Heterogeneous Nuclear Ribonucleoprotein F Mediates Insulin Inhibition of Bcl2-Modifying Factor Expression and Tubulopathy in Diabetic Kidney

Anindya Ghosh,John S D Chan,Shuiling Zhao,Chao-Sheng Lo,Shaaban Abdo,Janos G Filep,Muhammad Abdul Lateef,Shao-Ling Zhang,Isabelle Chenier,Julie R Ingelfinger,Hasna Maachi

doi:10.1038/s41598-019-43218-2

Abstract

We investigated the molecular mechanism(s) by which insulin prevents Bcl2-modifying factor (Bmf)-induced renal proximal tubular cell (RPTC) apoptosis and loss in diabetic mice. Transgenic mice (Tg) mice specifically overexpressing human BMF in RPTCs and non-Tg littermates were studied at 10 to 20 weeks of age. Non-diabetic littermates, diabetic Akita mice +/− insulin implant, Akita Tg mice specifically overexpressing heterogeneous nuclear ribonucleoprotein F (hnRNP F) in their RPTCs and immortalized rat renal proximal tubular cells (IRPTCs) were also studied. BMF-Tg mice exhibited higher systolic blood pressure, urinary albumin/creatinine ratio, RPTC apoptosis and urinary RPTCs than non-Tg mice. Insulin treatment in Akita mice and Akita mice overexpressing hnRNP F suppressed Bmf expression and RPTC apoptosis. In hyperinsulinemic-euglycemic wild type mice, renal Bmf expression was down-regulated with up-regulation of hnRNP F. In vitro, insulin inhibited high glucose-stimulation of Bmf expression, predominantly via p44/42 mitogen-activated protein kinase (MAPK) signaling. Transfection of p44/42 MAPK or hnRNP F small interfering RNA (siRNA) prevented insulin inhibition of Bmf expression. HnRNP F inhibited Bmf transcription via hnRNP F-responsive element in the Bmf promoter. Our results demonstrate that hnRNP F suppression of Bmf transcription is an important mechanism by which insulin protects RPTCs from apoptosis in diabetes.

Highlights

Www.nature.com/scientificreports apoptosis in type 1 diabetic (T1D) Agt-Transgenic mice (Tg) mice[18,19]
We previously reported that insulin inhibits high-glucose stimulation of renal rat Agt gene expression and renal proximal tubular cell (RPTC) hypertrophy through a putative insulin-responsive element (IRE) in the rat Agt gene promoter that interacts with 2 nuclear proteins, heterogeneous nuclear ribonucleoprotein F and K in vitro[25,26,27]
Tissue-specific analysis by RT-polymerase chain reaction (PCR) confirmed Human Bcl2 Modifying Factor (hBMF) mRNA expression in the kidney cortex and RPTs isolated from male hBMF-Tg mice, as well as in the kidney cortex of female hBMF-Tg mice implanted with testosterone pellet but not in other tissues from both male and female mice (Fig. 1c). hBMF transgene was detected in RPTs of male hBMF-Tg mice but not in non-Tg mice and can be differentiated from endogenous mouse Bcl2-modifying factor (Bmf) using primers specific for hBMF and mouse Bmf in RT-PCR, respectively (Fig. 1d)

Summary

Introduction

Www.nature.com/scientificreports apoptosis in type 1 diabetic (T1D) Agt-Tg mice[18,19]. Overexpression of hnRNP F inhibited renal Agt gene expression and attenuated hypertension, kidney hypertrophy and RPTC apoptosis in Akita (T1D) and db/db Tg mice[28,29]. We further reported that hnRNP F/K mediate insulin inhibition of renal Agt gene expression and that insulin stimulates hnRNP F/K expression through p44/42 MAPK signaling pathway but not through the phosphatidylinositol 3-kinase (PI-3K) pathway in diabetic mice[30,31]. These findings suggest that insulin inhibition of renal Agt gene transcription and RPTC apoptosis occurs via hnRNP F/K in diabetes. We investigated the impact of Bmf overexpression on RPTC apoptosis in BMF-Tg mice and examined whether hnRNP F mediates, at least in part, insulin regulation of Bmf actions in Akita mice and in rat immortalized RPTCs (IRPTCs) cultured in HG milieu

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