The role of mitochondrial dynamics and autophagy in pancreatic beta-cell response to nutrient stress

Mitochondrial dynamics includes the processes of fusion, fission, and motility. These processes form interdependent adaptive mechanisms that, together with autophagy, maintain mitochondrial function to meet cellular needs. Mitochondrial dynamics control function directly by inducing bioenergetic remodeling or indirectly by promoting turnover of mitochondria via autophagy. Importantly, mitochondrial dysfunction has been implicated in beta-cell failure during type 2 diabetes. This thesis will investigate the role of dynamics and autophagy in regulating mitochondrial and pancreatic beta-cell function during chronic exposure to excess glucose and fatty acids, termed glucolipotoxicity (GLT). It remains ill-defined what role fusion and motility play in determining mitochondrial turnover, as current methodologies to assess turnover lack subcellular resolution. To address this need we developed the use of MitoTimer, a mitochondrial fluorescent probe that undergoes a time-dependent green-to-red transition. Turnover was revealed by the integrated proportions of young (green) and old (red) MitoTimer protein. The results demonstrate that mitochondrial fusion and motility regulate turnover by promoting the distribution of newer protein to subsets of mitochondria in the network. GLT inhibits mitochondrial fusion and networking in pancreatic beta-cells. Since fusion is dependent on motility we tested the hypothesis that GLT impairs fusion by affecting motility. We determined that GLT arrests motility, which may contribute to mitochondrial and beta-cell dysfunction. We show that excess nutrients increase O-linked β-N-acetyl glucosamine (O-GlcNAc) modification of mitochondrial motor adaptor Milton1, which decreases its activity and results in arrest of motility and increased fission. Thus Milton1 O-GlcNAc modification acts as a nutrient-sensor linking fusion, fission, and motility to nutrient supply in the beta-cell. Finally, GLT inhibits autophagic flux with concurrent lysosomal pH increase in beta-cells. To address the hypothesis that impaired lysosomal acidification is a causative event inhibiting autophagic flux and beta-cell function, we developed lysosome-localizing nanoparticles that expand and acidify upon UV photo-activation. Increasing lysosomal acidity with the nanoparticles increased autophagic flux and restored beta-cell function under GLT, establishing lysosomal pH as a key mediator of nutrient-induced beta-cell dysfunction. In summary the work elucidates the interdependence and specific roles of mitochondrial fusion, fission, motility, and autophagy in dictating beta-cell responses to excess nutrient environment.2017-06-15T00:00:00

Lysosomal dysfunction and impaired autophagy underlie the pathogenesis of amyloidogenic light chain-mediated cardiotoxicity

AL amyloidosis is the consequence of clonal production of amyloidogenic immunoglobulin light chain (LC) proteins, often resulting in a rapidly progressive and fatal amyloid cardiomyopathy. Recent work has found that amyloidogenic LC directly initiate a cardio-toxic response underlying the pathogenesis of the cardiomyopathy; however, the mechanisms that contribute to this proteotoxicity remain unknown. Using human amyloidogenic LC isolated from patients with amyloid cardiomyopathy, we reveal that dysregulation of autophagic flux is critical for mediating amyloidogenic LC proteotoxicity. Restoration of autophagic flux by pharmacological intervention using rapamycin protected against amyloidogenic light chain protein-induced pathologies including contractile dysfunction and cell death at the cellular and organ level and also prolonged survival in an in vivo zebrafish model of amyloid cardiotoxicity. Mechanistically, we identify impaired lysosomal function to be the major cause of defective autophagy and amyloidogenic LC-induced proteotoxicity. Collectively, these findings detail the downstream molecular mechanisms underlying AL amyloid cardiomyopathy and highlight potential targeting of autophagy and lysosomal dysfunction in patients with amyloid cardiomyopathy

The US Program in Ground-Based Gravitational Wave Science: Contribution from the LIGO Laboratory

Recent gravitational-wave observations from the LIGO and Virgo observatories have brought a sense of great excitement to scientists and citizens the world over. Since September 2015,10 binary black hole coalescences and one binary neutron star coalescence have been observed. They have provided remarkable, revolutionary insight into the "gravitational Universe" and have greatly extended the field of multi-messenger astronomy. At present, Advanced LIGO can see binary black hole coalescences out to redshift 0.6 and binary neutron star coalescences to redshift 0.05. This probes only a very small fraction of the volume of the observable Universe. However, current technologies can be extended to construct "3rd Generation" (3G) gravitational-wave observatories that would extend our reach to the very edge of the observable Universe. The event rates over such a large volume would be in the hundreds of thousands per year (i.e. tens per hour). Such 3G detectors would have a 10-fold improvement in strain sensitivity over the current generation of instruments, yielding signal-to-noise ratios of 1000 for events like those already seen. Several concepts are being studied for which engineering studies and reliable cost estimates will be developed in the next 5 years

Modulation of mTOR signaling as a strategy for the treatment of Pompe disease

Abstract Mechanistic target of rapamycin (mTOR) coordinates biosynthetic and catabolic processes in response to multiple extracellular and intracellular signals including growth factors and nutrients. This serine/threonine kinase has long been known as a critical regulator of muscle mass. The recent finding that the decision regarding its activation/inactivation takes place at the lysosome undeniably brings mTOR into the field of lysosomal storage diseases. In this study, we have examined the involvement of the mTOR pathway in the pathophysiology of a severe muscle wasting condition, Pompe disease, caused by excessive accumulation of lysosomal glycogen. Here, we report the dysregulation of mTOR signaling in the diseased muscle cells, and we focus on potential sites for therapeutic intervention. Reactivation of mTOR in the whole muscle of Pompe mice by TSC knockdown resulted in the reversal of atrophy and a striking removal of autophagic buildup. Of particular interest, we found that the aberrant mTOR signaling can be reversed by arginine. This finding can be translated into the clinic and may become a paradigm for targeted therapy in lysosomal, metabolic, and neuromuscular diseases

Lysosome acidification by photoactivated nanoparticles restores autophagy under lipotoxicity

In pancreatic β-cells, liver hepatocytes, and cardiomyocytes, chronic exposure to high levels of fatty acids (lipotoxicity) inhibits autophagic flux and concomitantly decreases lysosomal acidity. Whether impaired lysosomal acidification is causally inhibiting autophagic flux and cellular functions could not, up to the present, be determined because of the lack of an approach to modify lysosomal acidity. To address this question, lysosome-localizing nanoparticles are described that, upon UV photoactivation, enable controlled acidification of impaired lysosomes. The photoactivatable, acidifying nanoparticles (paNPs) demonstrate lysosomal uptake in INS1 and mouse β-cells. Photoactivation of paNPs in fatty acid–treated INS1 cells enhances lysosomal acidity and function while decreasing p62 and LC3-II levels, indicating rescue of autophagic flux upon acute lysosomal acidification. Furthermore, paNPs improve glucose-stimulated insulin secretion that is reduced under lipotoxicity in INS1 cells and mouse islets. These results establish a causative role for impaired lysosomal acidification in the deregulation of autophagy and β-cell function under lipotoxicity