title: Nuclear localization of peptidylarginine deiminase V and histone deimination in granulocytes.
abstract: Peptidylarginine deiminase (PAD) deiminates arginine residues in proteins to citrulline residues Ca(2+) dependently. There are four types of PADs, I, II, III, and V, in humans. We studied the subcellular distribution of PAD V in HL-60 granulocytes and peripheral blood granulocytes. Expression of green fluorescent protein-tagged PADs in HeLa cells revealed that PAD V is localized in the nucleus, whereas PAD I, II, and III are localized in the cytoplasm. PAD V deletion mutants indicated that the sequence residues 45-74 have a nuclear localization signal (NLS). A sequence feature of this NLS is a three-lysine residue cluster preceded by a proline residue and is not found in the three other PADs. Substitution of the lysine cluster by an alanine cluster abrogated the nuclear import activity. These results suggested that the NLS is a classical monopartite NLS. HL-60 granulocytes, neutrophils, and eosinophils stained with antibody specific for PAD V exhibited distinct positive signals in the nucleus. Subcellular fractionation of HL-60 granulocytes also showed the nuclear localization of the enzyme. When neutrophils were stimulated with calcium ionophore, protein deimination occurred in the nucleus. The major deiminated proteins were identified as histones H2A, H3, and H4. The implication of PAD V in histone modifications is discussed.
authors.affiliation: Graduate School of Integrated Science, Yokohama City University, 22-2 Seto, Kanazawa-ku, Yokohama 236-0027, Japan.
authors.forename: Katsuhiko
authors.initials: K
authors.lastname: Nakashima
authors.forename: Teruki
authors.initials: T
authors.lastname: Hagiwara
authors.forename: Michiyuki
authors.initials: M
authors.lastname: Yamada
mesh_terms: D000409:Alanine / Q000737:chemistry; D000595:Amino Acid Sequence; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D017353:Gene Deletion; D006098:Granulocytes / Q000201:enzymology* / Q000378:metabolism; D049452:Green Fluorescent Proteins; D018922:HL-60 Cells; D006367:HeLa Cells; D006657:Histones / Q000378:metabolism*; D006801:Humans; D006867:Hydrolases / Q000096:biosynthesis* / Q000737:chemistry; D015151:Immunoblotting; D007150:Immunohistochemistry; D007476:Ionophores / Q000494:pharmacology; D008164:Luminescent Proteins / Q000378:metabolism; D008239:Lysine / Q000737:chemistry; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D009154:Mutation; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D000080002:Protein-Arginine Deiminase Type 4; D000076342:Protein-Arginine Deiminases; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism; D014162:Transfection
chemical_list: D006657:Histones; D007476:Ionophores; D008164:Luminescent Proteins; D011994:Recombinant Proteins; D049452:Green Fluorescent Proteins; D000001:Calcimycin; D006867:Hydrolases; C000629235:PADI4 protein, human; D000080002:Protein-Arginine Deiminase Type 4; D000076342:Protein-Arginine Deiminases; D008239:Lysine; D000409:Alanine; D002118:Calcium