstadt, Germany) to obtain the hp300-pcDNA3.1 expression plasmid.…”

Section: Plasmid Constructionmentioning

confidence: 99%

“…A1, A3 and GG2 box of À881 + 54hInsP promoter sequence were mutated as described earlier (Le Lay and Stein, 2006). The CACCC box within a À387 + 54hInsP promoter fragment was mutated from CCCCACCCC to CCCGATTCC as described earlier (Niu et al, 2007). All mutations were performed using the QuickChange Site-Directed Mutagenesis Kit (Stratagene, Heidelberg, Germany).…”

Section: Plasmid Constructionmentioning

confidence: 99%

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Human Krüppel-like factor 11 differentially regulates human insulin promoter activity in β-cells and non-β-cells via p300 and PDX1 through the regulatory sites A3 and CACCC box

Perakakis

Danassi

Alt

et al. 2012

Molecular and Cellular Endocrinology

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Keywords: KLF11 p300 PDX1 A3 element CACCC box Human insulin promoter a b s t r a c t Human Krüppel-like factor 11 (hKLF11) has been characterised to both activate and inhibit human insulin promoter (hInsP) activity. Since KLF11 is capable to differentially regulate genes dependent on recruited cofactors, we investigated the effects of hKLF11 on cotransfected hInsP in both b-cells and non-b-cells. hKLF11 protein interacts with hp300 but not with hPDX1. Overexpressed hKLF11 stimulates PDX1-transactivation of hInsP in HEK293 non-b-cells, but confers inhibition in INS-1E b-cells. Both hKLF11 functions can be neutralised by the p300 inhibitor E1A, increased hp300 levels (INS-1E), dominant negative (DN)-PDX1 and by mutation of the PDX1 binding site A3 or the CACCC box. In summary, hKLF11 differentially regulates hInsP activity depending on the molecular context via modulation of p300:PDX1 interactions with the A3 element and CACCC box. We postulate that KLF11 has a role in fine-tuning insulin transcription in certain cellular situations rather than representing a major transcriptional activator or repressor of the insulin gene.

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Section: Introductionsupporting

confidence: 78%

Section: Plasmid Constructionmentioning

confidence: 99%

Section: Plasmid Constructionmentioning

confidence: 99%

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Human Krüppel-like factor 11 differentially regulates human insulin promoter activity in β-cells and non-β-cells via p300 and PDX1 through the regulatory sites A3 and CACCC box

Perakakis

Danassi

Alt

et al. 2012

Molecular and Cellular Endocrinology

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“…Two recent studies have shown that artificial mutations of this CCACC element lead to decreased INS promoter activity (36,42). Although previous studies failed to identify protein complexes interacting with this region in beta-cell nuclear extracts (17,36), a recent study showed that in vitro translated GLIS3, a zinc-finger transcription factor that is mutated in patients with neonatal diabetes and congenital hypothyroidism (43), exhibits sequence-specific binding to this region (42). Multiple DNA binding factors may 367_-344del (-129_-106) c. bind to CCACC elements in vitro, and thus future studies are warranted to determine if additional factors act through this element in vivo in beta cells.…”

Section: Discussionmentioning

confidence: 99%

“…This sequence is conserved among a subset of mammalian species (Fig. 3A) and mutagenesis of multiple bases neighboring this dinucleotide impairs INS promoter activity (36). We constructed insulin promoter fragments carrying the point mutations c.-331(C > G, C > A) and c.-332C > G. The point mutations induced up to 90% reduction in transcriptional activity, while a control mutation, c.-339G > A, did not alter the transcriptional activity in pancreatic beta-cell lines (Fig.…”

Section: Recessive Ins Mutations Uncover Essential Regulatory Sequencmentioning

confidence: 99%

Recessive mutations in the INS gene result in neonatal diabetes through reduced insulin biosynthesis

