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Reduced Apaf-1 levels in cardiomyocytes engage strict regulation of apoptosis by endogenous XIAP.

Potts MB, Vaughn AE, McDonough H, Patterson C, Deshmukh M - J. Cell Biol. (2005)

Bottom Line: The cardiomyocyte resistance to cytochrome c is mediated by endogenous XIAP, as XIAP-deficient cardiomyocytes die rapidly with cytosolic cytochrome c alone.Importantly, we found that cardiomyocytes, like neurons, have markedly reduced Apaf-1 levels and that this decrease in Apaf-1 is directly linked to the tight regulation of caspase activation by XIAP.These data identify an important function of XIAP in cardiomyocytes and point to a striking similarity in the regulation of apoptosis in postmitotic cells.

View Article: PubMed Central - PubMed

Affiliation: Neuroscience Center, Department of Cell and Developmental Biology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.

ABSTRACT
Overexpression studies have identified X-linked inhibitor of apoptosis protein (XIAP) as a potent inhibitor of caspases. However, the exact function of endogenous XIAP in regulating mammalian apoptosis is less clear. Endogenous XIAP strictly regulates cytochrome c-dependent caspase activation in sympathetic neurons but not in many mitotic cells. We report that postmitotic cardiomyocytes, unlike fibroblasts, are remarkably resistant to cytosolic microinjection of cytochrome c. The cardiomyocyte resistance to cytochrome c is mediated by endogenous XIAP, as XIAP-deficient cardiomyocytes die rapidly with cytosolic cytochrome c alone. Importantly, we found that cardiomyocytes, like neurons, have markedly reduced Apaf-1 levels and that this decrease in Apaf-1 is directly linked to the tight regulation of caspase activation by XIAP. These data identify an important function of XIAP in cardiomyocytes and point to a striking similarity in the regulation of apoptosis in postmitotic cells.

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XIAP-deficient cardiomyocytes die with injection of cytochrome c. Cardiomyocytes isolated from XIAP-deficient (−/−) or wild-type (+/+) littermate mice were microinjected with 25 μg/μl of bovine cytochrome c. As a control, XIAP-deficient cardiomyocytes were also injected with 25 μg/μl of yeast cytochrome c. (A) Phase-contrast and fluorescence photographs of representative cells. Arrows mark the injected cells, identified by coinjection of rhodamine dextran along with cytochrome c. (B) Quantitation of cell survival at multiple times after the injections. Data shown are the mean ± SEM of three independent experiments.
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fig3: XIAP-deficient cardiomyocytes die with injection of cytochrome c. Cardiomyocytes isolated from XIAP-deficient (−/−) or wild-type (+/+) littermate mice were microinjected with 25 μg/μl of bovine cytochrome c. As a control, XIAP-deficient cardiomyocytes were also injected with 25 μg/μl of yeast cytochrome c. (A) Phase-contrast and fluorescence photographs of representative cells. Arrows mark the injected cells, identified by coinjection of rhodamine dextran along with cytochrome c. (B) Quantitation of cell survival at multiple times after the injections. Data shown are the mean ± SEM of three independent experiments.

Mentions: Western analysis indicated that cardiomyocytes express multiple IAPs, including XIAP, c-IAP-1, and c-IAP-2 (Fig. 2 A). Despite the expression of multiple IAPs in sympathetic neurons, the removal of XIAP alone is sufficient to permit cytochrome c–dependent caspase activation in neurons (Potts et al., 2003). To determine whether endogenous XIAP is an important regulator of caspase activation in cardiomyocytes, we examined the ability of cytochrome c to induce death in cardiomyocytes isolated from XIAP-deficient mice (Harlin et al., 2001). In contrast to wild-type cardiomyocytes that are resistant to cytochrome c, the XIAP-deficient cardiomyocytes were strikingly permissive and died with cytosolic microinjection of cytochrome c alone (Fig. 3 A). 50% of XIAP-deficient cardiomyocytes were dead within 2 h of cytochrome c injections, and <15% remained alive 9 h after the injections. In contrast, 80% of wild-type cardiomyocytes remained viable even 9 h after cytochrome c injections (Fig. 3 B). The XIAP-deficient cardiomyocytes were not generally susceptible to microinjection per se, as control injections with yeast cytochrome c did not induce death in these cells.


