Oncogene (2004) 23, 2881–2890 & 2004 Nature Publishing Group All rights reserved 0950-9232/04 $25.00 www.nature.com/onc Lysosomes in cell death Maria Eugenia Guicciardi1, Marcel Leist2 and Gregory J Gores*,1 1Mayo Clinic College of Medicine, Rochester, MN 55905, USA; 2H. Lundbeck A/S, DK-2500 Valby, Denmark For many years apoptosis research has focused on the magnitude of lysosomal permeabilization, and, caspases and their putative role as sole executioners of consequently, the amount of proteolytic enzymes programmed cell death. Accumulating information now released into the cytosol (Li et al., 2000). A complete suggests that lysosomal cathepsins are also pivotally breakdown of the organelle with release of high involved in this process, especially in pathological condi- concentrations of lysosomal enzymes into the cytosol tions. In particular, the role of lysosomes and lysosomal results in unregulated necrosis, whereas partial, selective enzymes in initiation and execution of the apoptotic permeabilization triggers apoptosis (Bursch, 2001; Turk program has become clear in several models, to the point et al., 2002). In both instances, the amount of released that the existence of a ‘lysosomal pathway of apoptosis’is enzymes is sufficient to overcome the protective barrier now generally accepted. This pathway of apoptosis can be of the endogenous inhibitors present in the cytosol (i.e. activated by death receptors, lipid mediators, and photo- cystatins), which usually prevent potential damage from damage. Lysosomal proteases can be released from the the spontaneous leakage of lysosomal enzymes naturally lysosomes into the cytosol, where they contribute to the occurring through the lysosomal membrane (Berg et al., apoptotic cascade upstream of mitochondria. This review 1995; Claus et al., 1998). Once in the cytosol, lysosomal focuses on the players and the molecular mechanisms enzymes can contribute to the execution of the apoptotic involved in the lysosomal pathway of apoptosis as well as program either by direct cleavage of key cellular on the importance of this pathway in development and substrates, or by acting in concert with the caspases in pathology. the signaling pathway (Leist and Jaattela, 2001a, b). Oncogene (2004) 23, 2881–2890. doi:10.1038/sj.onc.1207512 However, the precise mechanisms by which lysosomes are involved in apoptosis are still largely unknown and Keywords: cathepsins; Spi2A; TNFa currently under intense investigation. The most recent findings regarding the role of lysosomal proteases in apoptosis are described below. Introduction When does the lysosomal pathway of cell death occur? For many years, lysosomes have been thought to be solely involved in necrotic and autophagic cell death, Lysosomal enzymes were among the first proteases to be with their role in apoptosis being limited to the digestion associated with programmed cell death, for example, in of engulfed apoptotic bodies (Ferri and Kroemer, 2001a; the tadpole tail during the 1960s. However, this line of Leist and Jaattela, 2001a). Likewise, lysosomal protease research has not been actively perpetuated into the functions were believed to be limited to nonspecific modern age of apoptosis. While it is firmly established intracellular protein degradation occurring within the that autophagy, and herewith lysosomes, plays a role in lysosome. These concepts now seem to be outdated by a developmental cell death, the evidence for a pivotal role growing body of evidence that strongly points to a role of individual cathepsins in developmental apoptosis is of lysosomes in apoptosis that goes beyond that of poor or negative. Unlike many other pathways of simple ‘garbage disposals’. Indeed, partial lysosomal apoptosis, lysosomal cathepsins do not appear to permeabilization with subsequent release of proteolytic contribute to developmental or physiologic apoptosis enzymes into the cytosol, and their active contribution of the central nervous system (CNS), heart, liver, and to the signaling pathways, has been recently described in limbs. In contrast, various caspase knockout mice show several models of apoptosis. This phenomenon is so developmental defects in these tissues (Los et al., 1999). widely recognized that the ‘lysosomal pathway of The lysosomal pathway of apoptosis has been identified apoptosis’ is now accepted terminology. The key-factor primarily in pathological situations. For instance, in determining the type of cell death (necrosis vs cathepsins have been implicated in CNS apoptosis apoptosis) mediated by lysosomal enzymes seems to be following ischemia and during neurodegenerative pro- cesses (Yamashima et al., 1998; Tsuchiya et al., 1999; *Correspondence:GJ Gores, Mayo Medical School, Clinic, and Lieuallen et al., 2001; Houseweart