Akt/Protein Kinase B plays an important role in many biological responses including cell survival, proliferation, and nutrient metabolism. Activation of Akt/PKB by growth factors involves PIP3 binding, membrane translocation and the phosphorylation at Thr308 and Ser473 residues by PDK1 and PDK2, respectively. Dephosphorylation and AKT binding proteins were two inhibitory mechanisms for AKT. Carboxyl-terminal modulator protein (CTMP) has been demonstrated to bind to carboxyl-terminus of Akt and as a regulator on plasma membrane. However, the interaction region of CTMP to Akt is still unknown. On the other hand, contradict results of positive and negative effects to AKT activity were reported. In this study, we firstly demonstrated the CTMP protein level shows positive correlation with Akt phosphorylation and cell proliferation in human breast cancer cell. The interaction of CTMP and phosphorylation Akt by co-immunoprecipitation and immunofluorescence experiments shows that CTMP may be involved in regulating Akt phosphrylation. The results of GST pull down assay demonstrated that the direct binding and revealed more then one interaction region of CTMP to Akt. Interestingly, in HeLa cells, transiently or stably expressing CTMP can directly promote Akt phosphorylation even in basal state and significantly increase the phosphorylation at both Thr308 and Ser473 of Akt upon the insulin stimulation in the pre-starvation condition. When compared the cell growth of HeLa cells in 10% serum condition using cell proliferation and soft-agar colony forming assay, rapid cell proliferation and colony number increase were observed in cells stably expressing CTMP suggesting that CTMP may be involved in the regulation of Akt-mediated cell growth. The analogous results were also obtained from colony formation assay with different condition from 10% to 1% serum. The result of cell cycle distribution shows that the increasment of cell proliferation rate may due to the increasment of cell cycle progression in CTMP stable clones. Furthermore, CTMP stable clone promote tumor growth and metastasis in xenograft animal model. Together, these results indicated that the binding of CTMP positively modulate Akt and thus increase insulin sensitivity in HeLa cells.
| Identifer | oai:union.ndltd.org:NSYSU/oai:NSYSU:etd-0804108-223222 |
| Date | 04 August 2008 |
| Creators | Liao, Wern-chir |
| Contributors | Michael Hsiao, Yi-Ren Hong, Pei-jung Lu, Chung-Lung Cho, Jiin-Tsuey Cheng |
| Publisher | NSYSU |
| Source Sets | NSYSU Electronic Thesis and Dissertation Archive |
| Language | English |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | http://etd.lib.nsysu.edu.tw/ETD-db/ETD-search/view_etd?URN=etd-0804108-223222 |
| Rights | not_available, Copyright information available at source archive |
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