title: Mammalian ESCRT-III-related protein IST1 has a distinctive met-pro repeat sequence that is essential for interaction with ALG-2 in the presence of Ca2+.
abstract: ALG-2 is an EF-hand-type Ca(2+)-binding protein that interacts with a variety of intracellular proteins that possess Pro-rich regions (PRRs) in mammalian cells. IST1 is an endosomal sorting complex required for transport (ESCRT)-III-related charged multivesicular body protein (CHMP)-like protein, but unlike other ESCRT-III proteins, mammalian IST1 has a PRR and a distinctive sequence of Met-Pro repeats. We found that ALG-2 binds to IST1 by Far-Western analysis using biotinylated ALG-2 as probe, and that the Met-Pro repeat sequence is essential for interaction. The results of pulldown assays using Strep-tagged ALG-2 and lysates of cells expressing GFP-fused IST1 proteins indicated that the binding of ALG-2 to IST1 is Ca(2+)-dependent, and that it is enhanced by co-expression with CHMP1 proteins. Moreover, pulldown assays using various mutants of GST-ALG-2 revealed that the ability of IST1 to bind to mutants is different from those of known ALG-2-interacting proteins, suggesting that IST1 binds to ALG-2 by a different mode of recognition.
authors: Department of Applied Molecular Biosciences, Graduate School of Bioagricultural Sciences, Nagoya University, Furo-cho, Chikusa-ku, Nagoya, Japan.
authors: Mayumi
authors: M
authors: Okumura
authors: Takeshi
authors: T
authors: Takahashi
authors: Hideki
authors: H
authors: Shibata
authors: Masatoshi
authors: M
authors: Maki
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D051017:Apoptosis Regulatory Proteins / Q000235:genetics / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D056827:Endosomal Sorting Complexes Required for Transport / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D015513:Oncogene Proteins / Q000737:chemistry* / Q000378:metabolism*; D011485:Protein Binding; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D051381:Rats; D020449:Repetitive Sequences, Amino Acid
chemical_list: D051017:Apoptosis Regulatory Proteins; D002135:Calcium-Binding Proteins; D056827:Endosomal Sorting Complexes Required for Transport; C533705:IST1 protein, human; D015513:Oncogene Proteins; C097753:PDCD6 protein, human; D020033:Protein Isoforms; D002118:Calcium