Application of TEM: Dynamic changes of AKAP95-Cx43 complex during G1 phase of lung cancer cells

Abstract BackgroundAKAP95(A-kinase anchoring protein) and Cx43 (connexin 43) express abnormally in lung cancer cells. As potential tumor therapeutic targets, specific process of bindings and dynamic changes of Cx43-AKAP95 complex in lung cancer cells may guide further treatments and detections of lung cancer. However, the process remains unclear now. We are aiming at investigating the dynamic changes of expression, localization, and binding of AKAP95 and Cx43, as well as their interaction with cyclin D1 and cyclin E1 in lung cancer cells during G1 phase.MethodsA549 and Beas-2B cells were arrested at preliminary stage(P), middle stage(M) and restriction point(R) of G1 phase and Western blot(WB), Confocal laser scanning microscopy (CLSM) and Transmission electron microscope (TEM) were used in our study to detect proteins to provide further evidence of the correlation of these proteins.Results: 1) AKAP95 carries Cx43 into the nucleus through the nuclear pore by binding to it during G1 phase. Some AKAP95 and Cx43 can aggregate and form larger protein aggregates. The process happens mainly during R stage. 2) AKAP95 and Cx43 mainly bind to cyclin D1 during P and M stage while bind to cyclin E1 during R stage respectively. Complexes of AKAP95-cyclin D1 and Cx43-cyclin D1 cannot enter nucleus while AKAP95-cyclin E1 and Cx43-cyclin E1 can.Conclusions1) Binding process of AKAP95-Cx43 complexes/ aggregates can be summarize as ‘bind and aggregate -target and enter nucleus- keep binding/aggregating in nucleus’. 2) Cyclin E1 was involved in the binding/aggregation and nuclear entry of AKAP95 and Cx43 while cyclin D1 only binds to them respectively in the cytoplasm.