E-cadherin-mediated cell contact prevents apoptosis of spontaneously immortalized granulosa cells by regulating Akt kinase activity.
The present studies were designed to determine the role that homophilic E-cadherin binding plays in preventing apoptosis of spontaneously immortalized granulosa cells (SIGCs). Although the levels of E-cadherin were similar to serum control levels, the amount of E-cadherin at the plasma membrane was dramatically reduced by 5 h after serum withdrawal. To determine whether disrupting homophilic E-cadherin binding leads to apoptosis, SIGCs were cultured in serum in the presence of either EGTA or an E-cadherin antibody. Treatment with either EGTA, which disrupts all calcium-dependent contacts, or E-cadherin antibody, induced apoptosis. Exposure to EGTA reduced MEK and Akt kinase activity, whereas E-cadherin antibody only attenuated Akt kinase activity. Because Akt kinase controls caspase-3 activity, an important activator of apoptosis, caspase-3 activity was monitored. Caspase-3 activity increased after serum depletion, or EGTA or E-cadherin antibody treatment. Time-series analysis of caspase-3 activity within single cells revealed that during apoptosis cell contact was disrupted then caspase-3 activity was detected. Finally, the caspase inhibitor, Z-VAD-FMK, blocked apoptosis. These data taken together are consistent with the concept that E-cadherin-mediated cell contact, either directly or indirectly, promotes Akt kinase activity, which in turn, inhibits caspase-3 activation and thereby maintains SIGC viability.
Departments of Physiology and Obstetrics and Gynecology, University of Connecticut Health Center, Farmington, Connecticut 06030, USA. peluso@nso2.uchc.edu
J J
JJ
Peluso
A
A
Pappalardo
G
G
Fernandez
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000818:Animals; D000906:Antibodies / Q000494:pharmacology; D017209:Apoptosis; D015820:Cadherins / Q000032:analysis / Q000276:immunology / Q000502:physiology*; D053148:Caspase 3; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism; D002448:Cell Adhesion; D002461:Cell Line, Transformed; D002462:Cell Membrane / Q000737:chemistry; D016895:Culture Media, Serum-Free; D003598:Cytoskeletal Proteins / Q000032:analysis; D004533:Egtazic Acid / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D006107:Granulosa Cells / Q000166:cytology* / Q000201:enzymology*; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt; D015398:Signal Transduction; D015534:Trans-Activators; D051176:beta Catenin
D000590:Amino Acid Chloromethyl Ketones; D000906:Antibodies; D015820:Cadherins; D061945:Caspase Inhibitors; D016895:Culture Media, Serum-Free; D003598:Cytoskeletal Proteins; D004791:Enzyme Inhibitors; D011518:Proto-Oncogene Proteins; D015534:Trans-Activators; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; D051176:beta Catenin; D004533:Egtazic Acid; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D053148:Caspase 3; D020169:Caspases