Reciprocal regulatory balance within the CLEC16A-RNF41 mitophagy complex depends on an intrinsically disordered protein region.

CLEC16A is an E3 ubiquitin ligase that regulates mitochondrial quality control through mitophagy and is associated with over 20 human diseases. CLEC16A forms a complex with another E3 ligase, RNF41, and a ubiquitin-specific peptidase, USP8; however, regions that regulate CLEC16A activity or the assembly of the tripartite mitophagy regulatory complex are unknown. Here, we report that CLEC16A contains an internal intrinsically disordered protein region (IDPR) that is crucial for CLEC16A function and turnover. IDPRs lack a fixed secondary structure and possess emerging yet still equivocal roles in protein stability, interactions, and enzymatic activity. We find that the internal IDPR of CLEC16A is crucial for its degradation. CLEC16A turnover was promoted by RNF41, which binds and acts upon the internal IDPR to destabilize CLEC16A. Loss of this internal IDPR also destabilized the ubiquitin-dependent tripartite CLEC16A-RNF41-USP8 complex. Finally, the presence of an internal IDPR within CLEC16A was confirmed using NMR and CD spectroscopy. Together, our studies reveal that an IDPR is essential to control the reciprocal regulatory balance between CLEC16A and RNF41, which could be targeted to improve mitochondrial health in disease.

An intrinsically disordered protein region encoded by the human disease gene CLEC16A regulates mitophagy.

CLEC16A regulates mitochondrial health through mitophagy and is associated with over 20 human diseases. However, the key structural and functional regions of CLEC16A, and their relevance for human disease, remain unknown. Here, we report that a disease-associated CLEC16A variant lacks a C-terminal intrinsically disordered protein region (IDPR) that is critical for mitochondrial quality control. IDPRs comprise nearly half of the human proteome, yet their mechanistic roles in human disease are poorly understood. Using carbon detect NMR, we find that the CLEC16A C terminus lacks secondary structure, validating the presence of an IDPR. Loss of the CLEC16A C-terminal IDPR in vivo impairs mitophagy, mitochondrial function, and glucose-stimulated insulin secretion, ultimately causing glucose intolerance. Deletion of the CLEC16A C-terminal IDPR increases CLEC16A ubiquitination and degradation, thus impairing assembly of the mitophagy regulatory machinery. Importantly, CLEC16A stability is dependent on proline bias within the C-terminal IDPR, but not amino acid sequence order or charge. Together, we elucidate how an IDPR in CLEC16A regulates mitophagy and implicate pathogenic human gene variants that disrupt IDPRs as novel contributors to diabetes and other CLEC16A-associated diseases.Abbreviations : CAS: carbon-detect amino-acid specific; IDPR: intrinsically disordered protein region; MEFs: mouse embryonic fibroblasts; NMR: nuclear magnetic resonance.

A Selective Look at Autophagy in Pancreatic ß-Cells.

Insulin-producing pancreatic ß-cells are central to glucose homeostasis, and their failure is a principal driver of diabetes development. To preserve optimal health ß-cells must withstand both intrinsic and extrinsic stressors, ranging from inflammation to increased peripheral insulin demand, in addition to maintaining insulin biosynthesis and secretory machinery. Autophagy is increasingly being appreciated as a critical ß-cell quality control system vital for glycemic control. Here we focus on the underappreciated, yet crucial, roles for selective and organelle-specific forms of autophagy as mediators of ß-cell health. We examine the unique molecular players underlying each distinct form of autophagy in ß-cells, including selective autophagy of mitochondria, insulin granules, lipid, intracellular amyloid aggregates, endoplasmic reticulum, and peroxisomes. We also describe how defects in selective autophagy pathways contribute to the development of diabetes. As all forms of autophagy are not the same, a refined view of ß-cell selective autophagy may inform new approaches to defend against the various insults leading to ß-cell failure in diabetes.

Cell Cycle Regulation of the Pdx1 Transcription Factor in Developing Pancreas and Insulin-Producing ß-Cells.

