Abstract 166: MCL-1 Facilitates the Removal of Damaged Mitochondria via the Mitophagy Receptor BNIP3

Abstract

Myeloid Cell Leukemia-1 (MCL-1) is an anti-apoptotic BCL-2 family protein that is necessary to maintain cardiac homeostasis in the adult heart. MCL-1 localizes to two distinct mitochondrial locations in myocytes, both on the outer mitochondrial membrane (MCL-1 OM ) and in the mitochondrial matrix (MCL-1 Matrix ). Our lab previously showed that cardiac-specific ablation of MCL-1 at both of these mitochondrial locations in mice led to severe contractile dysfunction and compromised mitochondrial function. Intriguingly, these defects were accompanied by signs of necrotic, rather than apoptotic, cell death. This indicates that MCL-1 has an alternate role in maintaining mitochondrial homeostasis in cardiac myocytes. Unexpectedly, we found that MCL-1 induces mitochondrial clearance in response to treatment with the chemical uncoupler FCCP in a Parkin-independent manner. Hypoxia is also known to induce mitochondrial clearance, and overexpression of MCL-1 further enhances hypoxia-mediated mitophagy. Fluorescence imaging identified MCL-1-positive mitochondria sequestered inside autophagosomes. MCL-1-mediated clearance is abrogated in autophagy deficient Atg5-/- cells, confirming that clearance is occurring via the autophagy pathway. Mutation of MCL-1’s BH3 domain (G198E D199A) does not affect its ability to induce clearance, suggesting that this role may be independent of its anti-apoptotic function. Also, replacing MCL-1’s BH domains with those of BCL-2, does not affect its ability to induce mitophagy. Next, we investigated whether MCL-1 functions as a mitophagy receptor and promotes removal of damaged mitochondria by binding to directly to LC3 through one or more of its three putative LC3-Interacting Region (LIR) motifs. Endogenous MCL-1 and LC3 co-immunoprecipitate in response to stress induced by FCCP. However, mutating each of MCL-1’s individual LIR motifs, as well as generating combined mutations in all three, does not affect MCL-1-mediated mitophagy. Instead, we found that MCL-1 interacts with the known mitophagy receptor BNIP3 both in vitro and in vivo . Thus, our data suggest that MCL-1 promotes elimination of dysfunctional mitochondria by positively regulating the mitophagy receptor BNIP3.

Gypenosides exert cardioprotective effects by promoting mitophagy and activating PI3K/Akt/GSK-3β/Mcl-1 signaling

“…The potential hub targets included EGFR, MAPK, Src, VEGF, PI3KCA, and Mcl-1. Some of them play a significant role in governing mitochondrial homeostasis processes, including protein phosphorylation, mitophagy, and calcium homeostasis; as well as signaling pathways and targets of MAPK, PI3K/Akt, FOXO, and Mcl-1 ( Ajzashokouhi et al, 2023 ; Javadov, Jang & Agostini, 2014 ; Kim & Koh, 2017 ; Moyzis et al, 2019 ; Popov, 2023 ; Yang et al, 2017 ). Therefore, we further investigated the cardioprotective effect and potential mechanisms in Dox-stimulated rat cardiomyocytes based on the results from network pharmacology analysis.…”