Mitochondrial dysfunction induces aberrant insulin signalling and glucose utilisation in murine C2C12 myotube cells

Mitochondrial dysfunction is considered a critical component in the development of diabetes. The aim of this study was to elucidate the molecular mechanisms involved in the development of insulin resistance and diabetes through investigation of mitochondrial retrograde signalling. Mitochondrial function of C2C12 myotube cells was impaired by genetic (ethidium bromide) and metabolic (oligomycin) stress, and changes in target molecules related to insulin signalling were analysed. Concomitant with reductions in mitochondrial membrane potential (DeltaPsim) and ATP synthesis, production of IRS1 and solute carrier family 2 (facilitated glucose transporter), member 4 (SLC2A4, formerly known as GLUT4) were reduced. Moreover, serine phosphorylation of IRS1 increased, resulting in decreased tyrosine phosphorylation. This indicates that mitochondrial dysfunction decreases insulin-stimulated SLC2A4 translocation and glucose uptake. Mitochondrial stress activated c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase (p38 MAPK) signalling in a Ca(2+)-dependent manner, and removal of free Ca(2+) by BAPTA-AM, as well as inhibition of JNK and p38 MAPK, abrogated mitochondrial stress-induced reductions in IRS1 and SLC2A4 production. Mitochondrial dysfunction after oligomycin treatment significantly increased levels of activating transcription factor 3 (ATF3), which represses Irs1 promoter activity. Removal of the 5' flanking region of Irs1 demonstrated that the promoter region within 191 bases from the transcription site may be involved in the transcriptional repression of Irs1 by mitochondrial stress. We show distinct mitochondrial retrograde signalling, where Irs1 is downregulated through ATF3 in a Ca(2+)-, JNK- and p38 MAPK-dependent manner, and IRS1 is inactivated. Therefore, mitochondrial dysfunction causes aberrant insulin signalling and abnormal utilisation of glucose, as observed in many insulin resistance states.