Abstract
Using the recently developed ELISPOT cloning methodology, we obtained cDNA clone S35 coding for the Ag epitope recognized by a murine sarcoma Meth A-specific CTL clone AT-1. Analysis of truncated S35 constructs and overlapping peptides revealed that the peptide epitope was LGAEAIFRL. AT-1 CTL lysed peptide-pulsed CMS8 cells at a nanomolar concentration, and the peptide strongly stimulated IFN-gamma production in AT-1 CTL. Sequence homology indicated that the S35 was derived from a mouse homologue of human retinoic acid-regulated nuclear matrix-associated protein (ramp). The ramp gene consisted of 15 exons. The majority of the ramp mRNA was the transcript normally spliced between exons 14 and 15, but a minor population of mRNA with an extended exon 14 was also present in Meth A cells. The epitope was derived from the newly created open reading frame, which resulted from extension of exon 14 after splicing of the adjacent intronic sequence.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Amino Acid Sequence
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Animals
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Antigens, Neoplasm / biosynthesis
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Antigens, Neoplasm / genetics
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Antigens, Neoplasm / isolation & purification
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Antigens, Neoplasm / metabolism
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Base Sequence
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Carrier Proteins / biosynthesis
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Carrier Proteins / genetics
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Carrier Proteins / isolation & purification
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Carrier Proteins / metabolism
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Clone Cells
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Cloning, Molecular
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Cytotoxicity Tests, Immunologic
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DNA-Binding Proteins / biosynthesis
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DNA-Binding Proteins / genetics
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DNA-Binding Proteins / isolation & purification
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DNA-Binding Proteins / metabolism
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Enzyme-Linked Immunosorbent Assay
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Epitopes, T-Lymphocyte / biosynthesis*
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Epitopes, T-Lymphocyte / genetics*
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Epitopes, T-Lymphocyte / isolation & purification
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Epitopes, T-Lymphocyte / metabolism
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Exons / genetics*
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Exons / immunology
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Gene Library
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Humans
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Interferon-gamma / analysis
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Methylcholanthrene*
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Mice
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Mice, Inbred BALB C
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Mice, Inbred DBA
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Molecular Sequence Data
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Sarcoma, Experimental / chemically induced
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Sarcoma, Experimental / immunology*
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T-Lymphocytes, Cytotoxic / immunology*
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Transcription Factors
Substances
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Antigens, Neoplasm
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Carrier Proteins
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DNA-Binding Proteins
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Epitopes, T-Lymphocyte
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Transcription Factors
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ZMYM2 protein, human
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Zmym2 protein, mouse
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Methylcholanthrene
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Interferon-gamma