The mechanism of nucleophagy in rejuvenating the nuclear envelope

PROJECT SUMMARY Over the last 10 years, the lysosome-mediated degradation pathway macroautophagy has gained prominence in the study of aging-related disorders and extension of lifespan. Macroautophagy is an essential cellular pathway responsible for the elimination of cytosolic proteins, lipids, and organelles, and as such, the field has focused upon the role of macroautophagy in clearing protein aggregates or dysfunctional organelles (such as mitochondria) that specifically accumulate in the cytoplasm. Increasingly however, protein and toxic lipid accumulation and organelle dysfunction are observed to occur within the nucleus, apparently shielded from cytoplasmic processes by the double-membraned nuclear envelope. Furthermore, links between aging and nuclear envelope structural defects are emerging, including nuclear envelopathies caused by mutations in the envelope scaffolding lamins and associated integral inner nuclear membrane proteins. Recent work has also established that the nuclear envelope is a hotspot for lipid metabolism, raising the specter that the buildup of toxic lipid species may impact nuclear envelope function if they are not effectively cleared. There is compelling evidence that components of the nuclear envelope and the nucleus are subject to macroautophagy-dependent turnover (termed nucleophagy) but the molecular mechanisms remain unknown. Here, we leverage the S. cerevisiae model to define the molecular and ultrastructural steps that define a model nucleophagy pathway. In our published and preliminary data, we establish a kinetic and ultrastructural timeline of nucleophagy. We will use these data to integrate molecular machineries that we have already discovered into this timeline to understand the mechanisms that underpin each step. These experiments will be supported by in vitro reconstitution to provide a deep insight into how these factors remodel the nuclear membranes. Lastly, we test the innovative hypothesis that nucleophagy clears toxic lipid species in order to rejuvenate the nuclear membranes and ensure robust chronological lifespan. The latter will employ leading-edge lipidomics platforms to provide much needed insight into how the nuclear envelope lipidome is altered during aging.