Activation of the Small G Protein Arf6 by Dynamin2 through Guanine Nucleotide Exchange Factors in Endocytosis.
The small G protein Arf6 and the GTPase dynamin2 (Dyn2) play key roles in clathrin-mediated endocytosis (CME). However, their functional relationship remains obscure. Here, we show that Arf6 functions as a downstream molecule of Dyn2 in CME. Wild type of Dyn2 overexpressed in HeLa cells markedly activates Arf6, while a GTPase-lacking Dyn2 mutant does not. Of the Arf6-specific guanine nucleotide exchange factors, EFA6A, EFA6B, and EFA6D specifically interact with Dyn2. Furthermore, overexpression of dominant negative mutants or knockdown of EFA6B and EFA6D significantly inhibit Dyn2-induced Arf6 activation. Finally, overexpression of the binding region peptide of EFA6B for Dyn2 or knockdown of EFA6B and EFA6D significantly suppresses clathrin-mediated transferrin uptake. These results provide evidence for a novel Arf6 activation mechanism by Dyn2 through EFA6B and EFA6D in CME in a manner dependent upon the GTPase activity of Dyn2.
Department of Physiological Chemistry, Faculty of Medicine and Graduate School of Comprehensive Human Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba 305-8575, Japan.
Risa
R
Okada
Department of Physiological Chemistry, Faculty of Medicine and Graduate School of Comprehensive Human Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba 305-8575, Japan.
Yohei
Y
Yamauchi
Department of Physiological Chemistry, Faculty of Medicine and Graduate School of Comprehensive Human Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba 305-8575, Japan.
Tsunaki
T
Hongu
Department of Physiological Chemistry, Faculty of Medicine and Graduate School of Comprehensive Human Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba 305-8575, Japan.
Yuji
Y
Funakoshi
Department of Physiological Chemistry, Faculty of Medicine and Graduate School of Comprehensive Human Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba 305-8575, Japan.
Norihiko
N
Ohbayashi
Department of Physiological Chemistry, Faculty of Medicine and Graduate School of Comprehensive Human Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba 305-8575, Japan.
Hiroshi
H
Hasegawa
Department of Physiological Chemistry, Faculty of Medicine and Graduate School of Comprehensive Human Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba 305-8575, Japan.
Yasunori
Y
Kanaho
D000089982:ADP-Ribosylation Factor 6; D020727:ADP-Ribosylation Factors / Q000378:metabolism*; D001665:Binding Sites; D034285:Dynamin II / Q000502:physiology*; D004705:Endocytosis / Q000502:physiology*; D020662:Guanine Nucleotide Exchange Factors / Q000502:physiology*; D006367:HeLa Cells; D006801:Humans; D009419:Nerve Tissue Proteins / Q000378:metabolism
D000089982:ADP-Ribosylation Factor 6; D020662:Guanine Nucleotide Exchange Factors; D009419:Nerve Tissue Proteins; C109970:PSD protein, human; D020727:ADP-Ribosylation Factors; C000715348:ARF6 protein, human; D034285:Dynamin II