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Abbreviated junctional sequences impoverish antibody diversity in urodele amphibians
Insights
Axolotl heavy chain CDR3 diversity is exceptionally low due to limited somatic recombination and extensive germline gene integration. This molecular mechanism contributes to the poor antibody responses observed in urodele amphibians.
Area of Science:
- Immunology
- Molecular Biology
- Amphibian Biology
Background:
- The heavy chain CDR3 loop is crucial for antibody diversity and is highly variable in mammals and some amphibians.
- Urodele amphibians exhibit lifelong poor immunocompetence and larval-like antibody responses.
- The molecular basis for this immunodeficiency in urodeles remains largely unknown.
Purpose of the Study:
- To investigate the molecular mechanisms underlying the limited antibody diversity in axolotls.
- To analyze the junctional diversity of heavy chain CDR3 sequences in the axolotl.
- To compare axolotl antibody gene rearrangement with that of other species.
Main Methods:
- Cloning of germline VH genes from the axolotl.
- Analysis of heavy chain gene rearrangements using reverse-transcriptase PCR.
- Sequence analysis of CDR3 loop junctional diversity.
Main Results:
- Axolotl CDR3 sequences showed significantly lower variability compared to newborn mice and Xenopus tadpoles.
- Only 29% of the axolotl CDR3 loop comprised somatically generated sequences, compared to 39-57% in other species.
- Extensive integration of germline JH sequences and shorter junctional sequences were observed in axolotls.
Conclusions:
- The limited heavy chain CDR3 diversity in axolotls is a key molecular factor contributing to their deficient antibody responses.
- This finding provides insight into the poor immunocompetence observed in urodele amphibians.
- The study highlights distinct mechanisms of immune system development in amphibians.
Abstract:
Of the six complementarity-determining regions (CDR) forming the structure of the Ab combining site, CDR3 of heavy chain is the most variable in length and sequence. Diversity of this loop is determined by the number of gene segments involved, extent of addition to or deletion from the joining genes, and imprecision of the site of recombination. In neonatal mice and Xenopus tadpoles, the last two factors occur less frequently than in adults, which in tadpoles result in low affinity Ab responses that do not mature. In contrast, adult urodele amphibians make larval-like responses and are notorious for lifelong poor immunocompetence. The mechanism for this is not known, and in this study we cloned germline VH genes from the axolotl and obtained rearrangements to these VH gene segments by reverse-transcriptase PCR. These sequences were analyzed for heavy chain junctional diversity and found to be even less variable than that in newborn mouse or Xenopus tadpoles, although for different reasons. Only 29% of the CDR3 loop in the axolotl consisted of somatically generated sequences, compared with 44% in tadpole, 39% in newborn mice, and 57% in both adult mice and Xenopus. This distinguishing feature of axolotl CDR3 results not only from shorter junctional sequences, but also unusually extensive integration of germline JH sequence. As the CDR3 loop is the most important portion of the Ig sequence for determining Ab combining site diversity, our data provide the molecular basis for a contributing factor in the deficient urodele amphibian Ab responses.