Arthritis & Rheumatism, Volume 60,
October 2009 Abstract Supplement
The 2009 ACR/ARHP Annual Scientific Meeting
Philadelphia October 16-21, 2009.
Titration of CD45 Expression Reveals Differential Regulation of Antigen Receptor Signaling in T and B Cells and Defines a BCR Signaling Threshold for B Cell Negative Selection
Zikherman1, Julie, Doan1, Kristin, Jenne1, Craig, Watson1, Susan, Goodnow2, Christopher, Raschke3, William, Weiss1, Arthur
Negative selection during lymphocyte development is required to establish a self-tolerant antigen receptor (AgR) repertoire. Here we take advantage of a novel allelic series of CD45 expression in mice which permits precise titration of BCR signal intensity and reveals a BCR signaling threshold for negative selection during splenic B cell development.
The receptor-like tyrosine phosphatase CD45 positively regulates AgR signaling by dephosphorylating the inhibitory tyrosine of the src family kinases (SFKs). It has been previously shown that CD45 is required for normal T and B cell development. Paradoxically, very low levels of CD45 are sufficient to fully rescue T cell but not B cell development. We show here that a negative regulatory role for CD45 that is unique to T cells accounts for this phenomenon.
We have identified a novel allele of CD45, lightning, in which a single residue mutation produces low protein expression levels but normal isoform splicing. We additionally used previously described 'HE' mice that have supraphysiologic levels of CD45 expression. By crossing these two alleles to either wild type or null CD45 alleles, we have generated a series of mice in which CD45 expression levels are titrated across a broad range (0180% of wildtype).
Results and Conclusion:
Supraphysiologic doses of CD45 enhance Erk phosphorylation and calcium flux following BCR stimulation, while the opposite is true for T cells. Conversely, low CD45 expression enhances TCR signaling but impairs BCR signaling. We conclude that CD45 plays a negative regulatory role in TCR but not BCR signaling.
Increasing CD45 expression in both T and B cells results in progressive dephosphorylation of the inhibitory tyrosine of the SFKs. However, only in T cells is the activation loop tyrosine of SFKs dephosphorylated with increasing CD45 level. We propose that this provides a biochemical basis for differential regulation of T and B cell signaling by CD45 and, in turn, accounts for the differential capacity of low CD45 expression to rescue T and B cell development.
In mice with increasing CD45 expression and concurrently enhanced BCR signaling, we observe gradual reduction in the MZ B cell compartment, while B1 cell development is preserved. Although splenic T1 B cell numbers are similar across the allelic series, supraphysiologic CD45 expression produces sharp reductions in T2 and in mature (FO) B cell numbers relative to wild type. In contrast, B cell numbers are relatively preserved in CD45 deficient mice, with only a modest decline between T2 and FO B cell compartments. We conclude that the CD45 allelic series reveals a signaling threshold for negative selection at the splenic transitional stages of B cell development. This has important implications for the establishment of a self-tolerant B cell repertoire.
To cite this abstract, please use the following information:
Zikherman, Julie, Doan, Kristin, Jenne, Craig, Watson, Susan, Goodnow, Christopher, Raschke, William, et al; Titration of CD45 Expression Reveals Differential Regulation of Antigen Receptor Signaling in T and B Cells and Defines a BCR Signaling Threshold for B Cell Negative Selection [abstract]. Arthritis Rheum 2009;60 Suppl 10 :696