ApoReview-Introduction to Apoptosis:Page 12 of 26 4.4 Mitochondria as central regulators of intrinsic apoptosis pathways Besides amplifying and mediating extrinsic apoptotic pathways,mitochondria also play a central role in the integration and propagation of death signals originating from inside the cell such as DNA damage,oxidative stress,starvation,as well as those induced by chemotherapeutic drugs [Kaufmann, .201].Most apoptosis-inducing conditions involve the disruption of the mitochondria inner transmembrane potential (w)as well as the so called permeability transition (PT)a sudden increase of the inner mitochondrial membrane permeability to solutes with a molecular mass below approximately 1.5 kDa.Concomitantly,osmotic mitochondrial swelling has been observed by influx of water into the matrix with eventual rupture of theouter mitochondrial membrane,resulting in the release of proapoptotic proteins from the mitochondrial intermembrane space into the cytoplasm [Berardi,1999 Loeffler,2000].Released proteins include cytochrome c.which activates the apoptosome and therefore the caspase cascade,but alsoother factors such as the apoptosis-inducing factor AIF [Susin,1999],the endonuclease endoG [Li,2001].Smac/Diablo [Verhagen,2000],and Htr/Omi [Verhagen,2002].Interestingly,PT is always followed by Aw,but Aw is not always caused by PT,and cytochrome crelease has been observed even in absence of A [Bernardi,1999:Kroemer. addition to the release of mitochondrial factors,the dissipation of and PTalso cause a loss of the biochemical homeostasis of the cell:ATP synthesis is stopped,redox molecules such as NADH,NADPH,and glutathione are oxidized,and reactive oxygen species (ROS)are increasingly generated [Kroemer,2000.Kroemer,1997].Increased levels of ROS directly cause the oxidation of lipids,proteins,and nucleic acids,thereby enhancing the disruption of A as part of a positive feedback [Marchetti,1997].Several possible mechanisms for PT have been proposed,but there appears to exist consent that a so-called permeability transition pore(PTP)is formed consisting of the adenin nucleotide translocator(ANT)and the voltage-dependent anion channel (VDAC)as its cor components.ANT is the most abundant protein of the inner mitochondrial membrane and as a transmembrane channel is responsible for the export of ATP in exchange with ADP(antiport) Overexpression of ANT-1 in human cancer cell lines dominantly induces apoptosis with all its characteristic features whereas its closely conserved homologue ANT-2 does not,indicating a specific mechanistic role of ANT-1 in mitochondrial apoptosis events [Bauer,199 VDAC,porin, is the most abundant protein of the outer mitochondrial membrane and forms a non-selective pore through the outer membrane.Indicated by direct protein-protein interactions,VDAC-ANT complexes presumably connect inner and outer mitochondrial membrane to so-called 'contact sites corresponding to a close association of the two membranes and thereby possibly constituting the PT pore [Beutner,1998].Since PT,loss of Aw,and release of mitochondrial proteins are of central importance in mediating and enhancing apoptotic pathways,those mitochondrial events must be kept under strict of regulatory mechanisms which are in many ways dependent on members of the Bcl-2 family which will be discussed in the next paragraph. ApoReview - Introduction to Apoptosis: Page 12 of 26 4.4 Mitochondria as central regulators of intrinsic apoptosis pathways Besides amplifying and mediating extrinsic apoptotic pathways, mitochondria also play a central role in the integration and propagation of death signals originating from inside the cell such as DNA damage, oxidative stress, starvation, as well as those induced by chemotherapeutic drugs [Kaufmann, 2000; Wang, 2001]. Most apoptosis-inducing conditions involve the disruption of the mitochondrial inner transmembrane potential (∆ψ) as well as the so called permeability transition (PT), a sudden increase of the inner mitochondrial membrane permeability to solutes with a molecular mass below approximately 1.5 kDa. Concomitantly, osmotic mitochondrial swelling has been observed by influx of water into the matrix with eventual rupture of the outer mitochondrial membrane, resulting in the release of proapoptotic proteins from the mitochondrial intermembrane space into the cytoplasm [Bernardi, 1999; Loeffler, 2000]. Released proteins include cytochrome c, which activates the apoptosome and therefore the caspase cascade, but also other factors such as the apoptosis-inducing factor AIF [Susin, 1999], the endonuclease endoG [Li, 2001], Smac/Diablo [Verhagen, 2000], and Htr/Omi [Verhagen, 2002]. Interestingly, PT is always followed by ∆ψ, but ∆ψ is not always caused by PT, and cytochrome c release has been observed even in absence of ∆ψ [Bernardi, 1999; Kroemer, 2000]. In addition to the release of mitochondrial factors, the dissipation of ∆ψ and PT also cause a loss of the biochemical homeostasis of the cell: ATP synthesis is stopped, redox molecules such as NADH, NADPH, and glutathione are oxidized, and reactive oxygen species (ROS) are increasingly generated [Kroemer, 2000; Kroemer, 1997]. Increased levels of ROS directly cause the oxidation of lipids, proteins, and nucleic acids, thereby enhancing the disruption of ∆ψ as part of a positive feedback [Marchetti, 1997]. Several possible mechanisms for PT have been proposed, but there appears to exist consent that a so-called permeability transition pore (PTP) is formed consisting of the adenin nucleotide translocator (ANT) and the voltage-dependent anion channel (VDAC) as its core components. ANT is the most abundant protein of the inner mitochondrial membrane and as a transmembrane channel is responsible for the export of ATP in exchange with ADP (antiport). Overexpression of ANT-1 in human cancer cell lines dominantly induces apoptosis with all its characteristic features whereas its closely conserved homologue ANT-2 does not, indicating a specific mechanistic role of ANT-1 in mitochondrial apoptosis events [Bauer, 1999]. VDAC, also called porin, is the most abundant protein of the outer mitochondrial membrane and forms a non-selective pore through the outer membrane. Indicated by direct protein-protein interactions, VDAC-ANT complexes presumably connect inner and outer mitochondrial membrane to so-called ‘contact sites’, corresponding to a close association of the two membranes and thereby possibly constituting the PT pore [Beutner, 1998]. Since PT, loss of ∆ψ, and release of mitochondrial proteins are of central importance in mediating and enhancing apoptotic pathways, those mitochondrial events must be kept under strict control of regulatory mechanisms which are in many ways dependent on members of the Bcl-2 family which will be discussed in the next paragraph