The current model of apoptosis holds that upstream signals lead to activation of downstream effector caspases. We generated mice deficient in the two effectors, caspase 3 and caspase 7, which died immediately after birth with defects in cardiac development. Fibroblasts lacking both enzymes were highly resistant to both mitochondrial and death receptor-mediated apoptosis, displayed preservation of mitochondrial membrane potential, and had defective nuclear translocation of apoptosis-inducing factor (AIF). Furthermore, the early apoptotic events of Bax translocation and cytochrome c release were also delayed. We conclude that caspases 3 and 7 are critical mediators of mitochondrial events of apoptosis.Mitochondria play a central role in apoptosis. Mitochondrial outer membrane permeabilization (MOMP) leads to release of proapoptotic factors such as cytochrome c and AIF (1). Furthermore, loss of mitochondrial membrane potential (Δψ m ) is thought to contribute to cell death by disruption of normal mitochondrial function (2, 3). Interaction of members of the Bcl-2 family of proteins regulates MOMP, the key event of cytochrome c release into the cytoplasm (3, 4). What is less clear, however, is the precise role of caspase proteases in mitochondrial events of apoptosis. Although upstream caspases, such as caspase 2 and caspase 8, affect mitochondrial events in both death-receptor and mitochondrial pathways of apoptosis, either directly or through interaction with Bcl-2 family members, the role of presumed downstream "effector" caspases in this process is less clear (5, 6). Therefore, we studied the two highly related effectors, caspase 3 and caspase 7, to elucidate their functions in apoptosis.* To whom correspondence should be addressed. richard.flavell@yale.edu. We generated caspase 7 −/− mice ( fig. S1), which were born in ratios consistent with Mendelian inheritance. They had normal appearance, organ morphology, and lymphoid development. When caspase 7 −/− mouse embryonic fibroblasts (MEFs) were treated with inducers of apoptosis, they exhibited a slight survival advantage as compared with wild-type MEFs. Apoptosis caused by a range of insults in other caspase 7 −/− cells proceeded normally, however, including the death of activated T cells following stimulation of the T cell receptor, thymocyte apoptosis, Fas-mediated death of B cells, and Fas-mediated death of hepatocytes ( fig. S2). Supporting Online MaterialCaspase 3, which is structurally similar to caspase 7, might compensate for the lack of caspase 7, which would lead to this relatively mild antiapoptotic phenotype (7,8). Thus, we bred caspase 7 −/− mice to caspase 3 −/− mice previously described by our laboratory (9). The embryonic stem cells containing the mutation were from the 129/SvJ genetic background. Mice derived from these embryonic stem cells were backcrossed six generations onto the C57BL/6 background. We obtained no live caspase 3 −/− /caspase 7 −/− double-knockout (DKO) mice when progeny were genotyped at an age of 10 to 14 days. DKO mic...

The mechanism by which cells undergo death determines whether dying cells trigger inflammatory responses or remain immunologically silent. Mitochondria play a central role in the induction of cell death, as well as in immune signaling pathways. Here, we identify of a mechanism by which mitochondria and downstream pro-apoptotic caspases regulate the activation of antiviral immunity. In the absence of active caspases, mitochondrial outer membrane permeabilization by Bax and Bak results in the expression of type I interferons (IFNs). This induction is mediated by mitochondrial DNA-dependent activation of the cGAS/STING pathway and results in the establishment of a potent state of viral resistance. Our results show that mitochondria have the capacity to simultaneously expose a cell-intrinsic inducer of the IFN response, and to inactivate this response in a caspase-dependent manner. This mechanism provides a dual control, which determines whether mitochondria initiate an immunologically silent or a pro-inflammatory type of cell death.

The cJun NH2-terminal kinase (JNK) signaling pathway contributes to inflammation and plays a key role in the metabolic response to obesity, including insulin resistance. Macrophages are implicated in this process. To test the role of JNK, we established mice with selective JNK-deficiency in macrophages. We report that feeding a high fat diet to control and JNK-deficient mice caused similar obesity, but only mice with JNK-deficient macrophages remained insulin sensitive. The protection of mice with macrophage-specific JNK-deficiency against insulin resistance was associated with reduced tissue infiltration by macrophages. Immunophenotyping demonstrated that JNK was required for pro-inflammatory macrophage polarization. These studies demonstrate that JNK in macrophages is required for the establishment of obesity-induced insulin resistance and inflammation.

Caspase-3 is a critical enzyme for apoptosis and cell survival. Here we report delayed ossification and decreased bone mineral density in caspase-3-deficient (Casp3 -/-and Casp3 +/-) mice due to an attenuated osteogenic differentiation of bone marrow stromal stem cells (BMSSCs). The mechanism involved in the impaired differentiation of BMSSCs is due, at least partially, to the overactivated TGF-β/Smad2 signaling pathway and the upregulated expressions of p53 and p21 along with the downregulated expressions of Cdk2 and Cdc2, and ultimately increased replicative senescence. In addition, the overactivated TGF-β/Smad2 signaling may result in the compromised Runx2/Cbfa1 expression in preosteoblasts. Furthermore, we demonstrate that caspase-3 inhibitor, a potential agent for clinical treatment of human diseases, caused accelerated bone loss in ovariectomized mice, which is also associated with the overactivated TGF-β/Smad2 signaling in BMSSCs. This study demonstrates that caspase-3 is crucial for the differentiation of BMSSCs by influencing TGF-β/Smad2 pathway and cell cycle progression.