Mapping and regulation of the tumor-associated epitope recognized by monoclonal antibody RS-11.
We have previously described a rat monoclonal antibody, RS-11, which recognizes a tumor-associated antigen common to several species. In the present study, we have cloned and characterized the antigen recognized by RS-11. We screened a phage expression library prepared from HeLa cDNA and identified a clone that reacts with RS-11. DNA sequence analysis revealed that this clone contains sequences of keratin 18 (nucleotides 568-1196). We constructed several glutathione S-transferase fusion proteins and synthetic peptides based on this DNA sequence analysis and examined their reactivity with RS-11 to accurately map the RS-11 epitope. We determined that the epitope resides within a region of seven amino acids on the alpha-helix 2B domain of keratin 18 in which two amino acids (Leu(366) and Lys(370)) are completely conserved among intermediate filaments as well as other keratin members that are immunoreactive with RS-11. These two residues are sequentially discontinuous but spatially adjacent. The RS-11 epitope is constitutively present in human primary cultured hepatocytes; however, its immunoreactivity with RS-11 is up-regulated by malignant transformation or stimulation with either epidermal growth factor or transforming growth factor alpha.
Eto
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Division of Surgical Oncology, Massachusetts General Hospital and Harvard Medical School, Boston, Massachusetts 02114, USA.
Yoon
S S
SS
Bode
B P
BP
Kamidono
S
S
Makino
K
K
Saya
H
H
Nakamura
H
H
Tanabe
K K
KK
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D001483:Base Sequence; D015153:Blotting, Western; D002478:Cells, Cultured; D017931:DNA Primers; D004815:Epidermal Growth Factor / Q000494:pharmacology; D018604:Epitope Mapping; D000939:Epitopes / Q000737:chemistry / Q000276:immunology*; D006367:HeLa Cells; D006801:Humans; D008099:Liver / Q000166:cytology / Q000187:drug effects / Q000276:immunology; D008969:Molecular Sequence Data; D009369:Neoplasms / Q000276:immunology*; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D016211:Transforming Growth Factor alpha / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D000911:Antibodies, Monoclonal; D017931:DNA Primers; D000939:Epitopes; D012333:RNA, Messenger; D016211:Transforming Growth Factor alpha; D004815:Epidermal Growth Factor