Garin

Edghill²,

Akerman³

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

193

219

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Heterozygous coding mutations in the INS gene that encodes preproinsulin were recently shown to be an important cause of permanent neonatal diabetes. These dominantly acting mutations prevent normal folding of proinsulin, which leads to beta-cell death through endoplasmic reticulum stress and apoptosis. We now report 10 different recessive INS mutations in 15 probands with neonatal diabetes. Functional studies showed that recessive mutations resulted in diabetes because of decreased insulin biosynthesis through distinct mechanisms, including gene deletion, lack of the translation initiation signal, and altered mRNA stability because of the disruption of a polyadenylation signal. A subset of recessive mutations caused abnormal INS transcription, including the deletion of the C1 and E1 cis regulatory elements, or three different single base-pair substitutions in a CC dinucleotide sequence located between E1 and A1 elements. In keeping with an earlier and more severe beta-cell defect, patients with recessive INS mutations had a lower birth weight (−3.2 SD score vs. −2.0 SD score) and were diagnosed earlier (median 1 week vs. 10 weeks) compared to those with dominant INS mutations. Mutations in the insulin gene can therefore result in neonatal diabetes as a result of two contrasting pathogenic mechanisms. Moreover, the recessively inherited mutations provide a genetic demonstration of the essential role of multiple sequence elements that regulate the biosynthesis of insulin in man. (8-12). In contrast, abnormalities in chromosome 6q24 are the most common cause of TNDM (13), followed by mutations in the KCNJ11 and ABCC8 genes (14). Despite these advances, the etiology of neonatal diabetes is still not known in at least 30% of patients with PNDM, suggesting other genetic causes are still to be found (9).Insulin is secreted from islet beta cells of the pancreas. Insufficient secretion of insulin results in hyperglycemia and diabetes, whereas excessive secretion results in hypoglycemia. Insulin biosynthesis and secretion are therefore tightly regulated to maintain blood glucose levels within a narrow physiological range. Extensive studies have dissected an array of cis sequence elements in the INS promoter region and their cognate DNA binding factors, which together ensure the cellular specificity and rate of INS transcription (15)(16)(17)(18)(19)(20)(21)(22). In addition, insulin biosynthesis is strongly dependent on posttranscriptional regulatory mechanisms, including the modulation of translation and stability (23-25). The latter is largely mediated through sequences located in the untranslated regions of INS transcripts (26-28).

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Identification of Minimal Promoter and Genetic Variants of Kruppel-like Factor 11 Gene and Association Analysis with Type 2 Diabetes in Japanese

et al. 2009

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Abstract. Genetic analysis of the KLF11 gene revealed two rare variants, A347S and T220M, segregating in families with early-onset type 2 diabetes, and one frequent polymorphic Q62R variant significantly associated with type 2 diabetes in Northern Europeans. Furthermore, it has been reported that over-expression of KLF11 has a deleterious effect on insulin promoter activity. Thus, an altered expression level of KLF11 may contribute to the occurrence of type 2 diabetes. To investigate the contribution of KLF11 to type 2 diabetes in Japanese, we surveyed the 5' flanking region of KLF11 by reporter assay and identified the minimal promoter region of the gene. The promoter region from -250 to +162 bp including five Sp1 binding sites showed basal promoter activity both in MIN6-m9 and HepG2 cells. We also examined the entire region of KLF11 to detect genetic variants. A total of 19 polymorphisms, six of which are novel, were identified, but none of them showed association with the occurrence of type 2 diabetes. Two of the identified polymorphisms, R29Q and S124F, are novel coding variants. Functional analyses of these variants were performed, and similarly reduced effects on transcriptional activities of insulin, catalase1, and the Smad7 gene were found. We conclude that variants of KLF11 are not a major factor in the occurrence of type 2 diabetes in Japanese. The promoter region of KLF11 identified in the present study should be useful in further elucidation of the transcriptional regulation mechanism of the gene and genetic analyses of type 2 diabetes.

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Human Krüppel-like factor 11 inhibits human proinsulin promoter activity in pancreatic beta cells

Cited by 3 publications

References 0 publications

Human Krüppel-like factor 11 differentially regulates human insulin promoter activity in β-cells and non-β-cells via p300 and PDX1 through the regulatory sites A3 and CACCC box

Human Krüppel-like factor 11 differentially regulates human insulin promoter activity in β-cells and non-β-cells via p300 and PDX1 through the regulatory sites A3 and CACCC box

Recessive mutations in the INS gene result in neonatal diabetes through reduced insulin biosynthesis

Identification of Minimal Promoter and Genetic Variants of Kruppel-like Factor 11 Gene and Association Analysis with Type 2 Diabetes in Japanese

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