We hypothesized that the mitochondrial length may be altered according to changes in the sarcomere length, and that this relationship may be affected by exposure to hypoxia. Rat ventricular papillary muscles were isolated and immersed in normoxic or hypoxic solutions for 10 min. Sarcomeres of various lengths were obtained by fixing the papillary muscles in a slack or stretched state, or after exposure to a contracture solution containing saponin and CaCl(2). The mitochondrial length measured using electron microscopy significantly correlated to the length of the adjacent sarcomere in both the normoxic (n=767) and hypoxic (n=1145) groups (P<.0001). The slope of the regression line, however, was significantly less steep, and its intercept was significantly larger in the hypoxic group than in the normoxic group (analysis of covariance). When we analyzed the mitochondrial lengths among the three sarcomere-length subgroups (<1.5, 1.5-2.0, and >2.0 microm), the mitochondrial length was significantly shorter in the hypoxic condition than in the normoxic condition at sarcomere lengths greater than 2.0 microm. Staining for desmin, the major muscle-type intermediate filament, the longitudinal system of which connects the mitochondria with the Z bands of sarcomeres, showed a clear cross-striation pattern in both papillary muscles with and without the exposure to hypoxia, suggesting that desmin was preserved after the exposure to hypoxia. These data indicate that the mitochondrial length changes according to changes in the sarcomere length, suggesting the possible role of mitochondria as an internal load against myocyte contraction. It is also suggested that mitochondria exposed to hypoxia may be more resistive to both compression and stretch in a longitudinal direction than those in the normoxic condition.

Animals↗

PubMedDate not supplied

Microtubule-targeting drugs induce Bcl-2 phosphorylation and association with Pin1.

Bcl-2 is a critical suppressor of apoptosis that is overproduced in many types of cancer. Phosphorylation of the Bcl-2 protein is induced on serine residues in tumor cells arrested by microtubule-targeting drugs (paclitaxel, vincristine, nocodazole) and has been associated with inactivation of antiapoptotic function through an unknown mechanism. Comparison of a variety of pharmacological inhibitors of serine/threonine-specific protein kinases demonstrated that the cyclin-dependent kinase inhibitor, flavopiridol, selectively blocks Bcl-2 phosphorylation induced by antimicrotubule drugs. Bcl-2 could also be coimmunoprecipitated with the kinase Cdc2 in M-phase-arrested cells, suggesting that a Cdc2 may be responsible for phosphorylation of Bcl-2 in cells treated with microtubule-targeting drugs. Examination of several serine-->alanine substitution mutants of Bcl-2 suggested that serine 70 and serine 87 represent major sites of Bcl-2 phosphorylation induced in response to microtubule-targeting drugs. Both these serines are within sequence contexts suitable for proline-directed kinases such as Cdc2. Phosphorylated Bcl-2 protein was discovered to associate in M-phase-arrested cells with Pin1, a mitotic peptidyl prolyl isomerase (PPIase) known to interact with substrates of Cdc2 during mitosis. In contrast, phosphorylation of Bcl-2 induced by microtubule-targeting drugs did not alter its ability to associate with Bcl-2 (homodimerization), Bax, BAG1, or other Bcl-2-binding proteins. Since the region in Bcl-2 containing serine 70 and serine 87 represents a proline-rich loop that has been associated with autorepression of its antiapoptotic activity, the discovery of Pin1 interactions with phosphorylated Bcl-2 raises the possibility that Pin1 alters the conformation of Bcl-2 and thereby modulates its function in cells arrested with antimicrotubule drugs.

Antineoplastic Agents↗

PubMedDate not supplied

Microtubule-targeting drugs induce bcl-2 phosphorylation and association with Pin1.

Bcl-2 is a critical suppressor of apoptosis that is overproduced in many types of cancer. Phosphorylation of the Bcl-2 protein is induced on serine residues in tumor cells arrested by microtubule-targeting drugs (paclitaxel, vincristine, nocodazole) and has been associated with inactivation of antiapoptotic function through an unknown mechanism. Comparison of a variety of pharmacological inhibitors of serine/threonine-specific protein kinases demonstrated that the cyclin-dependent kinase inhibitor, flavopiridol, selectively blocks Bcl-2 phosphorylation induced by antimicrotubule drugs. Bcl-2 could also be coimmunoprecipitated with the kinase Cdc2 in M-phase-arrested cells, suggesting that Cdc2 may be responsible for phosphorylation of Bcl-2 in cells treated with microtubule-targeting drugs. Examination of several serine-->alanine substitution mutants of Bcl-2 suggested that serine 70 and serine 87 represent major sites of Bcl-2 phosphorylation induced in response to microtubule-targeting drugs. Both these serines are within sequence contexts suitable for proline-directed kinases such as Cdc2. Phosphorylated Bcl-2 protein was discovered to associate in M-phase-arrested cells with Pin1, a mitotic peptidyl prolyl isomerase (PPIase) known to interact with substrates of Cdc2 during mitosis. In contrast, phosphorylation of Bcl-2 induced by microtubule-targeting drugs did not alter its ability to associate with Bcl-2 (homodimerization), Bax, BAG1, or other Bcl-2-binding proteins. Since the region in Bcl-2 containing serine 70 and serine 87 represents a proline-rich loop that has been associated with autorepression of its antiapoptotic activity, the discovery of Pin1 interactions with phosphorylated Bcl-2 raises the possibility that Pin1 alters the conformation of Bcl-2 and thereby modulates its function in cells arrested with antimicrotubule drugs.

Antineoplastic Agents↗