Functional interplay between the transcription factors USF1 and PDX-1 and protein kinase CK2 in pancreatic \u03b2-cells

Sarah Spohrer,Rebecca Groß,Lisa Schwind,Lisa Nalbach,Emmanuel Ampofo,Michael D Menger,Claudia Götz,Heike Stumpf,Mathias Montenarh

doi:10.1038/s41598-017-16590-0

Abstract

Glucose homeostasis is regulated by insulin, which is produced in the β-cells of the pancreas. The synthesis of insulin is controlled by several transcription factors including PDX-1, USF1 and USF2. Both, PDX-1 and USF1 were identified as substrates for protein kinase CK2. Here, we have analysed the interplay of PDX-1, USF1 and CK2 in the regulation of PDX-1 gene transcription. We found that the PDX-1 promoter is dose-dependently transactivated by PDX-1 and transrepressed by USF1. With increasing glucose concentrations the transrepression of the PDX-1 promoter by USF1 is successively abrogated. PDX-1 binding to its own promoter was not influenced by glucose, whereas USF1 binding to the PDX-1 promoter was reduced. The same effect was observed after inhibition of the protein kinase activity by three different inhibitors or by using a phospho-mutant of USF1. Moreover, phosphorylation of USF1 by CK2 seems to strengthen the interaction between USF1 and PDX-1. Thus, CK2 is a negative regulator of the USF1-dependent PDX-1 transcription. Moreover, upon inhibition of CK2 in primary islets, insulin expression as well as insulin secretion were enhanced without affecting the viability of the cells. Therefore, inhibition of CK2 activity may be a promising approach to stimulate insulin production in pancreatic β-cells.

Highlights

Protein kinase CK2, which is composed of two catalytic α- or α′-subunits and two non-catalytic β-subunits, phosphorylates more than 400 different substrates of the human proteome
PDX-1 and USF1 are transcription factors deeply involved in the regulation of glucose homeostasis
Amemiya-Kudo et al reported a functional interplay of USF1 and PDX-1 at the PDX-1 promoter[16]

Summary

Introduction

Protein kinase CK2, which is composed of two catalytic α- or α′-subunits and two non-catalytic β-subunits, phosphorylates more than 400 different substrates of the human proteome. In another study it was shown that only a nuclear sub-population of CK2α and CK2β proteins bound to USFs13 One interpretation of these results might be that binding of CK2 to USFs facilitates phosphorylation of nuclear USF1. Expression of a dominant negative form of USF2 decreased the binding of USFs to the promoter, which resulted in a lower level of PDX-1 mRNA17. (g) INS-1 cells were transfected with the PDX-1 promoter construct −6500/+68-STF-luc and with FLAG-USF1 alone or in combination with the dominant-negative USF-mutant CMV566 A-USF (AUSF) or the empty vector (mock) as a control. We found that PDX-1 and USF1 interact functionally at the PDX-1 promoter in INS-1 cells The interaction of both proteins and the transcriptional activity are influenced by glucose and by the inhibition of CK2 activity. The measurable impact of CK2 inhibition in primary islets was an enhancement of insulin expression and secretion

Methods

Results

Conclusion