Type I interferon signaling acts directly on transitional T2 B cells to promote RNP-Ag tolerance loss in BXD2 mice

Abstract

Abstract In lupus, autoreactive B cells fail to be removed at the transitional B cell stage. Lupus prone BXD2 mice exhibit transitional B cell tolerance loss and production of autoAb against RNA binding proteins (RNP). The purpose of this study was to determine direct effects of IFNα on developing transitional B cells. Analysis of IFNαR expression in B6 and BXD2 mice revealed the highest expression levels on transitional B cells with lower levels on mature FO, MZ, and GC B cells. Interestingly, both in vitro and in vivo, peripheral B cell responsiveness to IFNα started at the IgMhiCD23lo T1 stage and was significantly upregulated at the T2 stage, as measured by CD69 induction (76±5% in B6 vs 89±3% in BXD2 T2). In vivo treatment of B6 and BXD2 mice with IFNα promoted B cell upregulation of CD69 and CD86 on T2 B cells. Using an antigen (Ag) tetramer approach for isolation and characterization of La and RNP (La13-27 and snRNP357-373) autoreactive B cells, we found that IFNα stimulation increased maturation of transitional RNP autoreactive B cells and expansion of the autoantigen engaged IgMlo T3 compartment in BXD2 mice. Consistent with abnormal signaling and activation in vivo, transitional B cells from spleens of BXD2 mice exhibited elevated basal phospho-tyrosine and BCR induced activation, beginning at the T2 stage. Single cell analysis of transitional T2 B cells from BXD2 mice revealed a heterogeneity of gene expression within this population and a negative correlation between the expression of IFNαR gene and Id3, an important transcription factor for B-cell anergy. Our results suggest that B-cell tolerance loss in BXD2 mice occurs at the transitional stage, and IFNα exhibits the potential to break tolerance to RNP in transitional B cells.