Evidence for Ig Light Chain Isotype Exclusion in Shark B Lymphocytes Suggests Ordered Mechanisms

摘要

Unlike most vertebrates, the shark IgL gene organization precludes secondary rearrangements that delete self-reactive VJ rearranged genes. Nurse sharks express four L chain isotypes, κ, λ, σ, and σ-2, encoded by 35 functional minigenes or clusters. The sequence of gene activation/expression and receptor editing of these isotypes have not been studied. We therefore investigated the extent of isotypic exclusion in separated B cell subpopulations. Surface Ig (sIg)κ–expressing cells, isolated with mAb LK14 that recognizes Cκ, carry predominantly nonproductive rearrangements of other L chain isotypes. Conversely, after depletion with LK14, sIgM⁺ cells contained largely nonproductive κ and enrichment for in-frame VJ of the others. Because some isotypic inclusion was observed at the mRNA level, expression in the BCR was examined. Functional λ mRNA was obtained, as expected, from the LK14-depleted population, but was also in sIgκ⁺ splenocytes. Whereas λ somatic mutants from the depleted sample displayed evidence of positive selection, the λ genes in sIgκ⁺ cells accumulated bystander mutations indicating a failure to express their products at the cell surface in association with the BCR H chain. In conclusion, a shark B cell expresses one L chain isotype at the surface and other isotypes as nonproductive VJ, sterile transcripts, or in-frame VJ whose products may not associate with the H chain. Based on the mRNA content found in the B cell subpopulations, an order of L chain gene activation is suggested as: σ-2 followed by κ, then σ and λ.