Monoclonal antibody toward lysosomal cathepsin B cross-reacts preferentially with distinct histone classes.
1. A set of monoclonal antibodies (Mab) was prepared against cathepsin B (CB) from rat preputial-gland, an organ characterized by rapidly-renewing cell populations, which is a uniquely enriched source of lysosomal enzymes, including CB. Minute amounts of CB are known to be transferred abruptly to the nuclear compartment in a variety of activated cells. 2. Since, on the basis of its stringent substrate requirements, CB was expected to function at limited protein loci in chromatin, Mab Line II-B4 was used to probe Western blots of chromatin fractions and selected proteins. 3. The Mab, which was not directed against the active site of CB, cross-reacted preferentially with histones 3 and 4 (H3 and H4) in acid-soluble fractions of chromatin from rat preputial-gland. Line II-B4 also recognized H3 and H4 selectively in calf thymus histones and among histones purified from a wide range of sources from yeast to man. HMG 1 was minimally immunoreactive among preputial gland constituents and carbonic anhydrase (CA) was also sensitive to the Mab. 4. The common determinants were not shared by any of the H1 series, nor by H2A, H2B, protein A24 or a wide range of natural and synthetic products. 5. Origin of the antigenicity was traced by chemical modifications of H3, H4 and CA to the critical contribution of arginine and hydrophobic amino acid residues in its immediate environment, indicating that Line II-B4 may be directed against an epitope comprising the specific binding-site of CB and its selective substrate(s). 6. These data suggest that certain highly conserved cellular constituents may be uniquely vulnerable to limited proteolysis in preproliferative cells responding to mitotic signals.
Seeler
B J
BJ
Department of Biology, University of California, Los Angeles 90024.
Horton
M J
MJ
Szego
C M
CM
DeLange
R J
RJ
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000917:Antibody Formation; D002256:Carbonic Anhydrases / Q000032:analysis / Q000276:immunology; D002401:Cathepsin B / Q000032:analysis / Q000276:immunology*; D002417:Cattle; D002843:Chromatin / Q000032:analysis / Q000276:immunology; D002987:Clitoris / Q000201:enzymology; D003429:Cross Reactions; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D000939:Epitopes / Q000032:analysis; D005260:Female; D006657:Histones / Q000276:immunology*; D008247:Lysosomes / Q000201:enzymology* / Q000276:immunology; D051381:Rats; D011919:Rats, Inbred Strains; D013045:Species Specificity; D013950:Thymus Gland / Q000201:enzymology
D000911:Antibodies, Monoclonal; D002843:Chromatin; D000939:Epitopes; D006657:Histones; D002401:Cathepsin B; D002256:Carbonic Anhydrases