Identification of ocular cicatricial pemphigoid antibody binding site(s) in human beta4 integrin.
OBJECTIVE
To identify specific site(s) on human ss4 molecule to which sera from ocular cicatricial pemphigoid (OCP) patients bind and to determine its role in the process of blister formation.


METHODS
Clone the fragments representing the extracellular and intracellular domain of ss4 molecule from normal human conjunctival mRNA into an expression vector; map the region to which sera from OCP patients bind by Western blot analysis. Determine the role of the immunodominant region in pathogenesis by demonstrating the ability of the rabbit antibody to the immunodominant region to produce separation of basement membrane zone (BMZ) from the basal epithelial layer when incubated with normal human conjunctiva in an in vitro organ culture model.


RESULTS
Majority of the OCP sera tested bound to the C-terminal end of the intracellular domain (IC3.0) of the human ss4 integrin. Further subcloning of IC3.0 demonstrated that a smaller fragment extending from 1489 aa to 1572 aa (IC3.4) was responsible for this binding. This region may have multiple antibody binding sites. Antibody to human IC3.0 and IC3.4 produced in rabbit, resulted in BMZ separation, histologically identical with that observed when normal human conjunctiva was cultured with OCP sera in an human conjunctival organ culture model.


CONCLUSIONS
These observations identify IC3.4 as the antibody binding site for sera of OCP patients and suggest a possible role for it in blister formation. Indirectly it highlights certain important aspects of the structural and functional dynamics of the biology of the hemidesmosomes and basement membranes.
Kumari
S
S
Department of Oral Medicine and Diagnostic Sciences, Harvard School of Dental Medicine, Boston, Massachusetts 02115, USA.
Bhol
K C
KC
Simmons
R K
RK
Razzaque
M S
MS
Letko
E
E
Foster
C S
CS
Razzaque Ahmed
A
A
Ahmed
A R
AR
D000818:Animals; D015703:Antigens, CD / Q000235:genetics / Q000378:metabolism*; D001323:Autoantibodies / Q000378:metabolism*; D001666:Binding Sites, Antibody; D015153:Blotting, Western; D002478:Cells, Cultured; D003228:Conjunctiva / Q000166:cytology / Q000378:metabolism; D003229:Conjunctival Diseases / Q000276:immunology*; D017931:DNA Primers / Q000737:chemistry; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D039663:Integrin beta4; D009924:Organ Culture Techniques; D010390:Pemphigoid, Benign Mucous Membrane / Q000276:immunology*; D010446:Peptide Fragments; D012313:RNA / Q000302:isolation & purification; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA
D015703:Antigens, CD; D001323:Autoantibodies; D017931:DNA Primers; D039663:Integrin beta4; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D012313:RNA