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Epitope-dependent selection of highly restricted or diverse T cell receptor repertoires in response to persistent infection by Epstein-Barr virus.

Campos-Lima PO, Levitsky V, Imreh MP, Gavioli R, Masucci MG - J. Exp. Med. (1997)

Bottom Line: The response to the subdominant epitope (EBNA4 399-408, designated AVF) was highly restricted with conserved Vbeta usage and identical length and amino acid motifs in the third complementarity-determining regions (CDR3), while a broad repertoire using different combinations of TCR-alpha/beta V and J segments and CDR3 regions was selected by the immunodominant epitope (EBNA4 416-424, designated IVT).Distinct patterns of interaction with the A11-peptide complex were revealed for each AVF- or IVT-specific TCR clonotype by alanine scanning mutagenesis analysis.Thus, comparison of the memory response to two epitopes derived from the same viral antigen and presented through the same MHC class I allele suggests that immunodominance may correlate with the capacity to maintain a broad TCR repertoire.

View Article: PubMed Central - PubMed

Affiliation: Microbiology and Tumor Biology Center, Karolinska Institute, S-171 77, Stockholm, Sweden.

ABSTRACT
The T cell receptor (TCR) repertoires of cytotoxic responses to the immunodominant and subdominant HLA A11-restricted epitopes in the Epstein-Barr virus (EBV) nuclear antigen-4 were investigated in four healthy virus carriers. The response to the subdominant epitope (EBNA4 399-408, designated AVF) was highly restricted with conserved Vbeta usage and identical length and amino acid motifs in the third complementarity-determining regions (CDR3), while a broad repertoire using different combinations of TCR-alpha/beta V and J segments and CDR3 regions was selected by the immunodominant epitope (EBNA4 416-424, designated IVT). Distinct patterns of interaction with the A11-peptide complex were revealed for each AVF- or IVT-specific TCR clonotype by alanine scanning mutagenesis analysis. Blocking of cytotoxic function by antibodies specific for the CD8 coreceptor indicated that, while AVF-specific TCRs are of high affinity, the oligoclonal response to the IVT epitope includes both low- and high-affinity TCRs. Thus, comparison of the memory response to two epitopes derived from the same viral antigen and presented through the same MHC class I allele suggests that immunodominance may correlate with the capacity to maintain a broad TCR repertoire.

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Nucleotide sequence of the α and β chains V(D)J junctional regions and deduced amino acid sequence. (A) AVF-specific TCRs; (B) IVT-specific TCRs. For each TCR type, the nucleotide sequence and the deduced amino acid sequence in single letter code of the CDR3 equivalent loop,  defined according to Chothia et al. (33), is shown putatively supported by two framework branches (FW). Only deviations from the consensus FW sequences are indicated. N nucleotide additions are underlined and TCRBD1 and BD2 germinal sequences are highlighted in bold. The conserved framework Cys residues in position 90 of the Vα and 92 of the Vβ chains are indicated.
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Figure 1: Nucleotide sequence of the α and β chains V(D)J junctional regions and deduced amino acid sequence. (A) AVF-specific TCRs; (B) IVT-specific TCRs. For each TCR type, the nucleotide sequence and the deduced amino acid sequence in single letter code of the CDR3 equivalent loop, defined according to Chothia et al. (33), is shown putatively supported by two framework branches (FW). Only deviations from the consensus FW sequences are indicated. N nucleotide additions are underlined and TCRBD1 and BD2 germinal sequences are highlighted in bold. The conserved framework Cys residues in position 90 of the Vα and 92 of the Vβ chains are indicated.

