Project: Proteomic analysis of the autophagosome

No organelle is defined by just one protein, but thus far, autophagosomes come close. Beyond the lipidated form of Atg8 (or LC3 in mammals) there are no known permanently resident proteins of the autophagic membrane. Despite this simplicity, autophagosomes move about the cell, engage common intracellular trafficking pathways and eventually fuse into the lysosome to deliver their cargo for degradation. Which proteins accomplish these myriad functions are uncertain.

We have updated classic organelle isolation approaches first devised by Seglen and colleagues to isolate autophagosomes and determine their proteomic character. From these analyses we have identified several potentially interesting candidate proteins involved in autophagosome trafficking within the cell. Cell biology and microscopy assays establish whether these candidate proteins are involved in constitutive or simulated autophagic processes and whether they influence the clearance of cellular toxins like the aggregated form of huntington protein. Our long term goal is to determine whether membrane directed functions of autophagy rely upon novel protein machineries or the repurposing of existing protein paradigms and then use this information to manipulate the rate and extent of autophagy-dependent toxin clearance.