The 105-kDa basement membrane autoantigen p105 is N-terminally homologous to a tumor-associated antigen.
Certain constitutive skin basement membrane components, such as bullous pemphigoid antigens and epidermolysis bullosa acquisita antigen, were discovered because they were targeted by an autoimmune reaction. We aimed to purify and characterize a 105-kDa skin basement membrane protein termed p105 recognized by autoantibodies (anti-p105) from patients with a unique immune-mediated subepidermal blistering skin disease. A simian virus 40-transformed human fibroblast cell line that synthesizes and secretes p105 was utilized as the protein source. p105 was partially purified by salt-gradient fractionation of serum-free conditioned medium through a Mono Q anion-exchange column and by examining each fraction with protein staining and immunoblotting against anti-p105. p105 was isolated from polyacrylamide gel electrophoresis gels, blotted onto polyvinylidene difluoride membrane, and subjected to protein microsequencing. The 20 microsequenced N-terminal amino acids exhibited no homology to known basement membrane proteins but exhibited a 70% homology to a 90-kDa tumor-associated antigen. Antibodies raised against a peptide generated from these amino acid sequences reacted to a 105-kDa western-blotted keratinocyte and fibroblast protein and a basement membrane component. p105 resisted digestion by glycosidases chondroitinase ABC, neuraminidase, and N-glycosidase F but was cleaved by protease V8 to antigenic fragments of 22 kDa and 14 kDa. The synthesis of p105 was inhibited by cycloheximide. We conclude that p105 is a unique basement membrane component produced by both keratinocytes and fibroblasts.
Chan
L S
LS
Immunodermatology Division, Department of Dermatology, Northwestern University Medical School, Chicago, Illinois 60611, U.S.A.
Woodley
D T
DT
D000595:Amino Acid Sequence; D000951:Antigens, Neoplasm / Q000235:genetics*; D001324:Autoantigens / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D001485:Basement Membrane / Q000276:immunology*; D002852:Chromatography, Ion Exchange; D006801:Humans; D008969:Molecular Sequence Data; D008970:Molecular Weight; D017386:Sequence Homology, Amino Acid
D000951:Antigens, Neoplasm; D001324:Autoantigens