Human Peptidylarginine Deiminase Type III: Molecular Cloning and Nucleotide Sequence of the cDNA, Properties of the Recombinant Enzyme, and Immunohistochemical.

Slides:

Advertisements

Similar presentations

Highly Complex Peptide Aggregates of the S100 Fused-Type Protein Hornerin Are Present in Human Skin Zhihong Wu, Ulf Meyer-Hoffert, Katrin Reithmayer,

Advertisements

Volume 32, Issue 1, Pages (October 2008)

Crucial Roles of MZF1 and Sp1 in the Transcriptional Regulation of the Peptidylarginine Deiminase Type I Gene (PADI1) in Human Keratinocytes Sijun Dong,

Characterization of an ADAMTS-5-mediated cleavage site in aggrecan in OSM- stimulated bovine cartilage M. Durigova, M.Sc., P. Soucy, B.Sc., K. Fushimi,

Skin-Specific Expression of ank-393, a Novel Ankyrin-3 Splice Variant

Gene Expression of Mouse S100A3, a Cysteine-Rich Calcium-Binding Protein, in Developing Hair Follicle Kenji Kizawa, Suguru Tsuchimoto, Keiko Hashimoto,

A Novel Human Type II Cytokeratin, K6hf, Specifically Expressed in the Companion Layer of the Hair Follicle Hermelita Winter, Martina Jacobs, Michael.

Characterization of Mouse Frizzled-3 Expression in Hair Follicle Development and Identification of the Human Homolog in Keratinocytes Betsy S. Hung,

Peroxiredoxin is Ubiquitously Expressed in Rat Skin: Isotype-Specific Expression in the Epidermis and Hair Follicle Jeong Eun Lee, Byung Dae Kwon, Jee-Bum.

Peptidylarginine Deiminase Isoforms 1–3 Are Expressed in the Epidermis and Involved in the Deimination of K1 and Filaggrin Rachida Nachat, Marie-Claire.

Basement Membrane Zone Remodeling During Appendageal Development in Human Fetal Skin. The Absence of Type VII Collagen is Associated with Gelatinase-A.

HKAP1.6 and hKAP1.7, Two Novel Human High Sulfur Keratin-Associated Proteins are Expressed in the Hair Follicle Cortex Yutaka Shimomura, Noriaki Aoki,

Transcription Factors C/EBPα, C/EBPβ, and CHOP (Gadd153) Expressed During the Differentiation Program of Keratinocytes In Vitro and In Vivo Edward V.

Overexpression of Serpin Squamous Cell Carcinoma Antigens in Psoriatic Skin Atsushi Takeda, Dousei Higuchi, Tadahito Takahashi, Masashi Ogo, Peter Baciu,

Coexpression of Integrin αvβ3 and Matrix Metalloproteinase-2 (MMP-2) Coincides with MMP-2 Activation: Correlation with Melanoma Progression Uta B. Hofmann,

Rose-Anne Romano, Barbara Birkaya, Satrajit Sinha

Bulge- and Basal Layer-Specific Expression of Fibroblast Growth Factor-13 (FHF-2) in Mouse Skin Mitsuko Kawano, Satoshi Suzuki, Masashi Suzuki, Junko.

Expression of Multiple Cytochrome P450 Enzymes and Multidrug Resistance- Associated Transport Proteins in Human Skin Keratinocytes Jens M. Baron, Daniela.

Isolation and Characterization of Human Repetin, a Member of the Fused Gene Family of the Epidermal Differentiation Complex Marcel Huber, Georges Siegenthaler,

Highly Complex Peptide Aggregates of the S100 Fused-Type Protein Hornerin Are Present in Human Skin Zhihong Wu, Ulf Meyer-Hoffert, Katrin Reithmayer,

EGF Upregulates, Whereas TGF-β Downregulates, the Hyaluronan Synthases Has2 and Has3 in Organotypic Keratinocyte Cultures: Correlations with Epidermal.

