Characterization of the antibodies to p62 nucleoporin in primary biliary cirrhosis using human recombinant antigen

Józefa Wȩsierska-Ga̧dek, Anna Klima, Carmen Ranftler, Oxana Komina, John Hanover, Pietro Invernizzi, Edward Penner

Research output: Contribution to journalArticle

10 Citations (Scopus)

Abstract

Reactivity of sera from patients with primary biliary cirrhosis (PBC) with a 60 kDa component of nuclear pore complexes (NPCs), purified by affinity chromatography on wheat-germ agglutinin (WGA)-Sepharose, was previously detected. Recently, clinical significance of the anti-NPC antibodies in PBC became evident. In the light of recent reports, indicating the correlation of the anti-NPC antibodies with severity and progression of the disease, the characterization of the reactive antigens is becoming essential in the clinical management of patients with PBC. Since accurate autoantibody detection represents one of the fundamental requirements for a reliable testing, we have generated a human recombinant p62 protein and validated an immunoprecipitation assay for the detection of anti-p62. We also demonstrated that the generated human recombinant p62 nucleoporin was modified by N-acetylglucosamine residues. More than50%of tested PBC sera precipitated 35S-radioactively labeled p62 recombinant nucleoporin and 40% recognized this recombinant antigen by immunoblotting. We compared the reactivity of PBC sera with rat and human nucleoporin. The incidence of anti-p62 nucleoporin positive PBC sera increased by 15% when human recombinant antigen was used. The titer of autoantibodies in p62-positive PBC samples strongly varied. Preadsorption of the PBC sera with p62 recombinant protein completely abolished their reactivity with the antigen. In conclusion, this study unequivocally proves that autoantibodies reacting with the 60 kDa component of NPCs target p62 nucleoporin and, more importantly, provide a better antigen source for future evaluations of the clinical role of anti-p62 in PBC.

Original languageEnglish (US)
Pages (from-to)27-37
Number of pages11
JournalJournal of Cellular Biochemistry
Volume104
Issue number1
DOIs
StatePublished - May 1 2008
Externally publishedYes

Fingerprint

Biliary Liver Cirrhosis
Antigens
Nuclear Pore
Nuclear Pore Complex Proteins
Antibodies
Autoantibodies
Serum
Affinity chromatography
Recombinant proteins
Wheat Germ Agglutinins
Acetylglucosamine
Recombinant Proteins
Sepharose
Rats
Assays
nuclear pore protein p62
Affinity Chromatography
Immunoprecipitation
Immunoblotting
Testing

Keywords

  • ANA
  • Autoimmunity
  • gp210 glycoprotein
  • Human recombinant p62 nucleoporin
  • Nuclear envelope
  • Nuclear pore complexes
  • p62 nucleoporin

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

Cite this

Wȩsierska-Ga̧dek, J., Klima, A., Ranftler, C., Komina, O., Hanover, J., Invernizzi, P., & Penner, E. (2008). Characterization of the antibodies to p62 nucleoporin in primary biliary cirrhosis using human recombinant antigen. Journal of Cellular Biochemistry, 104(1), 27-37. https://doi.org/10.1002/jcb.21595

Characterization of the antibodies to p62 nucleoporin in primary biliary cirrhosis using human recombinant antigen. / Wȩsierska-Ga̧dek, Józefa; Klima, Anna; Ranftler, Carmen; Komina, Oxana; Hanover, John; Invernizzi, Pietro; Penner, Edward.

In: Journal of Cellular Biochemistry, Vol. 104, No. 1, 01.05.2008, p. 27-37.

Research output: Contribution to journalArticle

Wȩsierska-Ga̧dek, J, Klima, A, Ranftler, C, Komina, O, Hanover, J, Invernizzi, P & Penner, E 2008, 'Characterization of the antibodies to p62 nucleoporin in primary biliary cirrhosis using human recombinant antigen', Journal of Cellular Biochemistry, vol. 104, no. 1, pp. 27-37. https://doi.org/10.1002/jcb.21595
Wȩsierska-Ga̧dek, Józefa ; Klima, Anna ; Ranftler, Carmen ; Komina, Oxana ; Hanover, John ; Invernizzi, Pietro ; Penner, Edward. / Characterization of the antibodies to p62 nucleoporin in primary biliary cirrhosis using human recombinant antigen. In: Journal of Cellular Biochemistry. 2008 ; Vol. 104, No. 1. pp. 27-37.
@article{9136c6d1a13848ab837652495c63ebd8,
title = "Characterization of the antibodies to p62 nucleoporin in primary biliary cirrhosis using human recombinant antigen",
abstract = "Reactivity of sera from patients with primary biliary cirrhosis (PBC) with a 60 kDa component of nuclear pore complexes (NPCs), purified by affinity chromatography on wheat-germ agglutinin (WGA)-Sepharose, was previously detected. Recently, clinical significance of the anti-NPC antibodies in PBC became evident. In the light of recent reports, indicating the correlation of the anti-NPC antibodies with severity and progression of the disease, the characterization of the reactive antigens is becoming essential in the clinical management of patients with PBC. Since accurate autoantibody detection represents one of the fundamental requirements for a reliable testing, we have generated a human recombinant p62 protein and validated an immunoprecipitation assay for the detection of anti-p62. We also demonstrated that the generated human recombinant p62 nucleoporin was modified by N-acetylglucosamine residues. More than50{\%}of tested PBC sera precipitated 35S-radioactively labeled p62 recombinant nucleoporin and 40{\%} recognized this recombinant antigen by immunoblotting. We compared the reactivity of PBC sera with rat and human nucleoporin. The incidence of anti-p62 nucleoporin positive PBC sera increased by 15{\%} when human recombinant antigen was used. The titer of autoantibodies in p62-positive PBC samples strongly varied. Preadsorption of the PBC sera with p62 recombinant protein completely abolished their reactivity with the antigen. In conclusion, this study unequivocally proves that autoantibodies reacting with the 60 kDa component of NPCs target p62 nucleoporin and, more importantly, provide a better antigen source for future evaluations of the clinical role of anti-p62 in PBC.",
keywords = "ANA, Autoimmunity, gp210 glycoprotein, Human recombinant p62 nucleoporin, Nuclear envelope, Nuclear pore complexes, p62 nucleoporin",
author = "J{\'o}zefa Wȩsierska-Ga̧dek and Anna Klima and Carmen Ranftler and Oxana Komina and John Hanover and Pietro Invernizzi and Edward Penner",
year = "2008",
month = "5",
day = "1",
doi = "10.1002/jcb.21595",
language = "English (US)",
volume = "104",
pages = "27--37",
journal = "Journal of Cellular Biochemistry",
issn = "0730-2312",
publisher = "Wiley-Liss Inc.",
number = "1",

