Selective Actions of Mitochondrial Fission/Fusion Genes on Metabolism-Secretion Coupling in Insulin-releasing Cells

Claes B. Wollheim,Piero Marchetti,Andreas Wiederkehr,Nicolas Demaurex,Yves Mattenberger,Kyu-Sang Park,Jean-Claude Martinou,Clare Kirkpatrick

doi:10.1074/jbc.m806251200

Claes B. Wollheim, Piero Marchetti + Show 6 more

Open Access

https://doi.org/10.1074/jbc.m806251200

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Abstract

Mitochondria form filamentous networks that undergo continuous fission/fusion. In the pancreatic beta-cells, mitochondria are essential for the transduction of signals linking nutrient metabolism to insulin granule exocytosis. Here we have studied mitochondrial networks in the insulinoma cell line INS-1E, primary rat and human beta-cells. We have further investigated the impact of mitochondrial fission/fusion on metabolism-secretion coupling in INS-1E cells. Overexpression of hFis1 caused dramatic mitochondrial fragmentation, whereas Mfn1 evoked hyperfusion and the aggregation of mitochondria. Cells overexpressing hFis1 or Mfn1 showed reduced mitochondrial volume, lowered cellular ATP levels, and as a consequence, impaired glucose-stimulated insulin secretion. Decreased mitochondrial ATP generation was partially compensated for by enhanced glycolysis as indicated by increased lactate production in these cells. Dominant-negative Mfn1 elicited mitochondrial shortening and fragmentation of INS-1E cell mitochondria, similar to hFis1. However, the mitochondrial volume, cytosolic ATP levels, and glucose-stimulated insulin secretion were little affected. We conclude that mitochondrial fragmentation per se does not impair metabolism-secretion coupling. Through their impact on mitochondrial bioenergetics and distribution, hFis1 and Mfn1 activities influence mitochondrial signal generation thereby insulin exocytosis.

Highlights

Overexpression of WT-mitofusin 1 (Mfn1) mitochondrial energy metabolism, hFis1-overexpressing cells reduced the ATP content at basal
Because of extremely low lactate dehydrogenase levels, more than 90% of glucose-derived carbons are funneled into the ␤-cell mitochondria [18, 19]. mtDNA-depleted ␤-cell lines showed complete absence of glucose-stimulated insulin secretion, illustrating the importance of the organelle in signal gen
Changes in fluorescence ratios from basal levels of hFis1, DN-mitofusin 1 (Mfn1), or Drp1 caused mitochondrial fragmentawere normalized to ratio differences between resting and maximal tion in a large fraction of transfected cells (Fig. 2A, supplemendepolarization induced by 10 ␮M FCCP, and expressed as the per- tal Fig. S2, A and B)

Summary

Introduction

Overexpression of WT-Mfn1 mitochondrial energy metabolism, hFis1-overexpressing cells reduced the ATP content at basal We tested whether hFis1 overexpression alters the mitochondrial calcium response to glucose or KCl. In noninduced control cells, glucose rapidly increased the average mitochondrial calcium concentration ([Ca2ϩ]mito) from 195 Ϯ 8 nM to a peak amplitude of 1062 Ϯ 75 nM, which thereafter gradually returned to resting levels (Fig. 5, D and E). The glucose-mediated area under the curve increase of [Ca2ϩ]mito was reduced by 33% in hFis1-overexpressing cells (p Ͻ 0.01; Fig. 5F).

Results

Conclusion