Reduced Apaf-1 levels in cardiomyocytes engage strict regulation of apoptosis by endogenous XIAP.

Potts MB, Vaughn AE, McDonough H, Patterson C, Deshmukh M - J. Cell Biol. (2005)

XIAP-deficient cardiomyocytes die with injection of cytochrome c. Cardiomyocytes isolated from XIAP-deficient (−/−) or wild-type (+/+) littermate mice were microinjected with 25 μg/μl of bovine cytochrome c. As a control, XIAP-deficient cardiomyocytes were also injected with 25 μg/μl of yeast cytochrome c. (A) Phase-contrast and fluorescence photographs of representative cells. Arrows mark the injected cells, identified by coinjection of rhodamine dextran along with cytochrome c. (B) Quantitation of cell survival at multiple times after the injections. Data shown are the mean ± SEM of three independent experiments.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2171313&req=5

fig3: XIAP-deficient cardiomyocytes die with injection of cytochrome c. Cardiomyocytes isolated from XIAP-deficient (−/−) or wild-type (+/+) littermate mice were microinjected with 25 μg/μl of bovine cytochrome c. As a control, XIAP-deficient cardiomyocytes were also injected with 25 μg/μl of yeast cytochrome c. (A) Phase-contrast and fluorescence photographs of representative cells. Arrows mark the injected cells, identified by coinjection of rhodamine dextran along with cytochrome c. (B) Quantitation of cell survival at multiple times after the injections. Data shown are the mean ± SEM of three independent experiments.
Mentions: Western analysis indicated that cardiomyocytes express multiple IAPs, including XIAP, c-IAP-1, and c-IAP-2 (Fig. 2 A). Despite the expression of multiple IAPs in sympathetic neurons, the removal of XIAP alone is sufficient to permit cytochrome c–dependent caspase activation in neurons (Potts et al., 2003). To determine whether endogenous XIAP is an important regulator of caspase activation in cardiomyocytes, we examined the ability of cytochrome c to induce death in cardiomyocytes isolated from XIAP-deficient mice (Harlin et al., 2001). In contrast to wild-type cardiomyocytes that are resistant to cytochrome c, the XIAP-deficient cardiomyocytes were strikingly permissive and died with cytosolic microinjection of cytochrome c alone (Fig. 3 A). 50% of XIAP-deficient cardiomyocytes were dead within 2 h of cytochrome c injections, and <15% remained alive 9 h after the injections. In contrast, 80% of wild-type cardiomyocytes remained viable even 9 h after cytochrome c injections (Fig. 3 B). The XIAP-deficient cardiomyocytes were not generally susceptible to microinjection per se, as control injections with yeast cytochrome c did not induce death in these cells.

Bottom Line: The cardiomyocyte resistance to cytochrome c is mediated by endogenous XIAP, as XIAP-deficient cardiomyocytes die rapidly with cytosolic cytochrome c alone.Importantly, we found that cardiomyocytes, like neurons, have markedly reduced Apaf-1 levels and that this decrease in Apaf-1 is directly linked to the tight regulation of caspase activation by XIAP.These data identify an important function of XIAP in cardiomyocytes and point to a striking similarity in the regulation of apoptosis in postmitotic cells.

View Article: PubMed Central - PubMed

Affiliation: Neuroscience Center, Department of Cell and Developmental Biology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.

ABSTRACT
Overexpression studies have identified X-linked inhibitor of apoptosis protein (XIAP) as a potent inhibitor of caspases. However, the exact function of endogenous XIAP in regulating mammalian apoptosis is less clear. Endogenous XIAP strictly regulates cytochrome c-dependent caspase activation in sympathetic neurons but not in many mitotic cells. We report that postmitotic cardiomyocytes, unlike fibroblasts, are remarkably resistant to cytosolic microinjection of cytochrome c. The cardiomyocyte resistance to cytochrome c is mediated by endogenous XIAP, as XIAP-deficient cardiomyocytes die rapidly with cytosolic cytochrome c alone. Importantly, we found that cardiomyocytes, like neurons, have markedly reduced Apaf-1 levels and that this decrease in Apaf-1 is directly linked to the tight regulation of caspase activation by XIAP. These data identify an important function of XIAP in cardiomyocytes and point to a striking similarity in the regulation of apoptosis in postmitotic cells.

Show MeSH
Related in: MedlinePlus