title: Identification of a novel role for dematin in regulating red cell membrane function by modulating spectrin-actin interaction.
abstract: The membrane skeleton plays a central role in maintaining the elasticity and stability of the erythrocyte membrane, two biophysical features critical for optimal functioning and survival of red cells. Many constituent proteins of the membrane skeleton are phosphorylated by various kinases, and phosphorylation of β-spectrin by casein kinase and of protein 4.1R by PKC has been documented to modulate erythrocyte membrane mechanical stability. In this study, we show that activation of endogenous PKA by cAMP decreases membrane mechanical stability and that this effect is mediated primarily by phosphorylation of dematin. Co-sedimentation assay showed that dematin facilitated interaction between spectrin and F-actin, and phosphorylation of dematin by PKA markedly diminished this activity. Quartz crystal microbalance measurement revealed that purified dematin specifically bound the tail region of the spectrin dimer in a saturable manner with a submicromolar affinity. Pulldown assay using recombinant spectrin fragments showed that dematin, but not phospho-dematin, bound to the tail region of the spectrin dimer. These findings imply that dematin contributes to the maintenance of erythrocyte membrane mechanical stability by facilitating spectrin-actin interaction and that phosphorylation of dematin by PKA can modulate these effects. In this study, we have uncovered a novel functional role for dematin in regulating erythrocyte membrane function.
authors.affiliation: Department of Biochemistry, School of Medicine, Tokyo Women's Medical University, 8-1 Kawada-cho, Shinjuku 162-8666, Tokyo, Japan.
authors.forename: Ichiro
authors.initials: I
authors.lastname: Koshino
authors.affiliation: New York Blood Center, New York, New York 10065.
authors.forename: Narla
authors.initials: N
authors.lastname: Mohandas
authors.affiliation: Department of Biochemistry, School of Medicine, Tokyo Women's Medical University, 8-1 Kawada-cho, Shinjuku 162-8666, Tokyo, Japan. Electronic address: takakuwa@research.twmu.ac.jp.
authors.forename: Yuichi
authors.initials: Y
authors.lastname: Takakuwa
mesh_terms: D000199:Actins / Q000378:metabolism*; D047388:Casein Kinases / Q000378:metabolism; D000242:Cyclic AMP / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000378:metabolism; D004910:Erythrocyte Membrane / Q000378:metabolism*; D005260:Female; D006801:Humans; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism; D008840:Microfilament Proteins / Q000378:metabolism*; D055503:Protein Multimerization / Q000502:physiology; D017434:Protein Structure, Tertiary; D013049:Spectrin / Q000378:metabolism*
chemical_list: D000199:Actins; D003598:Cytoskeletal Proteins; C044805:DMTN protein, human; D008565:Membrane Proteins; D008840:Microfilament Proteins; C028841:erythrocyte membrane band 4.1 protein; D013049:Spectrin; D000242:Cyclic AMP; D047388:Casein Kinases; D017868:Cyclic AMP-Dependent Protein Kinases