Mentions: The TCR α and β chain usage of a representative panel of 19 AVF-specific clones from the four donors and 24 IVT-specific clones from three of the donors was determined by polymerase chain reaction (PCR)-assisted cDNA amplification. All AVF-specific clones expressed TCRBV3S1 (Table 2). This was paired to TCRAV1S4 in clones derived from three of the donors (BK, CA, and ZA; AVF type I), while all clones from EA expressed the closely related TCRAV16S1 (AVF type II). Sequencing of the PCR products demonstrated the regular rearrangement of BV3S1 to BJ2S2, AV1S4 to AJ21, and AV16S1 to AJ40. Comparison of the predicted amino acid sequences of the 19 Vα chains revealed CDR3 regions of equal length and identical protein sequence in all clones expressing a given VJ combination, with variations in codon usage restricted to the N regions (Fig. 1 A). The codon for Asp at position 94 was generated by N additions and was consistently preceded by the codons for Gly in clones expressing AV1S4J21, or Ser in the AV16S1J40 clones. The Vβ sequences of clones expressing the AVF type I receptor also showed a remarkable similarity. A conserved Thr-Ser-Ala motif was generated in the NDN region of all but one sequence that carried a Thr to Ala substitution. This motif was not present in the junctional region of the AVF type II TCRs, although a strict requirement for length maintenance in the CDR3 loop was suggested by the replacement of two germ line encoded Ser residues at positions 94 and 95 with ND encoded Thr and Gly. A remarkable conservation of TCR Vβ usage was previously reported in EBV-specific CTLs that recognize the EBNA3 325-333 epitope. Identical amino acid and nucleotide sequences were detected in the Vβ CDR3 regions of the EBNA3 325-333-specific TCRs isolated from four individuals (18). However, the usage of germ line sequences indicates that a preferentially favored rearrangement may contribute to the conservation of this response. This is clearly not the case in the AVF-specific response. The significant differences between the α and β chain nucleotide sequences and the regular occurrence of N additions confirm the origin of AVF-specific TCRs from independent recombination events even in clones derived from the same individuals (Fig. 1, donor BK). This finding stresses the importance of the conserved features of the AVF-specific TCRs for interaction with the A11–peptide complex. A different scenario was revealed by analysis of the repertoire specific for the immunodominant IVT epitope. Each of the 24 IVT-specific clones analyzed expressed one of nine distinct TCR-α/β heterodimers (Table 2). Four IVT-specific clonotypes were identified in donor BK, one in CA, and four in EA with no identical or even similar TCR isolated from more than one donor. Sequence comparison showed no preferential usage of TCRJ segments and no apparent conservation of CDR3 amino acid composition. Different sets of N or NDN additions resulted in the generation of CDR3 loops of variable length and amino acid composition (Fig. 1 B).


Epitope-dependent selection of highly restricted or diverse T cell receptor repertoires in response to persistent infection by Epstein-Barr virus.

Campos-Lima PO, Levitsky V, Imreh MP, Gavioli R, Masucci MG - J. Exp. Med. (1997)

Nucleotide sequence of the α and β chains V(D)J junctional regions and deduced amino acid sequence. (A) AVF-specific TCRs; (B) IVT-specific TCRs. For each TCR type, the nucleotide sequence and the deduced amino acid sequence in single letter code of the CDR3 equivalent loop,  defined according to Chothia et al. (33), is shown putatively supported by two framework branches (FW). Only deviations from the consensus FW sequences are indicated. N nucleotide additions are underlined and TCRBD1 and BD2 germinal sequences are highlighted in bold. The conserved framework Cys residues in position 90 of the Vα and 92 of the Vβ chains are indicated.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2198955&req=5

