Candidate for a PhD in Molecular, Cellular, and Developmental Biology

Abstract: Macroautophagy (hereafter autophagy) is a cellular recycling process through which cytoplasmic contents are delivered into the lysosome/vacuole for degradation by double-membrane organelles, autophagosomes. Autophagy is essential for cell survival under stress; however, too much autophagy can also be detrimental. Autophagy activity can be regulated by modulating the size or the number of autophagosomes. Although there are more than 40 autophagy-related (ATG) genes that have been identified, it is not fully understood how most of these genes contribute to these aspects of regulation. Autophagy is highly conserved among eukaryotic cells, and its molecular machinery has been best characterized in the budding yeast Saccharomyces cerevisiae.

In my thesis studies, I use budding yeast as the model organism, taking advantage of its utility in genetic/genomic screening; in addition, high-throughput sequencing and powerful autophagy assays have been developed uniquely in the yeast system, to explore how autophagy is modulated through transcriptional regulation.
When I joined the Klionsky lab, I became involved in the study of a negative transcriptional regulator of autophagy, Ume6. Deletion of the UME6 gene results in an increase in the size, but not the number, of autophagosomes by increasing the expression of Atg8. From a subsequent genetic screen for autophagy modulators, I identified another transcription repressor of autophagy, Pho23. Intriguingly, Pho23 ended up being characterized as a specific regulator of the number, but not the size, of autophagosomes, or it can be viewed as controlling the rate of autophagosome formation by regulating Atg9 expression. These studies support a model whereby the size and numbers of autophagosomes are independently regulated through precise transcriptional regulation of different ATG genes.

Collaborating with Amélie Bernard, a postdoc in the lab, to further explore the transcriptional regulation network of autophagy, we analyzed 139 yeast strains each deleted for a single gene encoding a transcription factor; we profiled the transcription of several ATG genes in each strain. Through this screen we identified Gcn4, Slf1, Gat1 and Gln3 as transcriptional activators, and Spt10, Fyv5, and Rph1 as transcriptional repressors of autophagy. We also further investigated the detailed molecular mechanisms of the regulation of autophagy by Rph1.