Psoriasis Upregulated Phorbolin-1 Shares Structural but not Functional Similarity to the mRNA-Editing Protein Apobec-1 Peder Madsen, Julio E. Celis,

Both of the N-Terminal and C-Terminal Regions of Human Papillomavirus Type 16 E7 are Essential for Immortalization of Primary Rat Cells Toshiharu Yamashita,

A Novel Mouse Gene, Sh3yl1, is Expressed in the Anagen Hair Follicle

Upregulation of Class II β-Tubulin Expression in Differentiating Keratinocytes Woong-Hee Lee, Joo-Young Kim, Young-Sik Kim, Hye-Joon Song, Ki-Joon Song,

Transcriptional Control of the Mouse Col7a1 Gene in Keratinocytes: Basal and Transforming Growth Factor-β Regulated Expression Michael Naso, Jouni Uitto,

Analysis of Mouse Keratin 6a Regulatory Sequences in Transgenic Mice Reveals Constitutive, Tissue-Specific Expression by a Keratin 6a Minigene Donna.

Size Polymorphisms in the Human Ultrahigh Sulfur Hair Keratin-Associated Protein 4, KAP4, Gene Family Naoyuki Kariya, Yutaka Shimomura, Masaaki Ito

Volume 60, Issue 2, Pages (August 2001)

Characterization of Human Keratin-Associated Protein 1 Family Members

Fatty Acid Transport Protein 1 Can Compensate for Fatty Acid Transport Protein 4 in the Developing Mouse Epidermis Meei-Hua Lin, Jeffrey H. Miner Journal.

Inhibition of Hair Follicle Growth by a Laminin-1 G-Domain Peptide, RKRLQVQLSIRT, in an Organ Culture of Isolated Vibrissa Rudiment1 Kazuhiro Hayashi,

Proteolytic Activation Cascade of the Netherton Syndrome–Defective Protein, LEKTI, in the Epidermis: Implications for Skin Homeostasis Paola Fortugno,

Localization of Rat FGF-5 Protein in Skin Macrophage-like Cells and FGF-5S Protein in Hair Follicle: Possible Involvement of twoFgf-5 Gene Products in.

Characterization and Chromosomal Localization of Human Hair-Specific Keratin Genes and Comparative Expression During the Hair Growth Cycle Paul E. Bowden,

Changing Pattern of Deiminated Proteins in Developing Human Epidermis

Profilaggrin Requires both Linker and Filaggrin Peptide Sequences to Form Granules: Implications for Profilaggrin Processing In Vivo1 Melanie K. Kuechle,

Keratinocytes Synthesize Enteropeptidase and Multiple Forms of Trypsinogen during Terminal Differentiation Jotaro Nakanishi, Mami Yamamoto, Junichi Koyama,

Isolation and Characterization of a Putative Keratin-Associated Protein Gene Expressed in Embryonic Skin of Mice Mikiro Takaishi, Yoshimi Takata, Toshio.

Matthew A. Sleeman, James D. Watson, J. Greg Murison

Characterization of a Novel Isoform of α-Nascent Polypeptide-associated Complex as IgE-defined Autoantigen Roschanak Mossabeb, Susanne Seiberler, Irene.

Μ-Crystallin, Thyroid Hormone-binding Protein, is Expressed Abundantly in the Murine Inner Root Sheath Cells Noriaki Aoki, Kaoru Ito, Masaaki Ito Journal.

Characterization of Mouse Profilaggrin: Evidence for Nuclear Engulfment and Translocation of the Profilaggrin B-Domain during Epidermal Differentiation

Noritaka Oyama, Keiji Iwatsuki, Yoshimi Homma, Fumio Kaneko

Deimination of Human Hornerin Enhances its Processing by Calpain-1 and its Cross- Linking by Transglutaminases Chiung-Yueh Hsu, Géraldine Gasc, Anne-Aurélie.

Functional Analysis of the Profilaggrin N-Terminal Peptide: Identification of Domains that Regulate Nuclear and Cytoplasmic Distribution David J. Pearton,

Peptidylarginine Deiminase Isoforms Are Differentially Expressed in the Anagen Hair Follicles and Other Human Skin Appendages Rachida Nachat, Marie-Claire.

