Alpha-enolase is restricted to basal cells of stratified squamous epithelium.
We have developed a monoclonal antibody against a 50-kDa protein that binds preferentially to basal cells in the limbus of rat, rabbit, and human corneas (J. D. Zieske, G. Bukusoglu, and M. A. Yankauckas, Invest. Ophthalmol. Visual Sci. 33, 143-152, 1992). Here we report on the purification and identification of the antigen. The 50-kDa antigen was purified from rabbit limbal and corneal epithelium using HPLC methodology including anion exchange (DEAE) followed by reverse-phase (C18) chromatography. The purified 50-kDa protein was then digested with endoproteinase Lys-C, and a reproducible profile comprising approximately 20 peptides was observed by reverse-phase HPLC of the digest. Sequence analysis of five peptides ranging in length from 4 to 20 residues revealed that the 50-kDa protein was alpha-enolase, a glycolytic enzyme. Overall, 57 amino acids were identified with a 95% sequence homology. Localization of alpha-enolase in rat epithelium by immunofluorescence microscopy demonstrated that simple epithelium contained low or undetectable levels of the enzyme. Stratified squamous epithelium, however, showed high levels of alpha-enolase, which was localized specifically to cells of the basal layer. Epidermal, corneal limbal, oral mucosal, vaginal, and laryngeal epithelium all showed cytoplasmic binding specific to the basal cells. These data indicate that the glycolytic enzyme alpha-enolase is preferentially localized in the basal cell layer of stratified squamous epithelium and suggest that glycolytic activity is concentrated in these cells. The localization pattern suggests that a major change in metabolism occurs as cells leave the mitotically active basal cell layer and migrate toward terminal differentiation in the suprabasal cell layers.
Zieske
J D
JD
Eye Research Institute, Harvard Medical School, Boston, Massachusetts 02114.
Bukusoglu
G
G
Yankauckas
M A
MA
Wasson
M E
ME
Keutmann
H T
HT
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D002454:Cell Differentiation; D003315:Cornea / Q000201:enzymology; D003593:Cytoplasm / Q000201:enzymology; D004848:Epithelium / Q000201:enzymology*; D008969:Molecular Sequence Data; D009061:Mouth Mucosa / Q000201:enzymology; D010751:Phosphopyruvate Hydratase / Q000032:analysis* / Q000302:isolation & purification; D051381:Rats; D011919:Rats, Inbred Strains; D012867:Skin / Q000201:enzymology
D000911:Antibodies, Monoclonal; D010751:Phosphopyruvate Hydratase