VHL tumor suppressor gene : Identification its function for cell growth inhibition and cell ular apoptosis for kidne cancer to develop new modality of treatment

Our purpose in this grant is to identify specific proteins which affect tumor growth or invasion that are controlled by VHL tumor suppressor protein(PVHL)with cell biolgy and molecular biological methods.We first examined morphological change in 99VHL，lung cancer cell lines in which pVHL is specially inducible to 100-fold by ponasterone treatment。In the experiment for the induction of pVHL in that cell lines，almost of the cells are flattened and the cell adhesion ability to the cell culture dish is highly increased.The specific proteins for cellular adhesion，vinculin is increased in the adherent side of cell membrane.The total amount of cellular vinculin itself is not changed.The actin finbers are well polymerized at the same time with the movement of vinculin to the adherent membrane.The cell motitilty is dramatically decreased with these changes.According to these results，the function of pVHL is to reorganize vinculin to the adherent side of membranem，to polymerize acti…More n fiber and finally to decrease cell motility.We next examined the characteristics of germline mutation of the VHL gene in the whole world.We detected4mutation-clustered regions，that are the end of the exonl，the junction of the exon1and2，the middle portion of exon2and the first part of exon3.The first part of exon3is well known as the binding domain of pVHL with Elongin C.It is hly possible that other3regions have important function。As the results of several experiments，the 2^&lt；nd&gt；clustered region of mutation coinsides with the binding domain of pVHL with atyipcal protein kinase lambda(aPKC···)。This resion is located at the beta-domain of pVHL(Amine Acid114to122)。Since the essential function of pVHL is ubiqutination and final degradation of short-lived proteins，that bind the beta domain，one of the important candidate protein for function of pVHL is aPKC···Since aPKC···has·anti-apoptotic function and promote cell growth，pVHL is working to degrade aPKC…and induce cellular apoptosis.Our purpose in this grant is to identify specific protes to degrade aPKC…and induce cellular apoptosis.Our purpose in this grant is to identify specific protes to degrade aPKC…and induce cellular apoptosis.Our purpose in this grantWe first examined morphological change in 99 VHL.lung cancer cell lintes in which pVHL is specially inducible to 100-fold by ponasterone treatment.In the experiment for the induction of pVHL in that cell lines，almost of the cells are flattened and the cell adhesion ability to the cell culture dish is higltly increased.The specific proteins for cellular adhesion，vinculin is increased in the adherent side of cell membrane.The total amount of cellular vinculin itself is not changed.The actin finbers are well polymerized at the same time with the movement of vinculin to the adherent membrane.The cell motitilty is dramatically decreased with these changes.According to these results，the function of pVHL is to reorganize vinculin to the adherent side of membranem，to polymerize actin fiber and finally to decrease cell motility.As a summary，it is shown that the direct effect of pVHL is possible aPKC·degradation and indirect effect is to reorganize vinculin，to polymerize actin fiber and finally to reduce cell motility.Less：Less

Our purpose in this grant is to identify specific proteins which affect tumor growth or invasion that are controlled by VHL tumor suppressor protein (pVHL) with cell biolgy and molecular biological methods. We first examined morphological change in 99VHL, lung cancer cell lines in which pVHL is specially inducible to 100-fold by ponasterone treatment. In the experiment for the induction of pVHL in that cell lines, almost of the cells are flattened and the cell adhesion ability to the cell culture dish is highly increased. The specific proteins for cellular adhesion, vinculin is increased in the adherent side of cell membrane. The total amount of cellular vinculin itself is not changed. The actin finbers are well polymerized at the same time with the movement of vinculin to the adherent membrane. The cell motitilty is dramatically decreased with these changes. According to these results, the function of pVHL is to reorganize vinculin to the adherent side of membranem, to polymerize acti … More n fiber and finally to decrease cell motility.We next examined the characteristics of germline mutation of the VHL gene in the whole world. We detected 4 mutation-clustered regions, that are the end of the exonl, the junction of the exon 1 and 2, the middle portion of exon 2 and the first part of exon 3.The first part of exon 3 is well known as the binding domain of pVHL with Elongin C.It is highly possible that other 3 regions have important function. As the results of several experiments, the 2^<nd> clustered region of mutation coinsides with the binding domain of pVHL with atyipcal protein kinase lambda (aPKC・・). This resion is located at the beta-domain of pVHL (amine acid 114 to 122). Since the essential function of pVHL is ubiqutination and final degradation of short-lived proteins, that bind the beta domain, one of the important candidate protein for function of pVHL is aPKC・・・ Since aPKC・・・・ has ・anti-apoptotic function and promote cell growth, pVHL is working to degrade aPKC・・・ and induce cellular apoptosis.Our purpose in this grant is to identify specific proteins which affect tumor growth or invasion that are controlled by VIIL tumor suppressor protein (pVHL) with cell biolgy and molecular biological methods. We first examined morphological change in 99VHL.lung cancer cell lintes in which pVHL is specially inducible to 100-fold by ponasterone treatment. In the experiment for the induction of pVHL in that cell lines, almost of the cells are flattened and the cell adhesion ability to the cell culture dish is higltly increased. The specific proteins for cellular adhesion, vinculin is increased in the adherent side of cell membrane. The total amount of cellular vinculin itself is not changed. The actin finbers are well polymerized at the same time with the movement of vinculin to the adherent membrane. The cell motitilty is dramatically decreased with these changes. According to these results, the function of pVHL is to reorganize vinculin to the adherent side of membranem, to polymerize actin fiber and finally to decrease cell motility.As a summary, it is shown that the direct effect of pVHL is possible aPKC・・ degradation and indirect effect is to reorganize vinculin, to polymerize actin fiber and finally to reduce cell motility. Less