Corneal epithelial-specific cytokeratin 3 is an autoantigen in Wegener's granulomatosis-associated peripheral ulcerative keratitis.
OBJECTIVE
In a previous investigation it was demonstrated that circulating antibodies to a 66-kDa corneal epithelial antigen (BCEA-A) are associated with peripheral ulcerative keratitis (PUK) in patients with Wegener's granulomatosis (WG). The aim of this study was to identify BCEA-A.


METHODS
The 66-kDa antigen was purified from a bovine corneal epithelial protein extract, using DE52 ion exchange chromatography. Purified protein was used to raise rabbit polyclonal antibodies. These antibodies were used to screen a bovine corneal epithelial cDNA expression library. Positive clones were purified and sequenced. Clones were identified by DNA sequence homology searches of the GenBank DNA database.


RESULTS
A cDNA clone that demonstrated strong binding to both the rabbit polyclonal antibody and patient sera, showed 85% homology to rabbit cytokeratin 3 (K3). K3 is a basic cytokeratin specific to corneal epithelium. No bovine DNA sequence for K3 is available. However, bovine K3 is larger than rabbit K3, with a molecular weight of 66 kDa. Immunofluorescence using both patient sera and the rabbit antibody demonstrated a cytoplasmic binding pattern on human corneal epithelium.


CONCLUSIONS
This evidence suggests that the 66-kDa autoantigen (BCEA-A) associated with PUK in WG is cytokeratin 3, and this may form the basis of a diagnostic/prognostic test.
Reynolds
I
I
Musculoskeletal Research Group, University of Manchester, and Central Manchester Health Care NHS Trust, United Kingdom.
Tullo
A B
AB
John
S L
SL
Holt
P J
PJ
Hillarby
M C
MC
D000595:Amino Acid Sequence; D000818:Animals; D001324:Autoantigens / Q000378:metabolism*; D002417:Cattle; D002852:Chromatography, Ion Exchange; D003320:Corneal Ulcer / Q000378:metabolism*; D017931:DNA Primers / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D019573:Epithelium, Corneal / Q000737:chemistry / Q000378:metabolism*; D019084:Fluorescent Antibody Technique, Indirect; D014890:Granulomatosis with Polyangiitis / Q000378:metabolism*; D006801:Humans; D007633:Keratins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011817:Rabbits; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D012689:Sequence Homology, Nucleic Acid
D001324:Autoantigens; D017931:DNA Primers; D007633:Keratins