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  • Title: Analysis of the B-cell progenitor compartment at the level of single cells.
    Author: Ehlich A, Martin V, Müller W, Rajewsky K.
    Journal: Curr Biol; 1994 Jul 01; 4(7):573-83. PubMed ID: 7953531.
    Abstract:
    BACKGROUND: During B-cell development in the mouse, the VH, DH and JH elements of the immunoglobulin heavy chain (IgH) locus are rearranged, firstly by DH-JH joining, and then by VH-DHJH joining. In-frame ('productive') VHDHJH joints and DHJH joints in reading frame 2 (one of the three possible DH reading frames) allow the expression of mu and truncated mu chains (D mu proteins), respectively. The expression of such molecules from one of the two IgH loci of a cell is thought to interfere with VH-DHJH recombination on the other IgH locus, and to guide the cells through further development. RESULTS: We have developed a gene amplification assay that permits the examination of rearranged immunoglobulin genes in single cells. Using this assay, we monitored cells bearing DHJH and/or VHDHJH joints at early stages of development: in CD43+ B-cell progenitors, subdivided into fractions A, B, C and C' by flow cytometry, and in CD43- pre-B cells (fraction D). Fraction C was enriched for cells with two non-productive VHDHJH joints. Cells containing both a DHJH joint in DH reading frame 2 and a VHDHJH joint were not seen in any fraction. All fraction D cells harbored an in-frame VHDHJH joint. Cells with two productive VHDHJH joints appear to be selected against throughout development. CONCLUSIONS: Cells expressing D mu proteins appear to be arrested in development as a result of inhibited VH-DHJH joining. Expression of the mu chain is required for maturation into CD43- pre-B cells; accordingly, cells carrying two non-productive VHDHJH joints accumulate in the CD43+ compartment. Such a developmental arrest may also affect cells that express self-reactive VHDHJH antibody domains. Our results indicate further that allelic exclusion at the IgH locus is already established at the pre-B cell stage.
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