Ribophorin ii is involved in the tissue factor expression mediated by phosphatidylserinedependent antiprothrombin antibody on monocytes

Yuichiro Fujieda, Olga Amengual, Masaki Matsumoto, Kimiko Kuroki, Hidehisa Takahashi, Michihito Kono, Takashi Kurita, Kotaro Otomo, Masaru Kato, Kenji Oku, Toshiyuki Bohgaki, Tetsuya Horita, Shinsuke Yasuda, Katsumi Maenaka, Shigetsugu Hatakeyama, Keiichi Nakayama, Tatsuya Atsumi

Research output: Contribution to journalArticle

1 Citation (Scopus)

Abstract

Objective. Phosphatidylserine-dependent, also called aPS-PT, recognizes the phosphati dylserine prothrombin complex, which is associated with APS. We have previously reported that aPSPT induces tissue factor (TF) expression on monocytes through the p38 mitogen-activated protein kinase pathway. However, the cell surface interaction between prothrombin and aPS-PT, which is involved in the activation of cell-signalling pathways, has remained unknown. The objective of this study was to identify membrane proteins involved in the binding of prothrombin and aPS-PT to monocyte surfaces as well as the induction of TF expression. Methods. RAW264.7 cells with FLAG-tagged prothrombin were incubated and separated using affinity chromatography with anti-FLAG antibody-conjugated Sepharose beads. Immunopurified proteins were then analysed by an online nano-liquid chromatography tandem mass spectrometry. The binding between prothrombin and the identified protein, ribophorin II (RPN2), was analysed by ELISA and surface plasmon resonance. To elucidate the role of RPN2 in TF expression, the TF mRNA level in RAW264.7 cells treated with RPN2 small interfering RNA was determined by quantitative real-time PCR (qPCR). Results. RPN2 was identified as a candidate molecule involved in the binding of prothrombin to the cell surface. The binding between prothrombin and RPN2 was confirmed by ELISA and surface plasmon resonance. RAW264.7 cells treated with RPN2 small interfering RNA showed significant reduction of the TF expression mediated by prothrombin and a mouse monoclonal aPS-PT. Conclusion. We identified that RPN2 is one of the prothrombin-binding proteins on monocyte surfaces, suggesting that RPN2 is involved in the pathophysiology of thrombosis in patients with APS.

Original languageEnglish
Pages (from-to)1117-1126
Number of pages10
JournalRheumatology (United Kingdom)
Volume55
Issue number6
DOIs
Publication statusPublished - Jun 1 2016

Fingerprint

Thromboplastin
Prothrombin
Monocytes
Antibodies
Surface Plasmon Resonance
Small Interfering RNA
Enzyme-Linked Immunosorbent Assay
ribophorin
Phosphatidylserines
p38 Mitogen-Activated Protein Kinases
Tandem Mass Spectrometry
Affinity Chromatography
Cell Communication
Liquid Chromatography
Sepharose
Real-Time Polymerase Chain Reaction
Anti-Idiotypic Antibodies
Carrier Proteins
Membrane Proteins
Proteins

All Science Journal Classification (ASJC) codes

  • Rheumatology
  • Pharmacology (medical)

Cite this

Ribophorin ii is involved in the tissue factor expression mediated by phosphatidylserinedependent antiprothrombin antibody on monocytes. / Fujieda, Yuichiro; Amengual, Olga; Matsumoto, Masaki; Kuroki, Kimiko; Takahashi, Hidehisa; Kono, Michihito; Kurita, Takashi; Otomo, Kotaro; Kato, Masaru; Oku, Kenji; Bohgaki, Toshiyuki; Horita, Tetsuya; Yasuda, Shinsuke; Maenaka, Katsumi; Hatakeyama, Shigetsugu; Nakayama, Keiichi; Atsumi, Tatsuya.

In: Rheumatology (United Kingdom), Vol. 55, No. 6, 01.06.2016, p. 1117-1126.

