SIMULTANEOUS EXPRESSION OF GERMLINE GAMMA-1 AND EPSILON-IMMUNOGLOBULIN HEAVY-CHAIN TRANSCRIPTS IN SINGLE MURINE SPLENIC B-CELLS

While successive isotype switching can occur from IgM to IgE via IgG1, little is known about the pattern of germline transcript expression i n normal B cells committed to switching to multiple isotypes. In this study we define the relationship between germline transcript expressio n and immunoglobulin isotype expression and secretion in murine spleni c B cells. Following 7 days stimulation with LPS and IL-4, 10% of sing le cells secrete IgE and of these only 1 in 10 secrete IgE alone. In c ontrast, when cells were stimulated with LPS and IL-4 for 48 hr prior to sorting into clonal cultures, 71% secreted IgE alone. In an attempt to isolate switch intermediates, IgG1(+)IgM(-) cells were sorted and upon re-culture secreted predominantly IgG1 alone or IgG1 and IgE. The frequency of cells expressing germline epsilon transcripts was 34% fo r surface IgG1(+)IgM(-) expressing cells and 14% for surface IgG1(-)Ig M(+) negative cells. Furthermore, analysis of single surface IgG1(+)Ig M(-) cells demonstrated that these cells can co-express germline gamma 1 and epsilon transcripts. Sorting of IgG1(-)IgM(+) cells according t o surface expression of the integral membrane proteoglycan, Syndecan, demonstrated that IgM(+)Syndecan(+) cells had a lower frequency of ger mline transcript expression and a lower frequency of isotype switching (32%) compared to IgM(+)Syndecan(-) cells (83%). Collectively, these finding show that murine B cells can switch successively from IgM to I gE via IgG1 and that IgG1 expressing intermediates express germline tr anscripts. Furthermore, Syndecan expression appears to be linked to ge rmline C-H transcription and isotype switch commitment. (C) 1998 Elsev ier Science Ltd. All rights reserved.