The role of mitochondria in regulating the senescence-associated secretory phenotype during cellular senescence

Abstract

Senescent cells have been implicated in the ageing process and in the development of various age-related disorders. A key characteristic of senescent cells is the presence of dysfunctional mitochondria, which are crucial for the development of the SASP (Correia-Melo et al., 2016). However, at this stage the underlying mechanisms by which mitochondria drive the SASP have not yet been completely elucidated. In this thesis, I have discovered that mitochondria in senescent fibroblasts have an increased abundance of mitochondrial DNA (mtDNA) in the cytosol compared to young proliferating fibroblasts. Using super resolution microscopy, I discovered that a small subset of mitochondria in senescent cells display a recently identified phenomenon termed: limited mitochondrial outer membrane permeabilisation (miMOMP) (Ichim et al., 2015). This subset of mitochondria show activation of pro-apoptotic BAX and BAK which cause a limited release of cytochrome c and caspase-dependent DNA damage in the absence of cell-death in senescent cells. My data indicates that during senescence, miMOMP and the presence of BAX/BAK mitochondrial outer membrane pores allow the release of mtDNA to the cytosol. Consistent with a role for mitochondrial BAX/BAK and the release of mtDNA in cellular senescence, I found that both CRISPR/CAS9 deletion of BAX and BAK suppresses the SASP and the release of mtDNA during senescence, whilst maintaining the cell cycle arrest. Additionally, my data shows that cytosolic mtDNA binds to the DNA sensor cGAS, potentially activating the cGASSTING pathway which has been previously implicated in SASP regulation (Dou et al., 2017; Gluck et al., 2017; Yang et al., 2017). Moreover, genetic silencing of BAX/BAK in vivo prevents the upregulation of SASP related genes following the induction of senescence by irradiation. My data argue that the mitochondrial apoptotic pathway is a key SASP regulator. Finally, I have demonstrated that pharmacological inhibitors of either BAX or cGAS lead to a robust suppression of the SASP, without impairing the cell cycle arrest. These small molecule inhibitors may therefore pose as potential candidates for senostatic therapies in offsetting the detrimental effects associated with the SASP.