Role of prodomain in importin-mediated nuclear localization and activation of caspase-2.

Caspase-2 is unique among mammalian caspases because it localizes to the nucleus in a prodomain-dependent manner. The caspase-2 prodomain also regulates caspase-2 activity via a caspase recruitment domain that mediates oligomerization of procaspase-2 molecules and their subsequent autoactivation. In this study we sought to map specific functional regions in the caspase-2 prodomain that regulate its nuclear transport and also its activation. Our data indicate that caspase-2 contains a classical nuclear localization signal (NLS) at the C terminus of the prodomain which is recognized by the importin alpha/beta heterodimer. The mutation of a conserved Lys residue in the NLS abolishes nuclear localization of caspase-2 and binding to the importin alpha/beta heterodimer. Although caspase-2 is imported into the nucleus, mutants lacking the NLS were still capable of inducing apoptosis upon overexpression in transfected cells. We define a region in the prodomain that regulates the ability of caspase-2 to form dot- and filament-like structures when ectopically expressed, which in turn promotes cell killing. Our data provides a mechanism for caspase-2 nuclear import and demonstrate that association of procaspase-2 into higher order structures, rather than its nuclear localization, is required for caspase-2 activation and its ability to induce apoptosis.

A novel Apaf-1-independent putative caspase-2 activation complex.

Caspase activation is a key event in apoptosis execution. In stress-induced apoptosis, the mitochondrial pathway of caspase activation is believed to be of central importance. In this pathway, cytochrome c released from mitochondria facilitates the formation of an Apaf-1 apoptosome that recruits and activates caspase-9. Recent data indicate that in some cells caspase-9 may not be the initiator caspase in stress-mediated apoptosis because caspase-2 is required upstream of mitochondria for the release of cytochrome c and other apoptogenic factors. To determine how caspase-2 is activated, we have studied the formation of a complex that mediates caspase-2 activation. Using gel filtration analysis of cell lysates, we show that caspase-2 is spontaneously recruited to a large protein complex independent of cytochrome c and Apaf-1 and that recruitment of caspase-2 to this complex is sufficient to mediate its activation. Using substrate-binding assays, we also provide the first evidence that caspase-2 activation may occur without processing of the precursor molecule. Our data are consistent with a model where caspase-2 activation occurs by oligomerization, independent of the Apaf-1 apoptosome.

Role of Bcl-2 family of proteins in malignancy.

B cell lymphoma gene-2 (Bcl-2) is the prototypic member of a growing family of proteins that play evolutionarily conserved, key regulatory roles in apoptosis. The Bcl-2 family members are characterized by the presence of one or more Bcl-2 homology domains and are comprised of both the prosurvival and proapoptotic proteins. Bcl-2 itself is a prosurvival member of the family and its aberrant expression has been linked to a variety of different cancers, including several hematological malignancies. Although the exact mechanism of action of Bcl-2 family of proteins in regulating apoptosis is still a matter of some debate, these proteins appear to act upstream of caspase activation. Many recent studies have shown the therapeutic potential of targeting Bcl-2 family members for the treatment of cancer. This article summarizes what is currently known about Bcl-2-like proteins and how the evolving understanding of the biology of these proteins is paving way for the development of novel cancer therapeutics.