Rap1GAP Interacts With RET And Negatively Regulates GDNF Downstream Signaling Via Retarding RET Internalization

The neurotrophin receptor PET plays critical roles in the nervous system by recruiting signaling molecules that activate pathways required for the growth,survival and differentiation of neurons.It's recently knew RET receptor initiate cell body responses, like other RTKs,through internalization and transport of a ligand-receptor complex. However,the molecules mechanism for regulation this significant process are largely unknown.In this study,we identified a new interaction between PET and the adapter protein Rap1GAP,which is a GTPase-activating protein(GAP) for Rap1. Coimmunoprecipitation and coimmunostaining demonstrated endogenous Rap1GAP interaction with PET that is increased by GDNF exposure.Moreover,through gain or loss of Rap1GAP gene expression,functional analysis indicated that Rap1GAP suppressed GDNF-induced neurite outgrowth in both PC12-GFRα1-PET cells(that stably overexpress GDNF receptor GFRα1 and RET) and cultured mesencephalic neurons.More over, mechanism study performed from two way,firstly proved Rap1GAP could inactive downstream Rap1/ERK,the other and more importantly provided a novel role of Rap1GAP,which could prevent PET internalizing into cell body.Further more, Rap1GAP(N290A) mutant that are lose of GAP catalytic activity were recited to separate these two mechanism,experiment data show there is weakly different between Rap1GAP mutant and wild type Rap1GAP for PET signaling as well as GDNF-induced cell neurodiffiferenfiation,indicating Rap1GAP associated with PET and suppress GDNF's downstream independent GAP catalytic activity.In conclusion,our study is the first to demonstrate a role of Rap1GAP in signaling of RTKs and indicates a first molecular for mediating PET internalization.