RESPONSES OF Nf1Fcr/Nf1FcrB CELLS AND Nf1+/Nf1+B CELLS TO SURFACE IMMUNOGLOBULIN CROSSLINKING

Abstract

The aim of this research is to compare the cellular responses of neurofibromin deficient (Nf1-/-) B cells and neurofibromin positive (Nf1+/+) B cells to surface immunoglobulin (sIg) crosslinking, which mimics B cell activation by specific antigens. Mice heterozygous for a germline null mutation at the Nf1 gene (Nf1Fcr/+) were mated, and fetal liver cells from 13.5 day Nf1Fcr/Nf1Fcr or Nf1+/Nf1+ embryos were used to reconstitute the hematopoietic system of lethally irradiated recipient mice. After full reconstitution had taken place, splenic B cells were isolated and surface immunoglobulin was crosslinked with goat a-murine Ig antibodies. The rate of surface Ig patching and capping after crosslinking was identical in B cells of Nf1Fcr/Nf1Fcr and Nf1+/Nf1+ genotypes. 3H-Thymidine uptake, after crosslinking with α-IgG or α-IgM or with the addition of IL-5, was measured and found to be the same in B cells of both the Nf1Fcr/Nf1Fcr and Nf1+/Nf1+ genotypes. Furthermore, both Nf1 mutant and wild-type B cells exhibited the ability for Ras to cocap with slg. However, differences in the appearance of proteins phosphorylated on tyrosine after crosslinking of surface Ig were noted. When lysates from crosslinked B cells were probed with α-phosphotyrosine antibody a band of approximately 109 kd, present in wild-type lysate, was not present in the mutant lysate. There was also a difference between mutant and wildtype B cell lysates in the level of phosphorylation on tyrosine for other substrates. These results show that neurofibromin may not have a role in the initial activation of the signaling pathway in B cells, but it may play a role further downstream in the signaling pathway. While defects in B cells have not been noted in NF1 patients, who carry one mutant NF1 allele, these results may lead to new insights in neurofibromin function or help diagnose NF1 syndrome in some patients.