Homo-oligomerization of ALS2 through its unique carboxyl-terminal regions is essential for the ALS2-associated Rab5 guanine nucleotide exchange activity and its regulatory function on endosome trafficking.
Mutations in the ALS2 gene have been known to account for a juvenile recessive form of amyotrophic lateral sclerosis (ALS2), a rare juvenile recessive form of primary lateral sclerosis, and a form of hereditary spastic paraplegia (HSP), indicating that the ALS2 protein is essential for the maintenance of motor neurons. Recently, we have demonstrated that the ALS2 protein specifically binds to the small GTPase Rab5 and acts as a GEF (guanine nucleotide exchange factor) for Rab5. We have also shown that its Rab5GEF-requisite domain resides within the C-terminal 640-amino acid region spanning membrane occupation and recognition nexus motifs and the vacuolar protein sorting 9 domain. Transiently expressed ALS2 localized onto early endosomal compartments and stimulated endosome fusions in neuronal and non-neuronal cells in an Rab5GEF activity-dependent manner. These results indicate that the C-terminal region of ALS2 plays a crucial role in endosomal dynamics by its Rab5GEF activity. Here we delineate a molecular feature of the ALS2-associated function through the C-terminal region-mediated homo-oligomerization. A yeast two-hybrid screen for interacting proteins with the ALS2 C-terminal portion identified ALS2 itself. ALS2 forms a homophilic oligomer through its distinct C-terminal regions. This homo-oligomerization is crucial for the Rab5GEF activity in vitro and the ALS2-mediated endosome enlargement in the cells. Taken together, these results indicate that oligomerization of the ALS2 protein is one of the fundamental features for its physiological function involving endosome dynamics in vivo.
Kunita
Ryota
R
Solution Oriented Research for Science and Technology, Japan Science and Technology Agency, (Tokai University School of Medicine), Kanagawa, Japan.
Otomo
Asako
A
Mizumura
Hikaru
H
Suzuki
Kyoko
K
Showguchi-Miyata
Junko
J
Yanagisawa
Yoshiko
Y
Hadano
Shinji
S
Ikeda
Joh-E
JE
D020816:Amino Acid Motifs; D000818:Animals; D015153:Blotting, Western; D019556:COS Cells; D002462:Cell Membrane / Q000378:metabolism; D002850:Chromatography, Gel; D019281:Dimerization; D011992:Endosomes / Q000378:metabolism*; D020662:Guanine Nucleotide Exchange Factors / Q000737:chemistry* / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007150:Immunohistochemistry; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D009474:Neurons / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D014162:Transfection; D020798:Two-Hybrid System Techniques; D014617:Vacuoles / Q000378:metabolism; D020696:rab5 GTP-Binding Proteins / Q000378:metabolism*
C438187:ALS2 protein, human; D020662:Guanine Nucleotide Exchange Factors; D020696:rab5 GTP-Binding Proteins