Mitochondria-Endoplasmic Reticulum Contact Sites: Regulation and Roles in Coordinating Cell Fate Decisions
Mitochondria-endoplasmic reticulum (ER) contact sites (MERCs) form signalling platforms, which are required for cellular processes such as lipid metabolism, mitochondrial dynamics, mitochondrial Ca2+ signalling and apoptosis. MERCs are vital in coordinating cell fate decisions and their deregulation is associated to the aetiology of several diseases including cancer, diabetes and neurodegenerative disorders. This work focuses on the regulation of MERCs by a contact site regulator and the contribution of aberrant ER-to-mitochondria communication in cancer cell death and chemotherapy resistance. The first part investigates a previously uncharacterised role of a fatty aldehyde dehydrogenase (FALDH), which was identified in a proteomic screen performed in the McBride lab (McGill University, Canada), as a candidate regulator of MERCs and mitochondrial dynamics. FALDH has been documented to exist as two differentially localised isoforms, with one localised to the ER membrane (FALDH-ER) and the other localised to the peroxisomal membrane (FALDH-Px). Here, a novel mitochondrial localisation of FALDH-ER was ascribed and was demonstrated to be enriched in mitochondria-associated ER membranes (MAMs). Using mammalian cell culture models and state-of-the-art microscopy, FALDH-ER was shown to be required for MERCs homeostasis, with its loss inducing a reduction in MERCs and a mitochondrial elongation phenotype. Furthermore, FALDH-ER was shown to be required for the induction of MERCs during starvation. The catalytic activity of FALDH-ER was shown to be independent of its capacity to influence both MERCs and mitochondrial dynamics. Super-resolution microscopy and pure mitochondria isolation revealed that FALDH-ER homodimerization was necessary for mitochondrial localisation and both MERCs and mitochondrial network homeostasis. The loss of FALDH-ER-dependent MERCs did not impair the capacity of mitochondria to uptake Ca2+ from the ER upon stimulation. These observations elicit a paradigm shift in our current understanding of MERCs, indicating the existence of discrete classes of MERCs, with both molecular and functional specificities. Interactome analysis of FALDH-ER revealed a potential involvement of FALDH-ER in the regulation of ER homeostasis through the ubiquitin fold modifier (UFM)-ylation pathway. The loss of FALDH impacted steady state UFMylation and resulted in elevated ER stress. Together, ii this work has identified a novel regulator of MERCs, which is required for the coordination of a specific subset of MERCs, which are functionally distinct to previously described MERCs. The second part focuses on the mechanisms of chemotherapy resistance in a subset of aggressive ovarian and lung cancers. These cancers are driven by the loss of SMARCA4/2 expression, which are subunits of the chromatin remodelling complex, Switch/Non- fermentable (SWI/SNF). The loss of SMARCA4/2 induces the epigenetic silencing of ITPR3, which encodes the inositol-1,4,5-trisphosphate receptor-3 (IP3R3). Using live cell spinning disk confocal microscopy, the loss of IP3R3 was demonstrated to impair ER to mitochondria Ca2+ transfer in these cancers, which subsequently inhibits apoptosis. This is proposed to occur through defective cristae remodelling and inefficient cytochrome C release. Finally, the epigenetic reactivation of SMARCA2 by a histone deacetylase inhibitor (HDACi) was able to restore IP3R3 expression, ER-to-mitochondrial Ca2+ uptake and cancer cell death. Together, a SMARCA4/2-dependent mechanism of apoptosis induction was identified, which may be targeted to enhance chemotherapy response in SMARCA4/2-defficient cancers.
Medical Research Council (MC_UU_00015/7)