Current evidence indicates that proliferating ß-cells express lower levels of some functional cell identity genes, suggesting that proliferating cells are not optimally functional. Pdx1 is important for ß-cell specification, function, and proliferation and is mutated in monogenic forms of diabetes. However, its regulation during the cell cycle is unknown. Here we examined Pdx1 protein expression in immortalized ß-cells, maternal mouse islets during pregnancy, and mouse embryonic pancreas. We demonstrate that Pdx1 localization and protein levels are highly dynamic. In nonmitotic cells, Pdx1 is not observed in constitutive heterochromatin, nucleoli, or most areas containing repressive epigenetic marks. At prophase, Pdx1 is enriched around the chromosomes before Ki67 coating of the chromosome surface. Pdx1 uniformly localizes in the cytoplasm at prometaphase and becomes enriched around the chromosomes again at the end of cell division, before nuclear envelope formation. Cells in S phase have lower Pdx1 levels than cells at earlier cell cycle stages, and overexpression of Pdx1 in INS-1 cells prevents progression toward G2, suggesting that cell cycle-dependent regulation of Pdx1 is required for completion of mitosis. Together, we find that Pdx1 localization and protein levels are tightly regulated throughout the cell cycle. This dynamic regulation has implications for the dichotomous role of Pdx1 in ß-cell function and proliferation.

Mitophagy protects ß cells from inflammatory damage in diabetes.

Inflammatory damage contributes to ß cell failure in type 1 and 2 diabetes (T1D and T2D, respectively). Mitochondria are damaged by inflammatory signaling in ß cells, resulting in impaired bioenergetics and initiation of proapoptotic machinery. Hence, the identification of protective responses to inflammation could lead to new therapeutic targets. Here, we report that mitophagy serves as a protective response to inflammatory stress in both human and rodent ß cells. Utilizing in vivo mitophagy reporters, we observed that diabetogenic proinflammatory cytokines induced mitophagy in response to nitrosative/oxidative mitochondrial damage. Mitophagy-deficient ß cells were sensitized to inflammatory stress, leading to the accumulation of fragmented dysfunctional mitochondria, increased ß cell death, and hyperglycemia. Overexpression of CLEC16A, a T1D gene and mitophagy regulator whose expression in islets is protective against T1D, ameliorated cytokine-induced human ß cell apoptosis. Thus, mitophagy promotes ß cell survival and prevents diabetes by countering inflammatory injury. Targeting this pathway has the potential to prevent ß cell failure in diabetes and may be beneficial in other inflammatory conditions.

Clarifying the function of genes at the chromosome 16p13 locus in type 1 diabetes: CLEC16A and DEXI.

More than a decade after the discovery of a novel type 1 diabetes risk locus on chromosome 16p13, there remains complexity and controversy over the specific gene(s) that regulate diabetes pathogenesis. A new study by Nieves-Bonilla et al. shows that one of these genes, DEXI, is unlikely to contribute to type 1 diabetes pathogenesis and positions the endolysosomal E3 ubiquitin ligase CLEC16A as the primary culprit by which this gene locus influences diabetes risk.

Comprehensive review of genetic factors contributing to head and neck squamous cell carcinoma development in low-risk, nontraditional patients.

BACKGROUND: The past 2 decades have seen an increased incidence of head and neck squamous cell carcinoma (HNSCC) in a nontraditional, low-risk patient population (ie, ≤45 years of age, no substance use history), owing to a combination of human papillomavirus (HPV) infection and individual genetic variation. METHODS: Articles positing genetic variants as contributing factors in HNSCC incidence in low-risk, nontraditional patients were identified using a PubMed search, reviewed in detail, and concisely summarized herein. RESULTS: Recent data suggest that common polymorphisms in DNA repair enzymes, cell-cycle control proteins, apoptotic pathway members, and Fanconi anemia-associated genes likely modulate susceptibility to HNSCC development in low-risk, nontraditional patients. CONCLUSION: At present, there is a lack of robust, comprehensive data on genetic drivers of oncogenesis in low-risk patients and a clear need for further research on genetic alterations underlying the rising incidence of HNSCC in low-risk, nontraditional patients.