Imeglimin mitigates the accumulation of dysfunctional mitochondria to restore insulin secretion and suppress apoptosis of pancreatic ß-cells from db/db mice.

Mitochondrial dysfunction in pancreatic ß-cells leads to impaired glucose-stimulated insulin secretion (GSIS) and type 2 diabetes (T2D), highlighting the importance of autophagic elimination of dysfunctional mitochondria (mitophagy) in mitochondrial quality control (mQC). Imeglimin, a new oral anti-diabetic drug that improves hyperglycemia and GSIS, may enhance mitochondrial activity. However, chronic imeglimin treatment's effects on mQC in diabetic ß-cells are unknown. Here, we compared imeglimin, structurally similar anti-diabetic drug metformin, and insulin for their effects on clearance of dysfunctional mitochondria through mitophagy in pancreatic ß-cells from diabetic model db/db mice and mitophagy reporter (CMMR) mice. Pancreatic islets from db/db mice showed aberrant accumulation of dysfunctional mitochondria and excessive production of reactive oxygen species (ROS) along with markedly elevated mitophagy, suggesting that the generation of dysfunctional mitochondria overwhelmed the mitophagic capacity in db/db ß-cells. Treatment with imeglimin or insulin, but not metformin, reduced ROS production and the numbers of dysfunctional mitochondria, and normalized mitophagic activity in db/db ß-cells. Concomitantly, imeglimin and insulin, but not metformin, restored the secreted insulin level and reduced ß-cell apoptosis in db/db mice. In conclusion, imeglimin mitigated accumulation of dysfunctional mitochondria through mitophagy in diabetic mice, and may contribute to preserving ß-cell function and effective glycemic control in T2D.

Atg44/Mdi1/mitofissin facilitates Dnm1-mediated mitochondrial fission.

Mitochondria undergo fission and fusion, and their coordinated balance is crucial for maintaining mitochondrial homeostasis. In yeast, the dynamin-related protein Dnm1 is a mitochondrial fission factor acting from outside the mitochondria. We recently reported the mitochondrial intermembrane space protein Atg44/mitofissin/Mdi1/Mco8 as a novel fission factor, but the relationship between Atg44 and Dnm1 remains elusive. Here, we show that Atg44 is required to complete Dnm1-mediated mitochondrial fission under homeostatic conditions. Atg44-deficient cells often exhibit enlarged mitochondria with accumulated Dnm1 and rosary-like mitochondria with Dnm1 foci at constriction sites. These mitochondrial constriction sites retain the continuity of both the outer and inner membranes within an extremely confined space, indicating that Dnm1 is unable to complete mitochondrial fission without Atg44. Moreover, accumulated Atg44 proteins are observed at mitochondrial constriction sites. These findings suggest that Atg44 and Dnm1 cooperatively execute mitochondrial fission from inside and outside the mitochondria, respectively.Abbreviation: ATG: autophagy related; CLEM: correlative light and electron microscopy; EM: electron microscopy; ER: endoplasmic reticulum; ERMES: endoplasmic reticulum-mitochondria encounter structure; GA: glutaraldehyde; GFP: green fluorescent protein; GTP: guanosine triphosphate: IMM: inner mitochondrial membrane; IMS: intermembrane space; OMM: outer mitochondrial membrane; PB: phosphate buffer; PBS: phosphate-buffered saline; PFA: paraformaldehyde; RFP: red fluorescent protein; WT: wild type.

Mitophagy mediated by BNIP3 and NIX protects against ferroptosis by downregulating mitochondrial reactive oxygen species.

Mitophagy plays an important role in the maintenance of mitochondrial homeostasis and can be categorized into two types: ubiquitin-mediated and receptor-mediated pathways. During receptor-mediated mitophagy, mitophagy receptors facilitate mitophagy by tethering the isolation membrane to mitochondria. Although at least five outer mitochondrial membrane proteins have been identified as mitophagy receptors, their individual contribution and interrelationship remain unclear. Here, we show that HeLa cells lacking BNIP3 and NIX, two of the five receptors, exhibit a complete loss of mitophagy in various conditions. Conversely, cells deficient in the other three receptors show normal mitophagy. Using BNIP3/NIX double knockout (DKO) cells as a model, we reveal that mitophagy deficiency elevates mitochondrial reactive oxygen species (mtROS), which leads to activation of the Nrf2 antioxidant pathway. Notably, BNIP3/NIX DKO cells are highly sensitive to ferroptosis when Nrf2-driven antioxidant enzymes are compromised. Moreover, the sensitivity of BNIP3/NIX DKO cells is fully rescued upon the introduction of wild-type BNIP3 and NIX, but not the mutant forms incapable of facilitating mitophagy. Consequently, our results demonstrate that BNIP3 and NIX-mediated mitophagy plays a role in regulating mtROS levels and protects cells from ferroptosis.

Comprehensive analysis of non-selective and selective autophagy in yeast atg mutants and characterization of autophagic activity in the absence of the Atg8 conjugation system.

Most autophagy-related genes, or ATG genes, have been identified in studies using budding yeast. Although the functions of the ATG genes are well understood, the contributions of individual genes to non-selective and various types of selective autophagy remain to be fully elucidated. In this study, we quantified the activity of non-selective autophagy, the cytoplasm-to-vacuole targeting (Cvt) pathway, mitophagy, endoplasmic reticulum (ER)-phagy, and pexophagy in all Saccharomyces cerevisiae atg mutants. Among the mutants of the core autophagy genes considered essential for autophagy, the atg13 mutant and mutants of the genes involved in the two ubiquitin-like conjugation systems retained residual autophagic functionality. In particular, mutants of the Atg8 ubiquitin-like conjugation system (the Atg8 system) exhibited substantial levels of non-selective autophagy, the Cvt pathway, and pexophagy, although mitophagy and ER-phagy were undetectable. Atg8-system mutants also displayed intravacuolar vesicles resembling autophagic bodies, albeit at significantly reduced size and frequency. Thus, our data suggest that membranous sequestration and vacuolar delivery of autophagic cargo can occur in the absence of the Atg8 system. Alongside these findings, the comprehensive analysis conducted here provides valuable datasets for future autophagy research.