The role of Cav1 YAP and Rac in Ha-RasV12-induced multilayer cellular aggregates of Madin-Darby canine kidney cells

  • 韓 佳霖

Student thesis: Master's Thesis

Abstract

Two mechanisms are proposed to maintain homeostasis in epithelial cells monolayer: growth arrest caused by contact inhibition and cell extrusion caused by crowding induced delamination It is highly likely that perturbation of these homeostatic systems lead to the formation of epidermal masses in cancer development In this study we showed that IPTG-inducible expression of oncogenic Ha-RasV12 in MK4 cells (a clone of MDCK cells) causes the formation of multilayer cellular aggregation The Hippo-YAP pathway has recently been shown to be an important regulator of contact inhibition Induction of Ha-RasV12 triggered YAP nuclear translocation and subsequently YAP-targeted gene expression Expression of Ha-RasV12 induced elevation of LATS1 and MST1 suggesting that canonical Hippo pathway may not be the cause of Ha-RasV12-induced activation of YAP Next we showed inhibition of p-ERK or actomyosin cytoskeleton impeded Ha-RasV12-induced activation of YAP Treatment of Verteporfin (VP) a YAP/TEAD binding inhibitor failed to abolish Ha-RasV12 induced multilayer cellular aggregates indicating the involvement of other molecules Overexpression of Cav1 inhibited Ha-RasV12 induced cellular and mechanical transformation In addition we showed that overexpression of Cav1 inhibited Ha-RasV12-induced YAP activation and multicellular cell aggregates However knockdown of Cav1 in MDCK cells only resulted in activation of YAP but not cellular aggregates Moreover Rac and RhoA both associated with cell extrusion were increased in Ha-RasV12 overexpressed MK4 cells EHT1864 (Rac inhibitor) abolished multilayer cellular aggregates in Ha-RasV12-overexpressed MK4 cells whereas Y27632 (ROCK inhibitor) induced multilayer cellular aggregates in MK4 cells Inhibition of p-ERK and overexpression of Cav1 also inhibited Ha-RasV12 induced Rac activation Furthermore overexpression of Rac1 resulted in cellular aggregates Taken together these data indicate that Ha-RasV12-induced YAP activation is not required for multicellular aggregates On the other hand Rac activation is essential for Ha-RasV12-induced multilayer cellular aggregate
Date of Award2017 Aug 22
Original languageEnglish
SupervisorMing-Jer Tang (Supervisor)

Cite this

The role of Cav1 YAP and Rac in Ha-RasV12-induced multilayer cellular aggregates of Madin-Darby canine kidney cells
佳霖, 韓. (Author). 2017 Aug 22

Student thesis: Master's Thesis