Expression of T-Cadherin in Basal Keratinocytes of Skin

The Melanocortin 5 Receptor is Expressed in Human Sebaceous Glands and Rat Preputial Cells Diane Thiboutot, Aruntha Sivarajah, Kathryn Gilliland, Zhaoyuan.

Serotoninergic System in Hamster Skin

Human Elastase 1: Evidence for Expression in the Skin and the Identification of a Frequent Frameshift Polymorphism Ulvi Talas, John Dunlop, Sahera Khalaf,

Trangenic Misexpression of the Differentiation-Specific Desmocollin Isoform 1 in Basal Keratinocytes Frank Henkler, Molly Strom, Kathleen Mathers, Hayley.

Versican, a Major Hyaluronan-Binding Component in the Dermis, Loses its Hyaluronan- Binding Ability in Solar Elastosis Keiko Hasegawa, Masahiko Yoneda,

Protease-Activated Receptor 2, a Receptor Involved in Melanosome Transfer, is Upregulated in Human Skin by Ultraviolet Irradiation Glynis Scott, Cristina.

Regulation of the Expression of Peptidylarginine Deiminase Type II Gene (PADI2) in Human Keratinocytes Involves Sp1 and Sp3 Transcription Factors Sijun.

Expression of μ-Opiate Receptor in Human Epidermis and Keratinocytes

Volume 60, Issue 2, Pages (August 2001)

Bikunin, a Serine Protease Inhibitor, is Present on the Cell Boundary of Epidermis Cui Chang-Yi, Yoshinori Aragane, Akira Maeda, Piao Yu-Lan, Masae Takahashi,

Characterization of Human Keratin-Associated Protein 1 Family Members

Rab3a and SNARE Proteins: Potential Regulators of Melanosome Movement

Multiple Epidermal Connexins are Expressed in Different Keratinocyte Subpopulations Including Connexin 31 Wei-Li Di, Elizabeth L. Rugg, Irene M. Leigh,

I. Elisabeth Ekholm, Maria Brattsand, Torbjörn Egelrud

A Novel Gene Expressed in Human Keratinocytes with Long-Term In Vitro Growth Potential is Required for Cell Growth Laure Aurelian, Cynthia C. Smith,

IgG Autoantibodies from Bullous Pemphigoid (BP) Patients Bind Antigenic Sites on Both the Extracellular and the Intracellular Domains of the BP Antigen.

The Expression of Vitamin D-Upregulated Protein 1 in Skin and its Interaction with Sciellin in Cultured Keratinocytes Marie-France Champliaud, Alain.

Expression of FcRn, the MHC Class I-Related Receptor for IgG, in Human Keratinocytes Karla Cauza, Gabriele Hinterhuber, Ruth Dingelmaier-Hovorka, Karin.

Shigeru Kusuda, Cui Chang-Yi, Masae Takahashi, Tadashi Tezuka

Gli1 Protein is Expressed in Basal Cell Carcinomas, Outer Root Sheath Keratinocytes and a Subpopulation of Mesenchymal Cells in Normal Human Skin Lucy.

Yasushi Hanakawa, Hong Li, Chenyan Lin, John R

Staphylococcal Exfoliative Toxin B Specifically Cleaves Desmoglein 1

Presentation transcript:

