title: Rabaptin-5-independent membrane targeting and Rab5 activation by Rabex-5 in the cell.
abstract: Rabex-5 is a guanine nucleotide exchange factor (GEF) for Rab5. Here, we report the identification of a novel functional domain of Rabex-5 that is essential for its membrane targeting and Rab5 GEF activity in vivo. The data show that full-length Rabex-5 efficiently activates Rab5 in the cell. However, the GEF domain itself (residues 135-399) is inactive in this respect, despite its activity in vitro. Generation and characterization of a series of Rabex-5 constructs reveal that the GEF domain is unable to target to early endosomes and that a sequence N-terminal to the GEF domain can restore its early endosomal targeting and its ability to activate Rab5 in the cell. This region (residues 81-135) is termed membrane-binding motif, which together with the downstream helical bundle domain (residues 135-230) forms an early endosomal targeting (EET) domain necessary and sufficient for association with early endosomes. Furthermore, several active Rabex-5 constructs do not contain the Rabaptin-5-binding domain in the C-terminal region. Thus, Rabex-5 can target to early endosomes via the EET domain and activate Rab5 in a Rabaptin-5-independent manner in vivo. We discuss a model to reconcile these in vivo data with previous in vitro results on Rabex-5 function and its interaction with Rabaptin-5.
authors: Department of Biochemistry and Molecular Biology, University of Oklahoma Health Sciences Center, Oklahoma City, OK 73104, USA.
authors: Huaiping
authors: H
authors: Zhu
authors: Guangyu
authors: G
authors: Zhu
authors: Jay
authors: J
authors: Liu
authors: Zhimin
authors: Z
authors: Liang
authors: Xuejun C
authors: XC
authors: Zhang
authors: Guangpu
authors: G
authors: Li
mesh_terms: D000818:Animals; D002417:Cattle; D002462:Cell Membrane / Q000378:metabolism*; D006224:Cricetinae; D011992:Endosomes / Q000378:metabolism; D004789:Enzyme Activation; D020662:Guanine Nucleotide Exchange Factors / Q000737:chemistry / Q000378:metabolism*; D008561:Membrane Fusion; D008954:Models, Biological; D050505:Mutant Proteins / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D033921:Vesicular Transport Proteins / Q000737:chemistry / Q000378:metabolism; D020696:rab5 GTP-Binding Proteins / Q000378:metabolism*
chemical_list: D020662:Guanine Nucleotide Exchange Factors; D050505:Mutant Proteins; D033921:Vesicular Transport Proteins; D020696:rab5 GTP-Binding Proteins
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