MitofusinsMfn1andMfn2Are Required to Preserve Glucose- but Not Incretin-Stimulated β-Cell Connectivity and Insulin Secretion

Abstract

Mitochondrial glucose metabolism is essential for stimulated insulin release from pancreatic beta cells. Whether mitofusin gene expression, and hence mitochondrial network integrity, is important for glucose or incretin signalling has not previously been explored. Here, we generated mice with beta cell-selective, adult-restricted deletion of the mitofusin genes Mfn1 and Mfn2 (βMfn1/2 dKO). βMfn1/2 dKO mice displayed elevated fed and fasted glycaemia and a >five-fold decrease in plasma insulin. Mitochondrial length, glucose-induced polarisation, ATP synthesis, cytosolic and mitochondrial Ca2+ increases were all reduced in dKO islets. In contrast, oral glucose tolerance was more modestly affected in βMfn1/2 dKO mice and GLP-1 or GIP receptor agonists largely corrected defective GSIS through enhanced EPAC-dependent signalling. Correspondingly, cAMP increases in the cytosol, as measured with an Epac-camps based sensor, were exaggerated in dKO mice. Mitochondrial fusion and fission cycles are thus essential in the beta cell to maintain normal glucose, but not incretin, sensing. These findings broaden our understanding of the roles of mitofusins in beta cells, the potential contributions of altered mitochondrial dynamics to diabetes development and the impact of incretins on this process.

Molecular phenotyping of single pancreatic islet leader beta cells by “Flash-Seq”

“…Nevertheless, we cannot exclude the possibility that there are actions of the calmodulin moiety present in GCaMP6 on calcium-regulated proteins and processes in GCaMP6-expressing cells. Consistent with the concentration of calcium-binding sites being a key determinant in differing calcium dynamics, our preliminary data (not shown), as well as those of others 34 , using the bright Cal520 probe demonstrate a greater propensity towards wave-like, and less small-worlds behaviour. Importantly, we observed almost no overlap between leader cells and highly connected hub cells.…”

Retrograde mitochondrial signaling governs the identity and maturity of metabolic tissues

“…Reduced expression of core β-cell maturity markers and increased markers of immaturity were also apparent in islets from mice bearing β-cell-specific deletion of Mitofusins 1 and 2, a recently described model of impaired mitochondrial quality control that abrogates mitochondrial fusion (β-Mfn1/2 DKO mice; Figures S4D-E; 31 ). Notably, loss of Mfn1/2 also leads to reductions in β-cell mass with age 51 . Stem cell or early endocrine progenitor cell markers such as Oct4, Sox2, Nanog , and Ngn3 were not upregulated in islets from β-Clec16a KO , β-Tfam KO , or β-Mfn1/2 DKO mice (data not shown), indicating that loss of mitochondrial quality control induces markers of immaturity and loss of terminal β-cell identity without a signature associated with regression to early developmental precursors.…”

“…Reduced expression of core β-cell maturity markers and increased markers of immaturity were also apparent in islets from mice bearing β-cell-specific deletion of Mitofusins 1 and 2, a recently described model of impaired mitochondrial quality control that abrogates mitochondrial fusion (β-Mfn1/2 DKO mice; Figures S4D-E; 31 ). Notably, loss of Mfn1/2 also leads to reductions in β-cell mass with age 51 .…”

Differential CpG methylation atNnatin the early establishment of beta cell heterogeneity

“…Mice with global deletion of Nnat were used as previously described (67). Conditional Dnmt3a null mice (74), generated by crossing to Pdx1-Cre (75) have been described elsewhere (13, 15, 76). Mice containing the Dnmt3a floxed allele but lacking Pdx1-Cre were used as controls.…”

“…Signal binarisation-based connectivity and correlation analyses were performed as previously described (36). Connectivity between cells was explored using Pearson’s R-based correlation analyses in MATLAB as described (76). We note that although the Ca 2+ dye used here was not selectively targeted towards beta cells, at the concentration of glucose used (11 mM), pancreatic alpha cells are expected to be largely inactive.…”