Loss of RREB1 in pancreatic beta cells reduces cellular insulin content and affects endocrine cell gene expression

Katia K Mattis; Nicole A J Krentz; Christoph Metzendorf; Fernando Abaitua; Aliya F Spigelman; Han Sun; Jennifer M Ikle; Swaraj Thaman; Antje K Rottner; Austin Bautista; Eugenia Mazzaferro; Marta Perez-Alcantara; Jocelyn E Manning Fox; Jason M Torres; Agata Wesolowska-Andersen; Grace Z Yu; Anubha Mahajan; Anders Larsson; Patrick E MacDonald; Benjamin Davies; Marcel den Hoed; Anna L Gloyn

doi:10.1007/s00125-022-05856-6

Loss of RREB1 in pancreatic beta cells reduces cellular insulin content and affects endocrine cell gene expression

Diabetologia. 2023 Apr;66(4):674-694. doi: 10.1007/s00125-022-05856-6. Epub 2023 Jan 12.

Authors

Katia K Mattis^{1

2}, Nicole A J Krentz^{2

3}, Christoph Metzendorf⁴, Fernando Abaitua², Aliya F Spigelman^{5

6}, Han Sun³, Jennifer M Ikle³, Swaraj Thaman³, Antje K Rottner¹, Austin Bautista^{5

6}, Eugenia Mazzaferro⁴, Marta Perez-Alcantara², Jocelyn E Manning Fox^{5

6}, Jason M Torres^{2

7}, Agata Wesolowska-Andersen², Grace Z Yu¹, Anubha Mahajan^{2

8}, Anders Larsson⁹, Patrick E MacDonald^{5

6}, Benjamin Davies², Marcel den Hoed⁴, Anna L Gloyn^{10

11

12

13}

Affiliations

¹ Oxford Centre for Diabetes, Endocrinology and Metabolism, University of Oxford, Oxford, UK.
² Wellcome Centre for Human Genetics, University of Oxford, Oxford, UK.
³ Division of Endocrinology, Department of Pediatrics, Stanford School of Medicine, Stanford University, Stanford, CA, USA.
⁴ Beijer Laboratory and Department of Immunology, Genetics and Pathology, Uppsala University and SciLifeLab, Uppsala, Sweden.
⁵ Department of Pharmacology, University of Alberta, Edmonton, AB, Canada.
⁶ Alberta Diabetes Institute, University of Alberta, Edmonton, AB, Canada.
⁷ Clinical Trial Service Unit and Epidemiological Studies Unit, Nuffield Department of Population Health, University of Oxford, Oxford, UK.
⁸ Genentech, South San Francisco, CA, USA.
⁹ Department of Medical Sciences, Clinical Chemistry, Uppsala University, Uppsala, Sweden.
¹⁰ Oxford Centre for Diabetes, Endocrinology and Metabolism, University of Oxford, Oxford, UK. agloyn@stanford.edu.
¹¹ Wellcome Centre for Human Genetics, University of Oxford, Oxford, UK. agloyn@stanford.edu.
¹² Division of Endocrinology, Department of Pediatrics, Stanford School of Medicine, Stanford University, Stanford, CA, USA. agloyn@stanford.edu.
¹³ Oxford NIHR Biomedical Research Centre, Churchill Hospital, Oxford, UK. agloyn@stanford.edu.

Abstract

Aims/hypothesis: Genome-wide studies have uncovered multiple independent signals at the RREB1 locus associated with altered type 2 diabetes risk and related glycaemic traits. However, little is known about the function of the zinc finger transcription factor Ras-responsive element binding protein 1 (RREB1) in glucose homeostasis or how changes in its expression and/or function influence diabetes risk.

Methods: A zebrafish model lacking rreb1a and rreb1b was used to study the effect of RREB1 loss in vivo. Using transcriptomic and cellular phenotyping of a human beta cell model (EndoC-βH1) and human induced pluripotent stem cell (hiPSC)-derived beta-like cells, we investigated how loss of RREB1 expression and activity affects pancreatic endocrine cell development and function. Ex vivo measurements of human islet function were performed in donor islets from carriers of RREB1 type 2 diabetes risk alleles.

Results: CRISPR/Cas9-mediated loss of rreb1a and rreb1b function in zebrafish supports an in vivo role for the transcription factor in beta cell mass, beta cell insulin expression and glucose levels. Loss of RREB1 also reduced insulin gene expression and cellular insulin content in EndoC-βH1 cells and impaired insulin secretion under prolonged stimulation. Transcriptomic analysis of RREB1 knockdown and knockout EndoC-βH1 cells supports RREB1 as a novel regulator of genes involved in insulin secretion. In vitro differentiation of RREB1^KO/KO hiPSCs revealed dysregulation of pro-endocrine cell genes, including RFX family members, suggesting that RREB1 also regulates genes involved in endocrine cell development. Human donor islets from carriers of type 2 diabetes risk alleles in RREB1 have altered glucose-stimulated insulin secretion ex vivo, consistent with a role for RREB1 in regulating islet cell function.

Conclusions/interpretation: Together, our results indicate that RREB1 regulates beta cell function by transcriptionally regulating the expression of genes involved in beta cell development and function.

Keywords: Beta cell; CRISPR/Cas9; Diabetes; Differentiation; Human genetics; Pancreatic islet; RREB1; Stem cell; Transcription factor; Zebrafish.

Publication types

Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

MeSH terms

Animals
DNA-Binding Proteins / metabolism
Diabetes Mellitus, Type 2* / genetics
Diabetes Mellitus, Type 2* / metabolism
Gene Expression
Glucose / metabolism
Humans
Induced Pluripotent Stem Cells / metabolism
Insulin / metabolism
Insulin-Secreting Cells* / metabolism
Transcription Factors / genetics
Zebrafish / genetics

Substances

DNA-Binding Proteins
Glucose
Insulin
RREB1 protein, human
Transcription Factors

Abstract

Publication types

MeSH terms

Substances

Grants and funding