et al., 2003a). In the Foundation, 200 First Street SW, Rochester, MN 55905, USA; liver, a lysosomal-mediated pathway of apoptosis has E-mail:[email protected] been found to contribute to TNF-a signaling, liver Lysosomes in cell death ME Guicciardi et al 2882 damage following bile duct obstruction, and cold- Oncogenic activation sensitizes cancer cells to various ischemia/warm-reperfusion injury (Guicciardi et al., apoptotic stimuli. Transformation of cells, for example 2000; Guicciardi et al., 2001; Canbay et al., 2003). by Ras, strongly affects processing, trafficking and Another pathological situation favoring the activation subcellular localization of lysosomal enzymes (Nishi- of the lysosomal pathway is tumorigenic transformation mura et al., 1998; Demoz et al., 1999), and it is of cells. Many tumors express considerably increased conceivable that cathepsins might take part in the amounts of cathepsins and the TNF-dependent apop- transformation-induced death of tumor cells. Indeed, tosis pathway in tumor cells is often particularly Ras triggers cell death of human cancer cells by acti- cathepsin B dependent (Foghsgaard et al., 2002; vation of a caspase-independent cell death program (Chi Fehrenbacher et al., unpublished). et al., 1999), which may involve lysosomal proteases. Photodynamic therapy, a process employing UV-light The overexpression of cathepsins often observed in and photosensitizers to kill cancer cells, as well as accu- tumors should render cancer cells more susceptible to mulation of lysosomotropic agents within the organelle, cell death via the lysosomal pathway. It should be also triggers the lysosomal pathway of cell death (Boya possible to exploit this pathway to differentially et al., 2003a, b; Cirman et al., 2004; Li et al., 2003). As modulate susceptibility of cancer cells to cell death. this pathway becomes better understood, its role in For example, tumor necrosis factor-related apoptosis- additional pathologies will likely become apparent. inducing ligand (TRAIL) preferentially kills tumor cells both in vitro and in vivo (Ashkenazi et al., 1999; Walczak et al., 1999). Although the underlying mechanism of Role of the lysosomal pathway of cell death in cancer TRAIL selectivity toward tumor cells remains unclear, biology TRAIL, which can trigger lysosomal cell death (Foghs- gaard et al., 2001), might induce apoptosis in trans- Many lysosomal enzymes, including cathepsins, are over- formed cells via the lysosomal pathway. These concepts expressed in cancer (Koblinski et al., 2000). For example, merit further investigation. increased expression and activity of cathepsin B and/or cathepsin D have been observed in breast (Castiglioni et al., 1994), colorectal (Campo et al., 1994; Meyer et al., Mechanisms of lysosomal permeabilization 1997), gastric (Watanabe et al., 1989), lung (Sukoh et al., 1994), prostate (Sinha et al., 1995), and thyroid cancers Vesicle permeabilization in apoptosis is selective. (Shuja and Murnane, 1996), as well as in different brain Whereas lysosomes are permeabilized, endocytic vesicles tumors, including gliomas and meningiomas (Rempel are not, as the absence of total vesicular breakdown et al., 1994; Levicar et al., 2003), and the magnitude of implies the existence of a regulatable process. Consistent expression has often prognostic significance. Many with these observations, even lysosomal breakdown is authors have suggested that enhanced secretion of these partial and/or selective in this apoptotic pathway. proteases by exocytosis promotes cancer growth, inva- Indeed, two manuscripts have reported that in cells sion, and metastasis by promoting degradation of with lysosomes labeled with LysoTracker red, a extracellular connective matrices and angiogenesis (Ko- fluorescent dye that loads predominantly into lyso- blinski et al., 2000). Indeed, inhibition of cathepsin B by somes, release of the dye appears to occur in only a synthetic cysteine protease inhibitors has been shown to subpopulation of lysosomes (Werneburg et al., 2002; effectively reduce the invasiveness of glioblastoma cells Madge et al., 2003). Whether these observations (Demchik et al., 1999) and breast cancer cell lines (Xing represent a time-dependent phenomenon, partial release, et al., 1998). However, the role of the lysosomal pathway or targeting of mediators to a distinct subpopulation of of cell death in transformed cells remains largely ‘killer lysosomes’ remains unclear. unexplored. Tumor cell death is often caspase-indepen- It is unquestionable that lysosomal enzymes, in order dent (Foghsgaard et al., 2001; Leist and Jaattela, 2001a), to participate to the apoptotic process, need to be due to acquired defects in the classic caspase-dependent translocated to the cytosol (and, possibly, to