}

TY - JOUR

T1 - Characterization of the antibodies to p62 nucleoporin in primary biliary cirrhosis using human recombinant antigen

AU - Wȩsierska-Ga̧dek, Józefa

AU - Klima, Anna

AU - Ranftler, Carmen

AU - Komina, Oxana

AU - Hanover, John

AU - Invernizzi, Pietro

AU - Penner, Edward

PY - 2008/5/1

Y1 - 2008/5/1

N2 - Reactivity of sera from patients with primary biliary cirrhosis (PBC) with a 60 kDa component of nuclear pore complexes (NPCs), purified by affinity chromatography on wheat-germ agglutinin (WGA)-Sepharose, was previously detected. Recently, clinical significance of the anti-NPC antibodies in PBC became evident. In the light of recent reports, indicating the correlation of the anti-NPC antibodies with severity and progression of the disease, the characterization of the reactive antigens is becoming essential in the clinical management of patients with PBC. Since accurate autoantibody detection represents one of the fundamental requirements for a reliable testing, we have generated a human recombinant p62 protein and validated an immunoprecipitation assay for the detection of anti-p62. We also demonstrated that the generated human recombinant p62 nucleoporin was modified by N-acetylglucosamine residues. More than50%of tested PBC sera precipitated 35S-radioactively labeled p62 recombinant nucleoporin and 40% recognized this recombinant antigen by immunoblotting. We compared the reactivity of PBC sera with rat and human nucleoporin. The incidence of anti-p62 nucleoporin positive PBC sera increased by 15% when human recombinant antigen was used. The titer of autoantibodies in p62-positive PBC samples strongly varied. Preadsorption of the PBC sera with p62 recombinant protein completely abolished their reactivity with the antigen. In conclusion, this study unequivocally proves that autoantibodies reacting with the 60 kDa component of NPCs target p62 nucleoporin and, more importantly, provide a better antigen source for future evaluations of the clinical role of anti-p62 in PBC.

AB - Reactivity of sera from patients with primary biliary cirrhosis (PBC) with a 60 kDa component of nuclear pore complexes (NPCs), purified by affinity chromatography on wheat-germ agglutinin (WGA)-Sepharose, was previously detected. Recently, clinical significance of the anti-NPC antibodies in PBC became evident. In the light of recent reports, indicating the correlation of the anti-NPC antibodies with severity and progression of the disease, the characterization of the reactive antigens is becoming essential in the clinical management of patients with PBC. Since accurate autoantibody detection represents one of the fundamental requirements for a reliable testing, we have generated a human recombinant p62 protein and validated an immunoprecipitation assay for the detection of anti-p62. We also demonstrated that the generated human recombinant p62 nucleoporin was modified by N-acetylglucosamine residues. More than50%of tested PBC sera precipitated 35S-radioactively labeled p62 recombinant nucleoporin and 40% recognized this recombinant antigen by immunoblotting. We compared the reactivity of PBC sera with rat and human nucleoporin. The incidence of anti-p62 nucleoporin positive PBC sera increased by 15% when human recombinant antigen was used. The titer of autoantibodies in p62-positive PBC samples strongly varied. Preadsorption of the PBC sera with p62 recombinant protein completely abolished their reactivity with the antigen. In conclusion, this study unequivocally proves that autoantibodies reacting with the 60 kDa component of NPCs target p62 nucleoporin and, more importantly, provide a better antigen source for future evaluations of the clinical role of anti-p62 in PBC.

KW - ANA

KW - Autoimmunity

KW - gp210 glycoprotein

KW - Human recombinant p62 nucleoporin

KW - Nuclear envelope

KW - Nuclear pore complexes

KW - p62 nucleoporin

UR - http://www.scopus.com/inward/record.url?scp=46449135226&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=46449135226&partnerID=8YFLogxK

U2 - 10.1002/jcb.21595

DO - 10.1002/jcb.21595

M3 - Article

C2 - 17960595

AN - SCOPUS:46449135226

VL - 104

SP - 27

EP - 37

JO - Journal of Cellular Biochemistry

JF - Journal of Cellular Biochemistry

SN - 0730-2312

IS - 1

ER -