Biomedical Sciences ETDs

Publication Date

5-1-2015

Abstract

Caloric restriction (CR) increases lifespan and promotes both longevity and €˜healthy-aging. CR's benefits are dependent in part on NAD-dependent deacetylases known as sirtuins. NAD levels increase with CR promoting sirtuin-mediated deacetylation of lysine residues. Humans express seven sirtuin isoforms that localize to specific sub-cellular compartments. Sirtuins and acetylation affect numerous cellular processes including mitochondrial function. Sirt3 is the major mitochondrial deacetylase and regulates mitochondrial function namely ATP levels, oxidative stress and metabolism. Given the importance of mitochondrial acetylation and Sirt3, we hypothesized the existence of a mitochondrial-localized acetyltransferase that 1) localizes to the mitochondria; 2) has an acetyltransferase domain; and 3) counters known effects of Sirt3. We found that the protein GCN5L1 localizes to mitochondria, and its depletion or overexpression attenuates and augments mitochondrial protein acetylation, respectively. Sirt3 increases mitochondrial respiration and ATP production. Acute GCN5L1 knockdown also increases mitochondrial respiration and ATP production suggesting unopposed Sirt3 activity. GCN5L1 promotes the acetylation of the electron transport chain proteins NDUFA9 and ATP5α (known Sirt3 targets). In addition, Sirt3 knockdown leads to mitochondrial hyperacetylation, which is countered by concurrent GCN5L1 depletion. We propose that GCN5L1 is an essential component of a mitochondrial acetyltransferase complex. As acetylation levels affect the housekeeping program known as autophagy, we interrogated GCN5L1's role in this program. GCN5L1 knockdown causes autophagic proteins to accumulate on mitochondria, which can be reversed with concurrent Sirt3 depletion. This knockdown selectively promotes the autophagic clearance of mitochondria (mitophagy) without altering global autophagy. Chronic GCN5L1 depletion leads to the diminution of mitochondrial proteins/mass that is dependent on the autophagy proteins ATG5 and p62 but independent of the E3-ligase Parkin. This program is elevated in GCN5L1 knockout MEFs, which is reversed with reconstitution of GCN5L1. GCN5L1 depleted mitochondria generate lower ROS levels and are resistant to MPTP opening. Both mitophagy and mitochondrial biogenesis (i.e. mitochondrial turnover) are simultaneously elevated with GCN5L1 knockdown. Therefore, GCN5L1 functions as a critical component of the mitochondrial acetyltransferase machinery and regulates mitochondrial acetylation-dependent mitophagy/mitochondrial biogenesis. GCN5L1 may play a role in numerous pathologies with known mitochondrial defects such as neuro-degeneration, obesity and aging.

Keywords

aging, mitochondria, acetylation, Sirtuin, GCN5L1, mitophagy, autophagy

Sponsors

National Institutes of Health (NIH) National Heart Lung and Blood Institute (NHLBI)

Document Type

Dissertation

Language

English

Degree Name

Biomedical Sciences

Level of Degree

Doctoral

Department Name

Biomedical Sciences Graduate Program

First Advisor

Kanagy, Nancy

First Committee Member (Chair)

Sack, Michael

Second Committee Member

Parra, Karlett

Third Committee Member

Gonzalez-Bosc, Laura

Fourth Committee Member

Finkel, Toren

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