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Structure, Diversity, and Repertoire of V_H Families in the Mexican Axolotl¹

Comparative Immunology Group, National Centre for Scientific Research, Pierre and Marie Curie University, Paris, France

The Mexican axolotl V_H segments associated with the Igh Cµ and C isotypes were isolated from anchored PCR libraries prepared from spleen cell cDNA. The eight new V_H segments found bring the number of V_H families in the axolotl to 11. Each V_H had the canonical structural features of vertebrate V_H segments, including residues important for the correct folding of the Ig domain. The distribution of ser AGC/T (AGY) and TCN codons in axolotl V_H genes was biased toward AGY in complementarity-determining region-1 (CDR1) and TCN in framework region-1 (FR1); there were no ser residues in the FR2 region. Thus, the axolotl CDR1 region is enriched in DNA sequences forming potential hypermutation hot spots and is flanked by DNA sequences more resistant to point mutation. There was no significant bias toward AGY in CDR2. Southern blotting using family-specific V_H probes showed restriction fragments from 1 (V_H9) to 11–19 (V_H2), and the total number of V_H genes was 44 to 70, depending on the restriction endonuclease used. The V_H segments were not randomly used by the Hµ and H chains; V_H1, V_H6, and V_H11 were underutilized; and the majority of the V_H segments belonged to the V_H7, V_H8, and V_H9 families. Most of the nine J_H segments seemed to be randomly used, except J_H6 and J_H9, which were found only once in 79 clones.

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