The Role of ChREBP in [beta]-cell Function and Dysfunction

Research output: ThesisPh.D. thesis

Abstract

Type 2 diabetes mellitus (T2DM) is a pandemic-like disease, which globally decreases the quality of life for hundreds of millions. It has multiple etiologies, but deregulation of the glucose homeostasis by both decreased insulin secretion and sensitivity is a common theme. A new paradigm focused on the pancreatic β-cells has recently emerged. Glucose is a crucial fuel for β-cells, as it acutely stimulates insulin exocytosis. In contrast, long term exposure leads to dysfunction by so-called glucotoxic mechanisms. Thus, there is a critical balance between function and dysfunction in the response to glucose. The primary aim of this thesis is to dissect the transcriptional mechanisms underlying these pleiotropic effects of glucose.

The work in this thesis is split into three different parts. In Part 1, two different new bioinformatic tools are presented. The first, iRNA-seq, is a new tool that leverages reads mapping to introns to accurately estimate transcription. Comparison between iRNA-seq and current gold standard techniques shows that iRNA-seq is approximately as sensitive as the competing methods. However, unlike the competing methods, which are all very labor intensive and demanding in terms of sample material, iRNA-seq is cheap, easy and requires little sample. In conclusion, iRNA-seq is an attractive alternative to current methods for determination of changes in transcriptional activity on a genome-wide level. The second tool, IAMET, integrates motif searching, machine learning and analysis of gene expression data to identify putative transcriptional regulators. Comparison between IAMET and current alternative methods demonstrates that IAMET has the highest specificity. Furthermore, IAMET has the added benefits of using a vastly expanded library of transcription factor motifs, and directly identify causal transcription factors. In conclusion, IAMET is a powerful new integrated method for the identification of transcriptional regulators.

In Part 2, we utilize these novel tools and other techniques to dissect the glucose-induced transcriptional response in pancreatic β-cells. Two studies are presented. In the first study, we show that high glucose drastically changes the transcriptional landscape of β-cells in a biphasic manner. In the late phase, many genes that are involved in regulation of β-cell function are repressed, whereas a large proportion of the late induced genes are involved in proliferation. We provide evidence that the master regulator of this response is the carbohydrate response element binding protein (ChREBP): Overexpression of a constitutive active ChREBP mimics the transcriptional response to glucose, and ChREBP knock down blunts it. However, whereas ChREBP directly regulates the early induced genes, it is indirectly associated with the late genes. We find that that ChREBP induces a novel transcription factor, Rorγ, which is necessary for induction of proliferation in both INS-1E cells and in islet of Langerhans. In the second study, we examine the glucosesensitive interactome of Mediator complex subunit 1 (Med1) in β-cells. We find some transcriptional regulators that interact with Med1 in a glucose-dependent manner. Knock down of these reveal that they regulate different gene programs in β-cells. One of the factors, Mybbp1a is necessary for the induction of genes involved in proliferation, but not for the repression of genes involved in β-cell function. Mybbp1a interacts with several transcription factors, and based on gene expression and motif analysis, we hypothesize that it acts through either Myc or Zbtb7b. However, this needs further validation.

In Part 3, we study the dynamics of enhancer-promoter looping. The two key points of this study are that the enhancer-promoter interactions are functionally important, and are dynamically regulated. The potential implication of this work is that enhancer-promoter looping should be regarded as an active means of regulating transcription, rather than a passive process. This initial conceptual study was performed in 3T3-L1 adipocytes, and it still remains to be elucidated if similar regulatory mechanisms apply to β-cells.
Original languageEnglish
Awarding Institution
  • University of Southern Denmark
Supervisors/Advisors
  • Mandrup, Susanne, Supervisor
Place of PublicationOdense
Publisher
Publication statusPublished - 2017

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