The molecular basis for functional loss of T antigen cytotoxic T lymphocyte recognition sites from SV40-transformed antigenic site loss variant cells

Miscellaneous · Wed Jan 01 00:00:00 EST 1992

OSTI ID:6084777

Lill, N L

The 94 kD large tumor (T) antigen present in cells transformed by the DNA tumor virus SV40 contains four H-2D[sup b] restricted CTL recognition epitopes recognized by site-specific CTL clones Y-1, Y-2, Y- 3 and Y-5. Epitopes recognized by these CTL map to T antigen amino acid residues 207-215 (Site I), 223-231 (Sites II and III), and 489-497 (Site V), respectively. Antigenic site loss variant cells lacking one or more epitopes without altered H-2D[sup b] class I antigen expression were previously selected by coculturing SV40-transformed H-2D[sup b] cells with CTL clones. Clonally derived cell lines from variant cell populations were subjected to immunological and molecular analyses to identify the molecular basis for CTL recognition site loss. DNA amplification and direct sequencing of variant cell epitope coding regions were performed. Variant cells selected for resistance to CTL clone Y-1 (K-1; K-1,4,5; K-3,1) carry deleted SV40 genomes lacking Site I, II and III coding sequences. Point mutations exist within the Site II/Ill coding region of Y-2-/Y-3-resistant cell lines K-2 and K-3. Separate mutations were identified within independent Y-5-selected populations (K-5 and K-1,4,5). Mutations were reconstructed into wild type T antigen coding sequences by site-directed mutagenesis. The mutant T antigens were expressed in H-2D[sup b] cells. In cytotoxicity assays, each substitution induced loss of a functional CTL recognition epitope. The identical amino acid substitutions in synthetic peptides representing Sites II, III and V also induced loss of peptide recognition by site-specific CTL clones. Functional synthetic peptide competition studies indicated that mutant peptides could bind to MHC class I H-2D[sup b] as well as their wild type counterparts. Therefore, recognition of each MHC/substituted epitope complex by the CTL TCR was compromised. Genetic changes cause the loss of CTL recognition sites from tumor antigens, possibly allowing for transformed cell escape.

• Research Organization: Pennsylvania State Univ., McKeesport, PA (United States)
• OSTI ID: 6084777
• Resource Relation: Other Information: Thesis (Ph.D.)
• Country of Publication: United States
• Language: English

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
ANTIGENS
GENE MUTATIONS
NEOPLASMS
AMINO ACID SEQUENCE
DNA HYBRIDIZATION
DNA SEQUENCING
HISTOCOMPATIBILITY COMPLEX
LYMPHOCYTES
ONCOGENIC TRANSFORMATIONS
ANIMAL CELLS
BIOLOGICAL MATERIALS
BLOOD
BLOOD CELLS
BODY FLUIDS
CELL TRANSFORMATIONS
CONNECTIVE TISSUE CELLS
DISEASES
HYBRIDIZATION
LEUKOCYTES
MATERIALS
MOLECULAR STRUCTURE
MUTATIONS
SOMATIC CELLS
STRUCTURAL CHEMICAL ANALYSIS
550400* - Genetics
551000 - Physiological Systems