title: Alsin is a Rab5 and Rac1 guanine nucleotide exchange factor.
abstract: ALS2 is the gene mutated in a recessive juvenile form of amyotrophic lateral sclerosis (ALS2). ALS2 encodes a large protein termed alsin, which contains a number of predicted cell signaling and protein trafficking sequence motifs. To gain insight into the overall function of alsin and to begin to evaluate its role in motor neuron maintenance, we examined the subcellular localization of alsin and the biochemical activities associated with its individual subdomains. We found that the Vps9p domain of alsin has Rab5 guanine nucleotide exchange activity. In addition, alsin interacted specifically with and acted as a guanine nucleotide exchange factor for Rac1. Immunofluorescence and fractionation experiments in both fibroblasts and neurons revealed that alsin is a cytosolic protein, with a significant portion associated with small, punctate membrane structures. Many of these membrane structures also contained Rab5 or Rac1. Upon overexpression of full-length alsin, the overexpressed material was largely cytosolic, indicating that the association with membrane structures could be saturated. We also found that alsin was present in membrane ruffles and lamellipodia. These data suggest that alsin is involved in membrane transport events, potentially linking endocytic processes and actin cytoskeleton remodeling.
authors.affiliation: Department of Biochemistry and Molecular Biology, Mayo Clinic, Rochester, Minnesota 55905, USA.
authors.forename: Justin D
authors.initials: JD
authors.lastname: Topp
authors.forename: Noah W
authors.initials: NW
authors.lastname: Gray
authors.forename: Robert D
authors.initials: RD
authors.lastname: Gerard
authors.forename: Bruce F
authors.initials: BF
authors.lastname: Horazdovsky
mesh_terms: D000199:Actins / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002531:Cerebellum / Q000378:metabolism; D003599:Cytoskeleton / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D011992:Endosomes / Q000378:metabolism; D005260:Female; D049452:Green Fluorescent Proteins; D020662:Guanine Nucleotide Exchange Factors / Q000378:metabolism* / Q000502:physiology*; D006801:Humans; D015151:Immunoblotting; D007313:Insecta; D008164:Luminescent Proteins / Q000378:metabolism; D051379:Mice; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D041681:NIH 3T3 Cells; D009474:Neurons / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D011554:Pseudopodia / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction; D013347:Subcellular Fractions; D014162:Transfection; D020798:Two-Hybrid System Techniques; D020696:rab5 GTP-Binding Proteins / Q000378:metabolism*; D020744:rac GTP-Binding Proteins / Q000378:metabolism; D020830:rac1 GTP-Binding Protein / Q000378:metabolism*
chemical_list: C438187:ALS2 protein, human; D000199:Actins; C512358:Als2 protein, mouse; D020662:Guanine Nucleotide Exchange Factors; D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins; D020696:rab5 GTP-Binding Proteins; D020744:rac GTP-Binding Proteins; D020830:rac1 GTP-Binding Protein