When death starts with a caspase

When death starts with a caspase hen a cytotoxic lymphocyte delivers granzyme B to a target cell, current models emphasize that the enzyme induces apoptosis through a mitochondria-centered pathway, cleaving the proapoptotic Bcl-2 family member Bid to cause mitochondrial permeabilization. On page 875, Metkar et al. overturn this view, showing that in the dominant pathway granzyme B begins by processing procaspase-3, whereas mitochondria are secondary components that amplify the signal once caspase-3 has been activated. The work adds to recent evidence that caspases can initiate apoptosis, whereas mitochondria serve as signal amplifiers. The authors used a variety of strategies to avoid common pitfalls during granzyme-mediated apoptosis assays. In past assays, uncom-plexed granzyme B was added to cell extracts. Granzyme B in the new assays was delivered into cells using adenovirus particles, and was presented in a complex with the proteoglycan serglycin to mimic the enzyme's natural state. Doing the apoptosis assays with whole cells minimized the risk of proteolysis occurring at sites not normally frequented by granzyme B. In this system, Bid cleavage was not detected, and cells deficient in procaspase-3 failed to undergo mitochondrial depolarization or DNA fragmentation, indicating that granzyme B acts first through procaspase-3 rather than the mitochondria. Caspase-3–induced permeabilization of the mitochondria amplifies the death signal but, based on assays with cells harboring a Bax/Bak deletion, this amplification is not essential for apoptosis. ᭿ W Granzyme B delivered via adenovirus (AD) requires caspase 3 in order to kill cells. iu et al. (page 899) genetically disrupted the gene for paramyosin, a major component of invertebrate muscle thick filaments, in Drosophila melanogaster. In addition to the expected finding, that the disruption damages myofibril structure, the work identifies a surprising requirement for L paramyosin in myoblast fusion. Mutant flies in which a P-element has been excised from the paramyosin promoter region die during embryonic development. In addition to defects in myofibril assembly, the embryos exhibit sporadic failures in myoblast fusion, leading to the absence of some muscle fibers. Antibody localization experiments show paramyosin in discrete foci at Normal muscle fiber development (top) goes wrong without paramyosin (bottom). sites near the junctions of fusing myoblasts, and nonmuscle myosin appears to colocalize to the same sites. The authors propose that, in the fly, paramyosin functions as a cytoskeletal protein during early development. In this model, paramyosin and nonmuscle myosin form minifilaments that interact with the actin cytoskeleton and promote myoblast …