title: Proper cytoskeletal architecture beneath the plasma membrane of red blood cells requiresTtll4.
abstract: Mammalian red blood cells (RBCs) circulate through blood vessels, including capillaries, for tens of days under high mechanical stress. RBCs tolerate this mechanical stress while maintaining their shape because of their elastic membrane skeleton. This membrane skeleton consists of spectrin-actin lattices arranged as quasi-hexagonal units beneath the plasma membrane. In this study, we found that the organization of the RBC cytoskeleton requires tubulin tyrosine ligase-like 4 (Ttll4). RBCs fromTtll4-knockout mice showed larger average diameters in smear test. Based on the rate of hemolysis,Ttll4-knockout RBCs showed greater vulnerability to phenylhydrazine-induced oxidative stress than did wild-type RBCs. Ultrastructural analyses revealed the macromolecular aggregation of cytoskeletal components in RBCs ofTtll4-knockout mice. Immunoprecipitation using the anti-glutamylation antibody GT335 revealed nucleosome assembly protein 1 (NAP1) to be the sole target of TTLL4 in the RBCs, and NAP1 glutamylation was completely lost inTtll4-knockout RBCs. In wild-type RBCs, the amount of glutamylated NAP1 in the membrane was nearly double that in the cytosol. Furthermore, the absence of TTLL4-dependent glutamylation of NAP1 weakened the binding of NAP1 to the RBC membrane. Taken together, these data demonstrate thatTtll4is required for proper cytoskeletal organization in RBCs.
authors.affiliation: Department of Cellular and Molecular Anatomy and International Mass Imaging Center.
authors.forename: Faryal
authors.initials: F
authors.lastname: Ijaz
authors.affiliation: Mitsubishi Kagaku Institute of Life Sciences, Tokyo 194-8511, Japan.
authors.forename: Yasue
authors.initials: Y
authors.lastname: Hatanaka
authors.affiliation: Mitsubishi Kagaku Institute of Life Sciences, Tokyo 194-8511, Japan.
authors.forename: Takahiro
authors.initials: T
authors.lastname: Hatanaka
authors.affiliation: Department of Cellular and Molecular Anatomy and International Mass Imaging Center.
authors.forename: Koji
authors.initials: K
authors.lastname: Tsutsumi
authors.affiliation: Department of Pharmacology, Hamamatsu University School of Medicine, Hamamatsu, Shizuoka 431-3192, Japan.
authors.forename: Takayuki
authors.initials: T
authors.lastname: Iwaki
authors.affiliation: Department of Pharmacology, Hamamatsu University School of Medicine, Hamamatsu, Shizuoka 431-3192, Japan.
authors.forename: Kazuo
authors.initials: K
authors.lastname: Umemura
authors.affiliation: Department of Cellular and Molecular Anatomy and International Mass Imaging Center kikegami@hama-med.ac.jp setou@hama-med.ac.jp.|Mitsubishi Kagaku Institute of Life Sciences, Tokyo 194-8511, Japan.
authors.forename: Koji
authors.initials: K
authors.lastname: Ikegami
authors.affiliation: Department of Cellular and Molecular Anatomy and International Mass Imaging Center kikegami@hama-med.ac.jp setou@hama-med.ac.jp.|Mitsubishi Kagaku Institute of Life Sciences, Tokyo 194-8511, Japan.|Preeminent Medical Photonics Education & Research Center, Hamamatsu University School of Medicine, Hamamatsu, Shizuoka 431-3192, Japan.|Riken Center for Molecular Imaging Science, Kobe, Hyogo 650-0047, Japan.|Department of Anatomy, University of Hong Kong, Hong Kong.|Division of Neural Systematics, National Institute for Physiological Sciences, Okazaki, Aichi 444-8585, Japan.
authors.forename: Mitsutoshi
authors.initials: M
authors.lastname: Setou
mesh_terms: D008841:Actin Cytoskeleton / Q000378:metabolism; D000199:Actins / Q000378:metabolism; D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D003599:Cytoskeleton / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004912:Erythrocytes / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D056490:Nucleosome Assembly Protein 1; D010453:Peptide Synthases / Q000378:metabolism* / Q000502:physiology*; D013049:Spectrin
chemical_list: D000199:Actins; D056490:Nucleosome Assembly Protein 1; D013049:Spectrin; D010453:Peptide Synthases; C112698:tubulin polyglutamylase