Human Peptidylarginine Deiminase Type III: Molecular Cloning and Nucleotide Sequence of the cDNA, Properties of the Recombinant Enzyme, and Immunohistochemical Localization in Human Skin Takuya Kanno, Masakazu Shiraiwa, Hidenari Takahara Journal of Investigative Dermatology Volume 115, Issue 5, Pages 813-823 (November 2000) DOI: 10.1046/j.1523-1747.2000.00131.x Copyright © 2000 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 1 Cloning strategy of human PAD type III cDNA by reverse transcription–PCR, 5′-RACE and 3′-RACE. Products A, B, and C showed cDNA fragments produced by PCR amplification. The clones phPAD3–1, phPAD3–2, and phPAD3–3 were obtained by reverse transcription–PCR. phPAD3–4 and phPAD3–5 were cloned by 5′-RACE and 3′-RACE, respectively. Horizontal arrows denote primers used for reverse transcription–PCR, 5′-RACE, and 3′-RACE amplification. These sequences were combined to construct the composite cDNA in which the open box and lines indicate the coding and noncoding sequences, respectively. Only the relevant restriction site (KpnI) is shown. Journal of Investigative Dermatology 2000 115, 813-823DOI: (10.1046/j.1523-1747.2000.00131.x) Copyright © 2000 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 2 Nucleotide sequence and deduced amino acid sequence of human PAD type III. The DNA sequence is numbered from the first nucleotide of the cDNA. The numbers are on the right of each line. The deduced amino acid sequence is displayed below the nucleotide sequence in one letter code starting from the putative initiation methionine. A putative polyadenylation signal is underlined. The amino acid sequence indicated by wave line is that of a synthetic peptide for producing anti-hPAD3 antibody. Journal of Investigative Dermatology 2000 115, 813-823DOI: (10.1046/j.1523-1747.2000.00131.x) Copyright © 2000 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 3 Sequential fractionation of PAD activity from an extract of E. coli harboring pKKhPAD3. (A) DEAE-Sepharose Fast Flow column after an ammonium sulfate fractionation step. (B) Fractionation of the PAD peak from (A) on an aminohexyl-Bio Gel A15m column. PAD activity of each fraction was measured with Bz-L-Arg-O-Et as a substrate. The fractions indicated by horizontal bar in (B) were pooled and stored at 4°C until use. Journal of Investigative Dermatology 2000 115, 813-823DOI: (10.1046/j.1523-1747.2000.00131.x) Copyright © 2000 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 4 Amino acid sequence comparison of PAD type III of different species. The sequences are aligned by using the Malign program of GENETYX-MAC. The identical residues that conserved in at least three gene products are boxed. The Abbreviations used in the figure are: mPAD3, mouse PAD type III; rPAD3, rat PAD type III; sPAD, sheep PAD type III; hPAD, human PAD type III. The accession numbers of the sequence data cited for the comparison are as follows: mouse PAD type III, AB013849; rat PAD type III, D88034; sheep PAD type III, U85264; human PAD type III, AB026831. Journal of Investigative Dermatology 2000 115, 813-823DOI: (10.1046/j.1523-1747.2000.00131.x) Copyright © 2000 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 5 SDS–PAGE of fractions in each purification step of the recombinant human PAD type III. Aliquots of samples were subjected to SDS–PAGE and stained with Coomassie Brilliant Blue. Lane M, molecular weight markers; lane 1, crude extract (25 μg of protein); lane 2, ammonium sulfate fractionation step (25 μg); lane 3, DEAE-Sepharose Fast Flow fractionation step (25 μg); lane 4, aminohexyl-Bio Gel A15m fractionation step (2 μg). Purified recombinant PAD type III was indicated by arrowhead. Journal of Investigative Dermatology 2000 115, 813-823DOI: (10.1046/j.1523-1747.2000.00131.x) Copyright © 2000 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 6 Time course of deimination of filaggrin repeat unit and THH-8 by human PAD type III. Each protein was deiminated by the enzyme as described in Materials and Methods. At the indicated times, the reaction was terminated by ethylenediamine tetraacetic acid, and the citrulline residues in each preparations were determined by amino acid analysis. Conversion rate of arginine residues to citrulline residues of each protein (○, filaggrin repeat unit; •, THH-8) was shown. Journal of Investigative Dermatology 2000 115, 813-823DOI: (10.1046/j.1523-1747.2000.00131.x) Copyright © 2000 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 