Deimination of arginine residues in nucleophosmin/B23 and histones in HL-60 granulocytes.
Peptidylarginine deiminases (PADs) convert arginine residues in proteins into citrulline residues Ca(2+)-dependently. PAD V was recently found in granulocyte-differentiated HL-60 cells. To find a target of PAD V, we incubated HL-60 granulocytes with the calcium ionophore A23187 and studied deiminated proteins by immunocytochemistry and immunoblotting using a monospecific antibody to modified citrulline residues. Immunocytochemical signals were found in the nucleus upon incubation with A23187. Immunoblotting indicated that 40-, 18-, 17-, and 14-kDa proteins were preferentially deiminated. The 40-kDa protein, which was focused to pI 5.0 on two-dimensional gel electrophoresis, was identified as nucleophosmin/B23 by mass spectrometry. The 18-, 17-, and 14-kDa proteins extracted with 0.4 N H(2)SO(4) comigrated with histones H3, H2A, and H4, respectively, on two-dimensional gel electrophoresis specialized for histones. The citrulline content of histones amounted to about 10% of the histone molecules. We discuss the implications of deimination of these proteins for their nuclear functions.
Hagiwara
Teruki
T
Graduate School of Integrated Science, Yokohama City University, 22-2, Seto, Kanazawa-ku, Yokohama, 236-0027, Japan.
Nakashima
Katsuhiko
K
Hirano
Hisashi
H
Senshu
Tatsuo
T
Yamada
Michiyuki
M
D001120:Arginine / Q000378:metabolism*; D000001:Calcimycin / Q000494:pharmacology; D002956:Citrulline / Q000032:analysis / Q000276:immunology; D015180:Electrophoresis, Gel, Two-Dimensional; D006098:Granulocytes / Q000378:metabolism* / Q000648:ultrastructure; D018922:HL-60 Cells; D006657:Histones / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D006867:Hydrolases / Q000378:metabolism; D015151:Immunoblotting; D007150:Immunohistochemistry; D007476:Ionophores / Q000494:pharmacology; D007700:Kinetics; D009687:Nuclear Proteins / Q000737:chemistry* / Q000378:metabolism*; D000090243:Nucleophosmin; D011499:Protein Processing, Post-Translational; D000080002:Protein-Arginine Deiminase Type 4; D000076342:Protein-Arginine Deiminases
D006657:Histones; D007476:Ionophores; C000716967:NPM1 protein, human; D009687:Nuclear Proteins; D000090243:Nucleophosmin; D002956:Citrulline; D000001:Calcimycin; D001120:Arginine; D006867:Hydrolases; C000629235:PADI4 protein, human; D000080002:Protein-Arginine Deiminase Type 4; D000076342:Protein-Arginine Deiminases