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 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. How the cell responds to these nuclear insults is not well understood, but intriguingly, lamins appear to be subject to macroautophagy-dependent turnover. Thus, it is clear that we must refocus our attention on how nuclear quality control is executed and specifically on the mechanism(s) that governs nuclear content turnover in cytoplasmic autophagosomes. Thus, in this proposal, we focus on the fundamental question of how cytoplasmic autophagy machinery and nuclear envelope remodeling are coordinated. Using S. cerevisiae, where discovery of the molecular machinery driving nuclear autophagy is the most mature, we will reveal each of the complex membrane dynamics events that occur to move nuclear envelope fragments away from the nucleus and eventually into the vacuole for degradation. We will then establish the mechanism of membrane remodeling, using fully reconstituted systems that maintain the topologic identity of each of the two nuclear envelope membranes. Importantly, within these reconstituted systems we will also introduce molecular mimics of the growing autophagosome and recapitulate the formation of the nuclear envelope-autophagosome interface that governs sequestration of nuclear envelope fragments. With the completion of this project, we will further our mechanistic understanding of a key underappreciated macroautophagic process and further our understanding of how nuclear autophagy can impact aging-impaired proteostasis.
Aging related disorders are strongly associated with a reduced capacity to induce the autophagy stress response and increasingly are also associated with the accumulation of structural defects along the nuclear membranes. Neither the physiologic overlaps between these two phenotypes, nor the mechanistic underpinnings of autophagy-dependent nuclear envelope remodeling are well characterized. To identify how autophagy can help limit age-related nuclear envelope dysfunction, our proposal focuses on describing the structural and mechanistic events that drive nuclear autophagy.