Research output: Contribution to journalArticle

Fujieda, Y, Amengual, O, Matsumoto, M, Kuroki, K, Takahashi, H, Kono, M, Kurita, T, Otomo, K, Kato, M, Oku, K, Bohgaki, T, Horita, T, Yasuda, S, Maenaka, K, Hatakeyama, S, Nakayama, K & Atsumi, T 2016, 'Ribophorin ii is involved in the tissue factor expression mediated by phosphatidylserinedependent antiprothrombin antibody on monocytes', Rheumatology (United Kingdom), vol. 55, no. 6, pp. 1117-1126. https://doi.org/10.1093/rheumatology/kew005
Fujieda, Yuichiro ; Amengual, Olga ; Matsumoto, Masaki ; Kuroki, Kimiko ; Takahashi, Hidehisa ; Kono, Michihito ; Kurita, Takashi ; Otomo, Kotaro ; Kato, Masaru ; Oku, Kenji ; Bohgaki, Toshiyuki ; Horita, Tetsuya ; Yasuda, Shinsuke ; Maenaka, Katsumi ; Hatakeyama, Shigetsugu ; Nakayama, Keiichi ; Atsumi, Tatsuya. / Ribophorin ii is involved in the tissue factor expression mediated by phosphatidylserinedependent antiprothrombin antibody on monocytes. In: Rheumatology (United Kingdom). 2016 ; Vol. 55, No. 6. pp. 1117-1126.
@article{6a421a9d74a443e2800bb91b99803fa6,
title = "Ribophorin ii is involved in the tissue factor expression mediated by phosphatidylserinedependent antiprothrombin antibody on monocytes",
abstract = "Objective. Phosphatidylserine-dependent, also called aPS-PT, recognizes the phosphati dylserine prothrombin complex, which is associated with APS. We have previously reported that aPSPT induces tissue factor (TF) expression on monocytes through the p38 mitogen-activated protein kinase pathway. However, the cell surface interaction between prothrombin and aPS-PT, which is involved in the activation of cell-signalling pathways, has remained unknown. The objective of this study was to identify membrane proteins involved in the binding of prothrombin and aPS-PT to monocyte surfaces as well as the induction of TF expression. Methods. RAW264.7 cells with FLAG-tagged prothrombin were incubated and separated using affinity chromatography with anti-FLAG antibody-conjugated Sepharose beads. Immunopurified proteins were then analysed by an online nano-liquid chromatography tandem mass spectrometry. The binding between prothrombin and the identified protein, ribophorin II (RPN2), was analysed by ELISA and surface plasmon resonance. To elucidate the role of RPN2 in TF expression, the TF mRNA level in RAW264.7 cells treated with RPN2 small interfering RNA was determined by quantitative real-time PCR (qPCR). Results. RPN2 was identified as a candidate molecule involved in the binding of prothrombin to the cell surface. The binding between prothrombin and RPN2 was confirmed by ELISA and surface plasmon resonance. RAW264.7 cells treated with RPN2 small interfering RNA showed significant reduction of the TF expression mediated by prothrombin and a mouse monoclonal aPS-PT. Conclusion. We identified that RPN2 is one of the prothrombin-binding proteins on monocyte surfaces, suggesting that RPN2 is involved in the pathophysiology of thrombosis in patients with APS.",
author = "Yuichiro Fujieda and Olga Amengual and Masaki Matsumoto and Kimiko Kuroki and Hidehisa Takahashi and Michihito Kono and Takashi Kurita and Kotaro Otomo and Masaru Kato and Kenji Oku and Toshiyuki Bohgaki and Tetsuya Horita and Shinsuke Yasuda and Katsumi Maenaka and Shigetsugu Hatakeyama and Keiichi Nakayama and Tatsuya Atsumi",
year = "2016",
month = "6",
day = "1",
doi = "10.1093/rheumatology/kew005",
language = "English",
volume = "55",
pages = "1117--1126",
journal = "Rheumatology",
issn = "1462-0324",
publisher = "Oxford University Press",
number = "6",

}

TY - JOUR

T1 - Ribophorin ii is involved in the tissue factor expression mediated by phosphatidylserinedependent antiprothrombin antibody on monocytes