Figure 1: Nucleotide sequence of the α and β chains V(D)J junctional regions and deduced amino acid sequence. (A) AVF-specific TCRs; (B) IVT-specific TCRs. For each TCR type, the nucleotide sequence and the deduced amino acid sequence in single letter code of the CDR3 equivalent loop, defined according to Chothia et al. (33), is shown putatively supported by two framework branches (FW). Only deviations from the consensus FW sequences are indicated. N nucleotide additions are underlined and TCRBD1 and BD2 germinal sequences are highlighted in bold. The conserved framework Cys residues in position 90 of the Vα and 92 of the Vβ chains are indicated.
Mentions: The TCR α and β chain usage of a representative panel of 19 AVF-specific clones from the four donors and 24 IVT-specific clones from three of the donors was determined by polymerase chain reaction (PCR)-assisted cDNA amplification. All AVF-specific clones expressed TCRBV3S1 (Table 2). This was paired to TCRAV1S4 in clones derived from three of the donors (BK, CA, and ZA; AVF type I), while all clones from EA expressed the closely related TCRAV16S1 (AVF type II). Sequencing of the PCR products demonstrated the regular rearrangement of BV3S1 to BJ2S2, AV1S4 to AJ21, and AV16S1 to AJ40. Comparison of the predicted amino acid sequences of the 19 Vα chains revealed CDR3 regions of equal length and identical protein sequence in all clones expressing a given VJ combination, with variations in codon usage restricted to the N regions (Fig. 1 A). The codon for Asp at position 94 was generated by N additions and was consistently preceded by the codons for Gly in clones expressing AV1S4J21, or Ser in the AV16S1J40 clones. The Vβ sequences of clones expressing the AVF type I receptor also showed a remarkable similarity. A conserved Thr-Ser-Ala motif was generated in the NDN region of all but one sequence that carried a Thr to Ala substitution. This motif was not present in the junctional region of the AVF type II TCRs, although a strict requirement for length maintenance in the CDR3 loop was suggested by the replacement of two germ line encoded Ser residues at positions 94 and 95 with ND encoded Thr and Gly. A remarkable conservation of TCR Vβ usage was previously reported in EBV-specific CTLs that recognize the EBNA3 325-333 epitope. Identical amino acid and nucleotide sequences were detected in the Vβ CDR3 regions of the EBNA3 325-333-specific TCRs isolated from four individuals (18). However, the usage of germ line sequences indicates that a preferentially favored rearrangement may contribute to the conservation of this response. This is clearly not the case in the AVF-specific response. The significant differences between the α and β chain nucleotide sequences and the regular occurrence of N additions confirm the origin of AVF-specific TCRs from independent recombination events even in clones derived from the same individuals (Fig. 1, donor BK). This finding stresses the importance of the conserved features of the AVF-specific TCRs for interaction with the A11–peptide complex. A different scenario was revealed by analysis of the repertoire specific for the immunodominant IVT epitope. Each of the 24 IVT-specific clones analyzed expressed one of nine distinct TCR-α/β heterodimers (Table 2). Four IVT-specific clonotypes were identified in donor BK, one in CA, and four in EA with no identical or even similar TCR isolated from more than one donor. Sequence comparison showed no preferential usage of TCRJ segments and no apparent conservation of CDR3 amino acid composition. Different sets of N or NDN additions resulted in the generation of CDR3 loops of variable length and amino acid composition (Fig. 1 B).

Bottom Line: The response to the subdominant epitope (EBNA4 399-408, designated AVF) was highly restricted with conserved Vbeta usage and identical length and amino acid motifs in the third complementarity-determining regions (CDR3), while a broad repertoire using different combinations of TCR-alpha/beta V and J segments and CDR3 regions was selected by the immunodominant epitope (EBNA4 416-424, designated IVT).Distinct patterns of interaction with the A11-peptide complex were revealed for each AVF- or IVT-specific TCR clonotype by alanine scanning mutagenesis analysis.Thus, comparison of the memory response to two epitopes derived from the same viral antigen and presented through the same MHC class I allele suggests that immunodominance may correlate with the capacity to maintain a broad TCR repertoire.

View Article: PubMed Central - PubMed

Affiliation: Microbiology and Tumor Biology Center, Karolinska Institute, S-171 77, Stockholm, Sweden.

ABSTRACT
The T cell receptor (TCR) repertoires of cytotoxic responses to the immunodominant and subdominant HLA A11-restricted epitopes in the Epstein-Barr virus (EBV) nuclear antigen-4 were investigated in four healthy virus carriers. The response to the subdominant epitope (EBNA4 399-408, designated AVF) was highly restricted with conserved Vbeta usage and identical length and amino acid motifs in the third complementarity-determining regions (CDR3), while a broad repertoire using different combinations of TCR-alpha/beta V and J segments and CDR3 regions was selected by the immunodominant epitope (EBNA4 416-424, designated IVT). Distinct patterns of interaction with the A11-peptide complex were revealed for each AVF- or IVT-specific TCR clonotype by alanine scanning mutagenesis analysis. Blocking of cytotoxic function by antibodies specific for the CD8 coreceptor indicated that, while AVF-specific TCRs are of high affinity, the oligoclonal response to the IVT epitope includes both low- and high-affinity TCRs. Thus, comparison of the memory response to two epitopes derived from the same viral antigen and presented through the same MHC class I allele suggests that immunodominance may correlate with the capacity to maintain a broad TCR repertoire.

Show MeSH
Related in: MedlinePlus