Skewed primary Igκ repertoire and V-J joining in C57BL/6 mice: implications for recombination accessibility and receptor editing

J Immunol. 2012 Mar 1;188(5):2305-15. doi: 10.4049/jimmunol.1103484. Epub 2012 Jan 27.

Abstract

Previous estimates of the diversity of the mouse Ab repertoire have been based on fragmentary data as a result of many technical limitations, in particular, the many samples necessary to provide adequate coverage. In this study, we used 5'-coding end amplification of Igκ mRNAs from bone marrow, splenic, and lymph node B cells of C57BL/6 mice combined with amplicon pyrosequencing to assess the functional and nonfunctional Vκ repertoire. To evaluate the potential effects of receptor editing, we also compared V/J associations and usage in bone marrows of mouse mutants under constitutive negative selection or an altered ability to undergo secondary recombination. To focus on preimmune B cells, our cell sorting strategy excluded memory B cells and plasma cells. Analysis of ~90 Mbp, representing >250,000 individual transcripts from 59 mice, revealed that 101 distinct functional Vκ genes are used but at frequencies ranging from ~0.001 to ~10%. Usage of seven Vκ genes made up >40% of the repertoire. A small class of transcripts from apparently nonfunctional Vκ genes was found, as were occasional transcripts from several apparently functional genes that carry aberrant recombination signals. Of 404 potential V-J combinations (101 Vκs × 4 Jκs), 398 (98.5%) were found at least once in our sample. For most Vκ transcripts, all Jκs were used, but V-J association biases were common. Usage patterns were remarkably stable in different selective conditions. Overall, the primary κ repertoire is highly skewed by preferred rearrangements, limiting Ab diversity, but potentially facilitating receptor editing.

Publication types

  • Comparative Study
  • Research Support, N.I.H., Extramural

MeSH terms

  • Animals
  • Antibody Diversity / genetics
  • B-Lymphocyte Subsets / immunology
  • B-Lymphocyte Subsets / metabolism
  • Bone Marrow Cells / immunology
  • Bone Marrow Cells / metabolism
  • Female
  • Gene Rearrangement, B-Lymphocyte, Light Chain*
  • Immunoglobulin Joining Region / biosynthesis
  • Immunoglobulin Joining Region / genetics*
  • Immunoglobulin Joining Region / metabolism
  • Immunoglobulin Variable Region / biosynthesis
  • Immunoglobulin Variable Region / genetics*
  • Immunoglobulin Variable Region / metabolism
  • Immunoglobulin kappa-Chains / biosynthesis
  • Immunoglobulin kappa-Chains / genetics*
  • Immunoglobulin kappa-Chains / metabolism
  • Male
  • Mice
  • Mice, Inbred C57BL
  • Mice, Knockout
  • RNA Editing / genetics*
  • RNA Editing / immunology*
  • Recombination, Genetic / immunology*
  • Sequence Analysis, DNA

Substances

  • Immunoglobulin Joining Region
  • Immunoglobulin Variable Region
  • Immunoglobulin kappa-Chains