*6.3.2. Endoplasmic reticulum: Unfolded protein response*

The ER has primordial roles in normal physiological and survival processes. These include intracellular calcium homeostasis, protein secretion and lipid biosynthesis [58]. Apoptosis can be initiated as a consequence of stress in the ER. This stress condition can be caused by calcium homeostasis alteration, glucose deprivation, hypoxia, low redox potential, excessive protein synthesis or defective protein secretion. These insults can cause accumulation of

unfolded proteins in the lumen of the ER, triggering an evolutionary conserved signalling pathway known as the Unfolded Protein Response (UPR) which can culminate in cell death. UPR consists of global protein synthesis reduction, synthesis induction of chaperones and other proteins related to protein folding and retro-translocation of misfolded or unfolded proteins from the ER to the cytosol, where they will be degraded by the proteasome [59]. When the ER stress is sustained and ER function cannot be restored, UPR activates a specific apoptotic pathway. Caspase-12, which is localized in the ER, is activated by calciumdependent proteases known as calpains [60]. Once activated, caspase-12 activates caspase-9 without apoptosome intervention [61,62]. It has been postulated that ER stress can also activate caspase-8, which induces cytochrome c release through Bid processing [29,63]. In addition, ER stress can also initiate **autophagy,** a different type of cell death.

Cell Death and Cancer, Novel Therapeutic Strategies 79

when they become detached from the extracellular matrix or neighbouring cells [68,69]. Besides its well-established role in mitosis, cytoskeleton components can modulate mitochondria. For example, microtubules sequester BH3-only proteins Bim and Bmf, which interact with dynein [70]. Gelsolin has anti-apoptotic effect by closing the VDAC channel

Poly-ADP-Ribose Polymerase (PARP) is a family of 16 nuclear enzymes, among which, the best characterized is PARP-1. PARPs have several functions in cell proliferation, cell death, DNA recombination and DNA repair. PARP-1 is a 116 kDa nuclear protein involved in DNA repair mechanisms. PARP synthesis is activated when DNA is fragmented in the presence of nuclear poly-ADP ribosylated proteins. In an early apoptotic stage, caspases cleave PARP resulting in an 89 kDa and a 24 kDa fragments [73]. The smaller fragment irreversibly binds DNA fragment ends, impeding the access of DNA repair enzymes. Hence, PARP proteolysis facilitates nuclear disorganization and ensures irreversibility of the apoptotic process [74]. PARP cleaves also takes place during necrosis. However, the fragments obtained are of different size [75]. A role of PARP cleavage in autophagy induced by DNA damage has been recently suggested [76]. **Parthanatos** is a particular case of regulated necrosis in which PARP activation plays an important role. PAR polymer, the product of PARP-1 activation, translocates to the mitochondria and induces AIF release. Subsequently, AIF translocates to the nucleus and provokes chromatin condensation and DNA fragmentation [77]. Necroptosis or regulated necrosis also involves PARP activation [78]. Therefore, PARP can participate in different types of cell death and it is not exclusive of

**Autophagy** is a self-digestive physiological process that occurs in all eukaryotic cells, during which long-lived proteins and organelles are degraded by lysosomes to maintain cellular homeostasis [79]. To date, at least three types of autophagic pathways have been described: macroautophagy (simply called ''autophagy''), microautophagy and Chaperone-Mediated Autophagy (CMA). These forms differ in the mode the cargo is delivered to the lysosome.

**Macroautophagy** is a dynamic process in which portions of the cellular cytoplasm and organelles are sequestered in a double-membrane bound vesicle called autophagosome, which then fuses with the lysosome [80,81]. **Microautophagy** involves the sequestering by the lysosome itself of part of the cytoplasm. During microautophagy, the membrane of the lysosome invaginates, and then pinches off to form an internal vacuole that contains material derived from the cytoplasm [82]. The notable difference between macroautophagy and microautophagy is that in the latter, part of the cytoplasm is directly taken up into the lysosome. Both macroautophagy and microautophagy are basically nonselective degradation pathways in which bulk cytoplasm is randomly sequestered. However, in some

[71]. Actin can also modulate VDAC closure [69,72].

apoptosis, as previously thought.

**8. Autophagy** 

**7. Parp proteolysis as an indicator of cell death** 
