CHSU Discovery

Abstract:
The mitochondria are dynamic, vital, maternally-inherited organelles with their own genome
(mtDNA) that function as the powerhouse of the cell and as a signaling hub regulating various physiological processes including programmed cell death (PCD). Errors during replication of mtDNA accumulate over time leading to a multitude of diseases affecting up to 1 in 4300 people. Mitochondrial dysfunction is also associated with age-related dementia, myopathies, and heart diseases. Mitochondrial purifying selection (MPS) is the removal of damaged mitochondria in the germline that can occur at organismal, cellular, organellar, and molecular levels. We wanted to test the hypothesis that damaged mtDNA and mitochondria are eliminated by programmed cell death in maternal germlines. We assayed germ cell death using transgenic fluorescent markers and vital dyes in different PCD mutant backgrounds in adult C. elegans. Caspase activity was monitored using a genetically engineered biosensor with differentially localizing membrane RFP and nuclear GFP signals upon caspase activation. Copy number of wildtype and mutant mtDNA was quantified by digital droplet PCR. Presence of mutant mtDNA,uaDf5, led to a modest increase in germ cell death. This extra germ cell death required the activity of caspase activator APAF-
1/CED-4 and the caspases CED-3 and CSP-1. Unexpectedly, the cell death inhibitor CED-9/BCL2, which does not play a role in normal germ cell death, blocked the extra cell death in uaDf5 mutants suggesting that uaDf5 dependent cell death occurs via a non-canonical CED-9 dependent pathway. Abrogation of caspases resulted in an increase in mutant mtDNA levels. Contrary to our expectations, in APAF1/CED-4 loss of function mutants there was no significant increase in uaDf5 copy number even in the absence of all cell death suggesting a CED-4 independent, non-apoptotic role for caspases in MPS. Collectively, our findings suggest the role of caspases in eliminating damaged mitochondria through both apoptotic as well as non-apoptotic processes and suggests future avenues of investigation. Understanding the mechanism for MPS will allow for development of therapies for currently incurable mitochondrial and aging associated diseases leading to the possibility of extending human lifespan and healthspan.