Human keratinocytes contain keratin filaments that are glycosylated with keratan sulfate.
We have reported strong intracytoplasmic immunoreactivity for anti-keratan sulfate monoclonal antibodies in human keratinocytes. Consequently, ultrastructural immunogold studies were undertaken to identify the cytoplasmic components responsible for this immunoreactivity. Immunogold labeling of cultured keratinocytes identified keratin filaments as a source of keratan sulfate epitopes. Immunolabeling also marked desmosomal cytoplasmic plaques and amorphous electron-dense bodies. These observations were confirmed for epidermal keratinocytes of human skin. Further evidence was obtained that some keratins have epitopes for anti-keratan sulfate antibodies by Western blot analyses following fractionation of proteins by sodium dodecyl sulfate gel electrophoresis. Four bands were detected with apparent molecular weights of 58, 54, 50, and 48 kDa that reacted with anti-keratin monoclonal antibody mixture AE1/AE3. Keratins of 58 and 56 kDa immunoreacted with anti-keratan sulfate antibody 8C2 while all four keratins immunoreacted with anti-keratan sulfate antibody 5D4. Endo-beta-D-galactosidase and keratanase, enzymes that degrade keratan sulfate, removed all, or a portion, of specific keratan sulfate epitopes from keratin extracts. These results demonstrate that a portion of the cytoplasmic anti-keratan sulfate immunoreactivity is due to keratins that are glycosylated with carbohydrates that contain keratan sulfate epitopes or that keratan sulfate-containing molecules bind or comigrate in SDS-polyacrylamide gels with cytokeratins.
Schafer
I A
IA
Department of Pediatrics, Case Western Reserve University, MetroHealth Medical Center, Cleveland, Ohio 44109.
Sorrell
J M
JM
D008841:Actin Cytoskeleton / Q000737:chemistry* / Q000378:metabolism* / Q000648:ultrastructure; D000911:Antibodies, Monoclonal / Q000276:immunology; D015153:Blotting, Western; D002478:Cells, Cultured; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes / Q000276:immunology; D006031:Glycosylation; D006801:Humans; D007150:Immunohistochemistry; D007632:Keratan Sulfate / Q000276:immunology / Q000378:metabolism*; D015603:Keratinocytes / Q000166:cytology / Q000378:metabolism / Q000648:ultrastructure*; D007633:Keratins / Q000032:analysis* / Q000276:immunology / Q000378:metabolism*; D016253:Microscopy, Immunoelectron
D000911:Antibodies, Monoclonal; D000939:Epitopes; D007633:Keratins; D007632:Keratan Sulfate