cDNA cloning of the 210-kDa paraneoplastic pemphigus antigen reveals that envoplakin is a component of the antigen complex

Soo Chan Kim, Young Do Kwon, In Joon Lee, Il Joo Lee, Sung Nam Chang, Tae Gyu Lee

Research output: Contribution to journalArticle

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Abstract

Although the 210 and 190-kDa proteins are the most frequently detected antigens reacting with sera of patients with paraneoplastic pemphigus (PNP) in immunoblot analysis, there is still uncertainty as to the nature of these PNP antigens. To isolate and characterize a cDNA clone encoding the 210-kDa PNP antigen, we screened a human keratinocyte λ gt 11 cDNA expression library by the immunoperoxidase method with serum IgG from a PNP patient. The IgG used for the immunoscreening of a keratinocyte cDNA expression library recognized 210- and 190kDa antigens by immunoblotting. A single clone, called here the PNP clone, producing a fusion protein that reacted strongly with the patient's IgG, was further characterized. Only the PNP patient's IgG, but not IgG from a normal control, pemphigus foliaceus, or pemphigus vulgaris patients, bound the plaques of this positive clone. Furthermore, PNP IgG affinity purified on plaques of this clone, but not unrelated clones, bound to keratinocyte cell surfaces by immunofluorescence and reacted with the 210kDa PNP antigen by immunoblotting. EcoRI digestion of the clone's cDNA insert demonstrated a 1.4kbp fragment. This cDNA insert was placed into a M13 mp 18 vector and sequenced. Sequence analysis revealed that the cDNA insert of the PNP clone encodes a part of the central rod domain and the COOH- terminal C domain of envoplakin, a newly defined precursor of the cornified envelope that is homologous to desmoplakin. This result demonstrates that the 210-kDa PNP antigen is envoplakin and PNP is an autoimmune disease that produces autoantibodies against intermediate filament-associated proteins in desmosomes and hemidesmosomes, desmoplakin, bullous pemphigoid antigen 1 (BPAG 1), and envoplakin.

Original languageEnglish
Pages (from-to)365-369
Number of pages5
JournalJournal of Investigative Dermatology
Volume109
Issue number3
DOIs
Publication statusPublished - 1997 Jan 1

Fingerprint

Pemphigus
Cloning
Organism Cloning
Complementary DNA
Immunoglobulin G
Antigens
Clone Cells
Desmoplakins
Keratinocytes
Proteins
Gene Library
Immunoblotting
envoplakin
Autoantibodies
Fusion reactions
Hemidesmosomes
Intermediate Filament Proteins
Desmosomes
Serum
Autoimmune Diseases

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Dermatology
  • Cell Biology

Cite this

Kim, Soo Chan ; Kwon, Young Do ; Lee, In Joon ; Lee, Il Joo ; Chang, Sung Nam ; Lee, Tae Gyu. / cDNA cloning of the 210-kDa paraneoplastic pemphigus antigen reveals that envoplakin is a component of the antigen complex. In: Journal of Investigative Dermatology. 1997 ; Vol. 109, No. 3. pp. 365-369.
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abstract = "Although the 210 and 190-kDa proteins are the most frequently detected antigens reacting with sera of patients with paraneoplastic pemphigus (PNP) in immunoblot analysis, there is still uncertainty as to the nature of these PNP antigens. To isolate and characterize a cDNA clone encoding the 210-kDa PNP antigen, we screened a human keratinocyte λ gt 11 cDNA expression library by the immunoperoxidase method with serum IgG from a PNP patient. The IgG used for the immunoscreening of a keratinocyte cDNA expression library recognized 210- and 190kDa antigens by immunoblotting. A single clone, called here the PNP clone, producing a fusion protein that reacted strongly with the patient's IgG, was further characterized. Only the PNP patient's IgG, but not IgG from a normal control, pemphigus foliaceus, or pemphigus vulgaris patients, bound the plaques of this positive clone. Furthermore, PNP IgG affinity purified on plaques of this clone, but not unrelated clones, bound to keratinocyte cell surfaces by immunofluorescence and reacted with the 210kDa PNP antigen by immunoblotting. EcoRI digestion of the clone's cDNA insert demonstrated a 1.4kbp fragment. This cDNA insert was placed into a M13 mp 18 vector and sequenced. Sequence analysis revealed that the cDNA insert of the PNP clone encodes a part of the central rod domain and the COOH- terminal C domain of envoplakin, a newly defined precursor of the cornified envelope that is homologous to desmoplakin. This result demonstrates that the 210-kDa PNP antigen is envoplakin and PNP is an autoimmune disease that produces autoantibodies against intermediate filament-associated proteins in desmosomes and hemidesmosomes, desmoplakin, bullous pemphigoid antigen 1 (BPAG 1), and envoplakin.",
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cDNA cloning of the 210-kDa paraneoplastic pemphigus antigen reveals that envoplakin is a component of the antigen complex. / Kim, Soo Chan; Kwon, Young Do; Lee, In Joon; Lee, Il Joo; Chang, Sung Nam; Lee, Tae Gyu.

In: Journal of Investigative Dermatology, Vol. 109, No. 3, 01.01.1997, p. 365-369.

Research output: Contribution to journalArticle

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T1 - cDNA cloning of the 210-kDa paraneoplastic pemphigus antigen reveals that envoplakin is a component of the antigen complex

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N2 - Although the 210 and 190-kDa proteins are the most frequently detected antigens reacting with sera of patients with paraneoplastic pemphigus (PNP) in immunoblot analysis, there is still uncertainty as to the nature of these PNP antigens. To isolate and characterize a cDNA clone encoding the 210-kDa PNP antigen, we screened a human keratinocyte λ gt 11 cDNA expression library by the immunoperoxidase method with serum IgG from a PNP patient. The IgG used for the immunoscreening of a keratinocyte cDNA expression library recognized 210- and 190kDa antigens by immunoblotting. A single clone, called here the PNP clone, producing a fusion protein that reacted strongly with the patient's IgG, was further characterized. Only the PNP patient's IgG, but not IgG from a normal control, pemphigus foliaceus, or pemphigus vulgaris patients, bound the plaques of this positive clone. Furthermore, PNP IgG affinity purified on plaques of this clone, but not unrelated clones, bound to keratinocyte cell surfaces by immunofluorescence and reacted with the 210kDa PNP antigen by immunoblotting. EcoRI digestion of the clone's cDNA insert demonstrated a 1.4kbp fragment. This cDNA insert was placed into a M13 mp 18 vector and sequenced. Sequence analysis revealed that the cDNA insert of the PNP clone encodes a part of the central rod domain and the COOH- terminal C domain of envoplakin, a newly defined precursor of the cornified envelope that is homologous to desmoplakin. This result demonstrates that the 210-kDa PNP antigen is envoplakin and PNP is an autoimmune disease that produces autoantibodies against intermediate filament-associated proteins in desmosomes and hemidesmosomes, desmoplakin, bullous pemphigoid antigen 1 (BPAG 1), and envoplakin.

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