RESUMO
The Mexican axolotl V(H) segments associated with the Igh C mu and C nu 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(H)9) to 11-19 (V(H)2), 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 H mu and H nu chains; V(H)1, V(H)6, and V(H)11 were underutilized; and the majority of the V(H) segments belonged to the V(H)7, V(H)8, and V(H)9 families. Most of the nine J(H) segments seemed to be randomly used, except J(H)6 and J(H)9, which were found only once in 79 clones.
Assuntos
Ambystoma mexicanum/imunologia , Diversidade de Anticorpos , Cadeias Pesadas de Imunoglobulinas/química , Região Variável de Imunoglobulina/química , Família Multigênica/imunologia , Ambystoma mexicanum/genética , Sequência de Aminoácidos , Animais , Diversidade de Anticorpos/genética , Rearranjo Gênico do Linfócito B/imunologia , Humanos , Cadeias Pesadas de Imunoglobulinas/genética , Região de Junção de Imunoglobulinas/química , Região de Junção de Imunoglobulinas/genética , Região Variável de Imunoglobulina/genética , Cadeias mu de Imunoglobulina/química , Cadeias mu de Imunoglobulina/genética , Camundongos , Dados de Sequência Molecular , Filogenia , XenopusRESUMO
The immune capacity of young and adult axolotls (Ambystoma mexicanum) was evaluated by examining the combinatorial and junctional diversity of the VH chain. A large number of VDJ rearrangements isolated from 2.5-, 3.5-, 10-, and 24-month-old animals were sequenced. Six JH segments were identified with the canonical structure of all known vertebrate JHs, including the conserved Trp103-Gly104-X-Gly106 motif. Four core DH-like sequences were used by most (80%) of the VDJ junctions. These G-rich sequences had structures reminiscent of the TCRB DB sequences, and were equally used in their three reading frames. About 25% of the Igh, VDJ junctions from 3.5-month-old axolotls were out of frame, but most rearrangements were in frame at 10 and 24 months, suggesting that there is active selection of the productively rearranged Igh chains in the developing animals. There was no significant difference between the size of CDR3 in young (3.5 months) and subadult (10 months) axolotls (mean: 8.5 amino acids). However, the CDR3 loop was 1 amino acid longer in 2-year-old adult animals (mean: 9.5 residues). Several pairs of identical VDJ/CDR3 sequences were shared between 3.5-month-old individually analyzed axolotls, or between groups of axolotl of different ages. These identical rearrangements might be provided by the selection of some B-cell clones important for species survival, although the probability that different 3.5-month-old axolotl larvae would produce identical junctions seems very low, considering their limited number of B cells (less than 10(5)). The high frequency of tyrosine residues and the paucity of charged residues in the axolotl CDR3 loops may explain the polyreactivity of natural antibodies, and also clarify why it is so difficult to raise specific antibodies against soluble antigens.
Assuntos
Ambystoma/genética , Rearranjo Gênico , Cadeias Pesadas de Imunoglobulinas/genética , Região de Junção de Imunoglobulinas/genética , Região Variável de Imunoglobulina/genética , Ambystoma/crescimento & desenvolvimento , Ambystoma/imunologia , Animais , Variação Genética , Cadeias mu de Imunoglobulina/genética , Imunoglobulinas/genética , Especificidade da EspécieRESUMO
Cholera is a topical infection of the intestinal tract and rarely causes extraintestinal disease. The gallbladder has been proposed to be the reservoir of this organism. We present a patient from Bolivia who developed symptoms of acute cholecystitis and whose bile culture grew Vibrio cholerae 01 El tor.