All ETDs from UAB

Advisory Committee Chair

David Bedwell

Advisory Committee Members

Kim Keeling

David Schneider

James Collawn

John Shacka

Document Type

Dissertation

Date of Award

2017

Degree Name by School

Doctor of Philosophy (PhD) Heersink School of Medicine

Abstract

The process of bulk degradation of cytoplasmic contents, called macroautophagy or simply autophagy, is a heavily conserved cellular process from yeast to humans. In yeast, it involves more than 30 different autophagy related genes (ATG) that coordinate the process of building a double membrane vesicle, called the autophagosome, that fuses with the vacuole to allow degradation and recycling of its contents. This process can also be selective, involving the recruitment of the autophagic machinery to specific organelles. In this work, we used RNA sequencing (RNA-seq) to identify a subset of ATG genes that are heavily upregulated in response to nitrogen starvation, a potent inducer of autophagy. This subset of genes includes several ATG genes that are essential to autophagy and play critical roles in the early formation of the autophagosome which suggests there may be one or more mechanisms to rapidly increase the abundance of these ATG mRNAs upon the induction of autophagy. We examined the transcription and mRNA decay of two of these genes, ATG1 and ATG8, which showed a 4 and 8-fold increase in steady-state abundance respectively. We also investigated the transcription and degradation of the ATG27 mRNA, a gene that showed little or no change during nitrogen starvation, and a housekeeping gene, ACT1. We found that while transcription was upregulated 2 to 3-fold for ATG8, ATG1 transcription did not increase at all. Interestingly, we found a major increase in mRNA half-life of these two genes during nitrogen starvation, suggesting that mRNA stability may be altered in order to increase ATG abundance in response to nitrogen starvation. Analysis of both 5’-3’ and 3’-5’ mRNA decay mutants demonstrated that this increase in steady-state abundance was mediated by alterations in the 5’-3’ decay pathway only. When we disrupted this pathway by creating a dcp2Δ decapping mutant the increase in mRNA half-life was largely abrogated during nitrogen starvation. Further, the dcp2Δ mutant showed an upregulation of autophagy in nutrient-rich conditions which supports the hypothesis that the 5’-3’ mRNA decay pathway regulates autophagy.

Share

COinS