Macroautophagy (hereafter autophagy) is an umbrella term for vesicle transport pathways that target cytoplasmic components to the lysosome for destruction. There are two classes of autophagy: nonselective and selective. During nonselective autophagy, bulk cytoplasm gets packaged into specialized vesicles, called autophagosomes, which fuse with the lysosome to degrade their encapsulated contents. By contrast, during selective autophagy, specific toxic structures ? such as ubiquitinated protein aggregates ? are recognized by autophagy receptors, which also interact with the nascent autophagosome membrane to enable vesicle engulfment of the receptor-target complex. In a recently published work, we discovered that autophagy receptors also have an unanticipated regulatory role as activators of the dedicated kinase that controls initiation of autophagosome formation. This work fits nicely with other studies showing that receptor activity is not constitutive but rather directly controlled by general kinases. To gain further insights into receptor function, we have hypothesized that receptors enable transduction of selective autophagy signals down kinase cascades ? from general kinases to the dedicated autophagy kinase ? to make decisions about the destruction of selective autophagy targets ? which specific target should be destroyed and when? This proposal uses selective autophagy of peroxisomes (pexophagy) to build a case study for the kinase signal cascade hypothesis. The bulk of the proposed work will be performed using budding yeast because this model organism has facile genetics, is tractable biochemically, and accessible cell biologically to imaging by fluorescence microscopy. The remainder of the work comprises pioneering genetic screens in mammalian cells for pexophagy mutants. Our strong track record in doing follow- up mechanistic work on the GET pathway components, which were identified originally by yeast high- throughput screens, will help guide us from screen hits towards new mammalian pexophagy components and mechanistic insights that are missing from the current picture. This latter work also has direct medical relevance because in certain Zellweger Spectrum patients, peroxisomes can form normally, but are degraded by autophagy.
This project addresses how cells make decisions that culminate in the destruction of large toxic structures in the cytoplasm, such as damaged organelles and protein aggregates. Defects in this decision-making process are associated with many human diseases, including neurodegeneration. This grant seeks to uncover the nuts and bolts of this decision-making process to facilitate discovery of better drug targets that will improve human health.
| Kamber, Roarke A; Shoemaker, Christopher J; Denic, Vladimir (2015) A molecular switch for selective autophagosome formation. Autophagy 11:2132-2133 |
| Kamber, Roarke A; Shoemaker, Christopher J; Denic, Vladimir (2015) Receptor-Bound Targets of Selective Autophagy Use a Scaffold Protein to Activate the Atg1 Kinase. Mol Cell 59:372-81 |
