Open Access Dissertation
Date of Award
Miriam L. Greenberg
Cardiolipin (CL), the signature phospholipid of mitochondrial membranes, is important for cardiovascular health. Perturbation of CL metabolism is implicated in cardiovascular disease (CVD). The link between CL and CVD may be explained by the physiological roles of CL in pathways that are cardioprotective, such as autophagy/mitophagy and the mitogen-activated protein kinase (MAPK) pathways. My dissertation work focuses on elucidating how CL influences mitophagy and MAPK pathways.
crd1Δ was synthetically lethal/sick with the general autophagy mutants atg8Δ, atg18Δ and mitophagy mutant atg32Δ, suggesting that autophagy/mitophagy may be deficient in cells lacking CL. Microscopic examination of mitophagy revealed decreased translocation of GFP-tagged mitochondrial proteins into the vacuole of crd1Δ cells. This was confirmed by a decreased level of free GFP generated by cleavage of GFP-tagged mitochondrial protein after delivery into the vacuole by mitophagy. These findings indicated that mitophagy is decreased in CL-deficient cells. Expression of ATG8 was increased in crd1Δ cells at 37˚C, suggesting that nonselective autophagy was upregulated to compensate for decreased mitophagy.
The PKC and HOG MAPK pathways are known to be required for mitophagy. crd1Δ growth defects are exacerbated by deletion of HOG pathway genes SHO1, SSK1, STE50 and HOG1, and rescued by stimulating the HOG pathway and upregulating the PKC pathway. These findings suggested the possibility that MAPK pathways are defective in crd1Δ cells. Phosphorylation of Slt2p and Hog1p in response to stimulants was decreased in crd1Δ, consistent with defective activation of these MAPK pathways. Interestingly, upregulating PKC by transforming the cell with a vector expressing a constitutively activated Pkc1p rescued defective mitophagy in crd1Δ.
These results suggest that the mechanism underlying defective mitophagy caused by loss of CL is a defective PKC pathway.
Shen, Zheni, "Cardiolipin Regulates Mitophagy Through The Pkc Pathway" (2015). Wayne State University Dissertations. 1319.