title: Structure and Regulation of the Movement of Human Myosin VIIA.
abstract: Human myosin VIIA (HM7A) is responsible for human Usher syndrome type 1B, which causes hearing and visual loss in humans. Here we studied the regulation of HM7A. The actin-activated ATPase activity of full-length HM7A (HM7AFull) was lower than that of tail-truncated HM7A (HM7AΔTail). Deletion of the C-terminal 40 amino acids and mutation of the basic residues in this region (R2176A or K2179A) abolished the inhibition. Electron microscopy revealed that HM7AFull is a monomer in which the tail domain bends back toward the head-neck domain to form a compact structure. This compact structure is extended at high ionic strength or in the presence of Ca(2+). Although myosin VIIA has five isoleucine-glutamine (IQ) motifs, the neck length seems to be shorter than the expected length of five bound calmodulins. Supporting this observation, the IQ domain bound only three calmodulins in Ca(2+), and the first IQ motif failed to bind calmodulin in EGTA. These results suggest that the unique IQ domain of HM7A is important for the tail-neck interaction and, therefore, regulation. Cellular studies revealed that dimer formation of HM7A is critical for its translocation to filopodial tips and that the tail domain (HM7ATail) markedly reduced the filopodial tip localization of the HM7AΔTail dimer, suggesting that the tail-inhibition mechanism is operating in vivo. The translocation of the HM7AFull dimer was significantly less than that of the HM7AΔTail dimer, and R2176A/R2179A mutation rescued the filopodial tip translocation. These results suggest that HM7A can transport its cargo molecules, such as USH1 proteins, upon release of the tail-dependent inhibition.
authors.affiliation: From the Department of Microbiology and Physiological Systems, University of Massachusetts Medical School, Worcester, Massachusetts 01605, the Department of Cellular and Molecular Biology, University of Texas Health Science Center at Tyler, Tyler, Texas 75708.
authors.forename: Tsuyoshi
authors.initials: T
authors.lastname: Sakai
authors.affiliation: the Division of Electron Microscopic Research, Korea Basic Science Institute, 169-148 Gwahak-ro, Daejeon 305-333, Korea, and the Department of Biochemistry, College of Natural Sciences, Kangwon National University, 1, Kangwondaehak-gil, Chuncheon-si, Gangwon-do 200-701, Korea.
authors.forename: Hyun Suk
authors.initials: HS
authors.lastname: Jung
authors.affiliation: From the Department of Microbiology and Physiological Systems, University of Massachusetts Medical School, Worcester, Massachusetts 01605, the Department of Cellular and Molecular Biology, University of Texas Health Science Center at Tyler, Tyler, Texas 75708.
authors.forename: Osamu
authors.initials: O
authors.lastname: Sato
authors.affiliation: From the Department of Microbiology and Physiological Systems, University of Massachusetts Medical School, Worcester, Massachusetts 01605.
authors.forename: Masafumi D
authors.initials: MD
authors.lastname: Yamada
authors.affiliation: the Division of Electron Microscopic Research, Korea Basic Science Institute, 169-148 Gwahak-ro, Daejeon 305-333, Korea, and.
authors.forename: Dong-Ju
authors.initials: DJ
authors.lastname: You
authors.affiliation: From the Department of Microbiology and Physiological Systems, University of Massachusetts Medical School, Worcester, Massachusetts 01605, the Department of Cellular and Molecular Biology, University of Texas Health Science Center at Tyler, Tyler, Texas 75708.
authors.forename: Reiko
authors.initials: R
authors.lastname: Ikebe
authors.affiliation: From the Department of Microbiology and Physiological Systems, University of Massachusetts Medical School, Worcester, Massachusetts 01605, the Department of Cellular and Molecular Biology, University of Texas Health Science Center at Tyler, Tyler, Texas 75708, Mitsuo.Ikebe@uthct.edu.
authors.forename: Mitsuo
authors.initials: M
authors.lastname: Ikebe
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D002147:Calmodulin / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D046529:Microscopy, Electron, Transmission; D020409:Molecular Motor Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D000081412:Myosin VIIa; D009218:Myosins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D055503:Protein Multimerization; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017384:Sequence Deletion; D017386:Sequence Homology, Amino Acid; D052245:Usher Syndromes / Q000235:genetics / Q000378:metabolism
chemical_list: D002147:Calmodulin; C000631455:MYO7A protein, human; D020409:Molecular Motor Proteins; D000081412:Myosin VIIa; D011993:Recombinant Fusion Proteins; D009218:Myosins
keywords: ATPase; Usher syndrome; actin; calmodulin (CaM); filopodia; intracellular trafficking; molecular motor; myosin; myosin VIIA