| E-cadherin mediates adhesion between cells, while integrins mediates adhesion between cells and ECM. Tumor cells detach from the primary tumor tissue, metasising to distal sites and adhesing to distal organs, dissolving the ECM and forming new vascules. This serial precesses should be accompanied the cross-talks between E-cadherin and integrins. The cross-talks between E-cadherin and integrins should also take active part in normal cells and the embryo development process. In this experiment, wild-typed E-cadherin cDNA construct which can be expressed in eukaryotic cells, was transfected into two E-cadherin-negative breast carcinoma cell lines, MDA-MB-231 and MDA-MB-435. Positive transfectants were selected by G418 and named as E-cad-231 and E-cad-435, correspondingly. At the same time, pcDNA3, the empty vector, was also transfected and selected by G418 and named as Mock-231 and Mock-435 as negative control. E-cad-435 and E-cad-231 adhesion ability to fibronectin and laminin decreased compared with the control cells, ewpecially fibronectin. Integrin α5β1 is the typical receptor of fibronectin. Flow cytometry, Western blot and RT-PCR showed that bothα5 and β1 integrins protein level and mRNA decreased compared with the control cells. In negative cells, β-catenin accumulated in the nucleus, while β-catenin nearly dispeared from the necleus and mainly accumulated near the membrane and cytosol. Since β-catenin is one of important participants in Wnt pathway, binding with mombers of TCF/LEF family and initiating or inhibiting transcription of many genes. In order to cerfiticate whether E-cadherin Could influence integrins through β-catenin, LiCl treatement was used to inhibit β-catenin protein degradation. After LiCl treatment, β-catenin and integrin α5 and β1 protein level increased. It suggested that E-cadherin could influence integrinα5β1 expression through β-catenin. Furthermore, positive expression of E-cadherin could inhibit integrin downstream signal molecules, such as FAK,PKB and ILK and a cell cycle regulator, cyclinD1. LiCl treatment could increase FAK and PKB protein level, which suggested that E-cadherin could inhibit FAK and PKB.Δ71, mutant E-cadherin cDNA which lacks the core sequence binding with β-catenin, was also transfected into the two cell lines mention above. G418 was used to select the positive transfectants. Δ71, the mutant E-cadherin, could bind with β-catenin, but could still decrease β-catenin protein level and relocate β-catenin from mainly nucleus tocytosol. The results showed that E-cadherin could decrease β-catenin protein level and relocate β-catenin without the help of direct binding.Further research showed thatΔ71 could inhibit bothα5 and β1 integrin protein levels, especially in MDA-MB-435, in which positive expression ofΔ71 could inhibit mostα5 integrin expression. This result was consistent with other results and furtherly confirmed that E-cadherin could inhibit bothα5 and β1 integrin, but was not fulfilled by simply binding with β-catenin and influencing its location.It has been reported that PKB can phosphorylate GSK-3 and inhibit β-catenin protein degradation. Wortmannin is an inhibitor of PI3K. With the increase of Wortmannin concentration, Phosphorylation of PKB 473-threnine and β-catenin protein level decreased. It seemed to mean that β-catenin protein degradation system was intact and β-catenin accumulated in the nucleus was due to inhibition of high protein level of PKB in both MDA-MB-231 and MDA-MB-435. β-catenin and PKB protected each other or promote the other's expression in a loop. In addition, positive expression of E-cadherin could inhibit the estrogen secretion ability in the human carcinoma cells. It has been reported that estrogen could active PKB through non-receptor pathway, such as GPCR30 and influence integrins expression. This provided another explanation for how E-cadherin influences integrin pathway. Above all, the positive expression of E-cadherin in human breast carcinoma cells not only binded with ?...
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