Role of Ceveolae on Translocation of Glucose Transporter

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I made GFP fusion protein to have full length of wild type Caveolin-1(human) which was the protein which constituted caveolae toward and transfected into a 3T3-L1 cell and I got selected this by antibiotics stably expressed cells. I was able to get GFP fusion proteins stably expressed cells to have N terminal portion deleted type of this protein. In addition, N terminal deletion type of Caveolin-3 and Caveolin-1 DNA in two places of amino acids mutation of apart of scaffolding domain of Caveolin-1.About a glucose transporter, I compared reaction for insulin stimulation after having guided fat cell differentiation with the 3T3-L1 cell which I transfected a GLUT4-GFP fusion type. As the factor which restrained the formation of caveolae, I transiently tranfected a N terminal portion deleted type (DGV) of Caveolin-3. As the factor which restrained the clathrin formation, I tranfected a C terminal part of AP180 DNA. I stimulated these two cells by insulin. As a result, a translocation of GLUT4 to a cell membrane was restrained by both protein expressions. This got a conclusion that both existence of clathrin coated vesicles and caveolae was restrained in a translocation to a cell membrane of a glucose transporter.I examined a translocation and recycling of aquaporin-2 to compare it with a translocation of GLUT4. I understood that a translocation of aquaporin-2 was obstructed by functional restraint of raft caused by a defect of glycosphingolipid. Furthermore, after recycling, aquaporin retrieved endosomes with Caveolin-1. I understood what was obstructed return to this cell when I transfected caveolin-1 which had variation into two places of amino acids of a part of scaffolding domain which restrained the formation of caveolae.