7 PAGE analyses of deimination of filaggrin repeat unit and THH-8 by human PAD type III. Each protein was deiminated by the enzyme as described in Materials and Methods. After the appropriate incubation period, an aliquot of each reaction mixture was subjected to SDS–PAGE (A, C) and native PAGE (B, D) and stained with Coomassie Brilliant Blue. (A, B) PAGE patterns of filaggrin repeat unit incubated with human PAD type III. (C, D) PAGE patterns of THH-8 incubated with human PAD type III. Lane M, molecular weight markers; lane 1, 0 min; lane 2, 10 min; lane 3, 20 min; lane 4, 40 min; lane 5, 80 min; lane 6, 360 min; lane 7, 720 min; lane 8, 1440 min. –, cathode; +, anode. Journal of Investigative Dermatology 2000 115, 813-823DOI: (10.1046/j.1523-1747.2000.00131.x) Copyright © 2000 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 8 Monospecific recognition of anti-hPAD3 antibody toward PAD type III isoform. Four different recombinant mouse PAD isoforms and recombinant human PAD type III (0.3 μg of each protein), extracts from cultured human epidermal keratinocyte and hair follicle (10 μg of each protein) were resolved on SDS–PAGE and blotted on to a polyvinylidene difluoride membrane. The membrane was stained with Ponceau S (A) for protein and immunoblotted with anti-hPAD3 antibody (B). Lane 1, mouse PAD type I, lane 2, mouse PAD type II; lane 3, mouse PAD type III; lane 4, mouse PAD type IV; lane 5, human PAD type III; lane 6, extract from cultured human epidermis; lane 7, extract from human hair follicle. Arrowhead shows the immunoreactive band of human PAD type III. The positions of protein standards run in the same experiment are indicated on the left: myosin (200 kDa), β-galactosidase (116 kDa), phosphorylase b (97 kDa), bovine serum albumin (66 kDa), ovalbumin (45 kDa), and carbonic anhydrase (31 kDa). Journal of Investigative Dermatology 2000 115, 813-823DOI: (10.1046/j.1523-1747.2000.00131.x) Copyright © 2000 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 9 Immunofluorescence staining of human hair follicle with anti-hPAD3, anti-trichohyalin, and anti-keratin 14 antibodies. (A–C, H–J) Longitudinal sections of hair follicles. (D–G, K–N) Transverse sections that are roughly corresponded to the area stated by numerals in C and J, respectively. The correspondences of each transverse section with the longitudinal sections are as follows: 1–D, 2–E, 3–F, 4–G in (C); 1–K, 2–L, 3–M, 4–N in (J). (A) Immunofluorescence staining with anti-hPAD3 antibody. (B) Immunofluorescence staining with anti-trichohyalin antibody. (C–G) Double immunofluorescence stainings with anti-hPAD3 and anti-trichohyalin antibodies. (H) Control staining with the preimmune serum. (I) Immunofluorescence staining with anti-keratin 14 antibody. (J–N) Double immunofluorescence stainings with anti-hPAD3 and anti-keratin 14 antibodies. Notes: PAD type III was coexpressed with trichohyalin in the IRS between supramatrix and keratogenous zone and in medulla. The type III enzyme was also localized to the ORS and cuticle of the hair. irs, inner root sheath; ors, outer root sheath; m, medulla; ch, cuticle of the hair; hs, hair shaft. Scale bar: 50 μm. Journal of Investigative Dermatology 2000 115, 813-823DOI: (10.1046/j.1523-1747.2000.00131.x) Copyright © 2000 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 10 Immunofluorescence staining of human epidermis and ORS at the infundibulum region with anti-hPAD3, anti-keratin 14, and anti-filaggrin antibodies. (A) Immunofluorescence staining with anti-hPAD3 antibody. (B) Double immunofluorescence stainings with anti-hPAD3 and anti-keratin 14 antibodies. The yellow color in the basal layer of the epidermis indicates that the expression of the PAD type III starts at the early stage of epidermal proliferation. (C) Double immunofluorescence stainings with anti-hPAD3 and anti-filaggrin antibodies. Most of the granular and cornified cells containing filaggrin show red color. A part of the granular cells facing out on the spinous cell layer turned yellow color, suggesting that the PAD type III enzyme partly colocalizes with filaggrin in these cells. epi, epidermis; ors, outer root sheath, ifb, infundibulum region; b, basal cell layer; g, granular cell layer; c, cornified cell layer. Scale bar, 200 μm. Journal of Investigative Dermatology 2000 115, 813-823DOI: (10.1046/j.1523-1747.2000.00131.x) Copyright © 2000 The Society for Investigative Dermatology, Inc Terms and Conditions