AU - Fujieda, Yuichiro

AU - Amengual, Olga

AU - Matsumoto, Masaki

AU - Kuroki, Kimiko

AU - Takahashi, Hidehisa

AU - Kono, Michihito

AU - Kurita, Takashi

AU - Otomo, Kotaro

AU - Kato, Masaru

AU - Oku, Kenji

AU - Bohgaki, Toshiyuki

AU - Horita, Tetsuya

AU - Yasuda, Shinsuke

AU - Maenaka, Katsumi

AU - Hatakeyama, Shigetsugu

AU - Nakayama, Keiichi

AU - Atsumi, Tatsuya

PY - 2016/6/1

Y1 - 2016/6/1

N2 - Objective. Phosphatidylserine-dependent, also called aPS-PT, recognizes the phosphati dylserine prothrombin complex, which is associated with APS. We have previously reported that aPSPT induces tissue factor (TF) expression on monocytes through the p38 mitogen-activated protein kinase pathway. However, the cell surface interaction between prothrombin and aPS-PT, which is involved in the activation of cell-signalling pathways, has remained unknown. The objective of this study was to identify membrane proteins involved in the binding of prothrombin and aPS-PT to monocyte surfaces as well as the induction of TF expression. Methods. RAW264.7 cells with FLAG-tagged prothrombin were incubated and separated using affinity chromatography with anti-FLAG antibody-conjugated Sepharose beads. Immunopurified proteins were then analysed by an online nano-liquid chromatography tandem mass spectrometry. The binding between prothrombin and the identified protein, ribophorin II (RPN2), was analysed by ELISA and surface plasmon resonance. To elucidate the role of RPN2 in TF expression, the TF mRNA level in RAW264.7 cells treated with RPN2 small interfering RNA was determined by quantitative real-time PCR (qPCR). Results. RPN2 was identified as a candidate molecule involved in the binding of prothrombin to the cell surface. The binding between prothrombin and RPN2 was confirmed by ELISA and surface plasmon resonance. RAW264.7 cells treated with RPN2 small interfering RNA showed significant reduction of the TF expression mediated by prothrombin and a mouse monoclonal aPS-PT. Conclusion. We identified that RPN2 is one of the prothrombin-binding proteins on monocyte surfaces, suggesting that RPN2 is involved in the pathophysiology of thrombosis in patients with APS.

AB - Objective. Phosphatidylserine-dependent, also called aPS-PT, recognizes the phosphati dylserine prothrombin complex, which is associated with APS. We have previously reported that aPSPT induces tissue factor (TF) expression on monocytes through the p38 mitogen-activated protein kinase pathway. However, the cell surface interaction between prothrombin and aPS-PT, which is involved in the activation of cell-signalling pathways, has remained unknown. The objective of this study was to identify membrane proteins involved in the binding of prothrombin and aPS-PT to monocyte surfaces as well as the induction of TF expression. Methods. RAW264.7 cells with FLAG-tagged prothrombin were incubated and separated using affinity chromatography with anti-FLAG antibody-conjugated Sepharose beads. Immunopurified proteins were then analysed by an online nano-liquid chromatography tandem mass spectrometry. The binding between prothrombin and the identified protein, ribophorin II (RPN2), was analysed by ELISA and surface plasmon resonance. To elucidate the role of RPN2 in TF expression, the TF mRNA level in RAW264.7 cells treated with RPN2 small interfering RNA was determined by quantitative real-time PCR (qPCR). Results. RPN2 was identified as a candidate molecule involved in the binding of prothrombin to the cell surface. The binding between prothrombin and RPN2 was confirmed by ELISA and surface plasmon resonance. RAW264.7 cells treated with RPN2 small interfering RNA showed significant reduction of the TF expression mediated by prothrombin and a mouse monoclonal aPS-PT. Conclusion. We identified that RPN2 is one of the prothrombin-binding proteins on monocyte surfaces, suggesting that RPN2 is involved in the pathophysiology of thrombosis in patients with APS.

UR - http://www.scopus.com/inward/record.url?scp=84974793178&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84974793178&partnerID=8YFLogxK

U2 - 10.1093/rheumatology/kew005

DO - 10.1093/rheumatology/kew005

M3 - Article

C2 - 26895716

AN - SCOPUS:84974793178

VL - 55

SP - 1117

EP - 1126

JO - Rheumatology

JF - Rheumatology

SN - 1462-0324

IS - 6

ER -