Sustained Activation of Lyn Tyrosine Kinase In Vivo Leads to Autoimmunity

Abstract

Genetic ablation of the Lyn tyrosine kinase has revealed unique inhibitory roles in B lymphocyte signaling. We now report the consequences of sustained activation of Lyn in vivo using a targeted gain-of-function mutation (Lynup/up mice). Lynup/up mice have reduced numbers of conventional B lymphocytes, down-regulated surface immunoglobulin M and costimulatory molecules, and elevated numbers of B1a B cells. Lynup/up B cells are characterized by the constitutive phosphorylation of negative regulators of B cell antigen receptor (BCR) signaling including CD22, SHP-1, and SHIP-1, and display attributes of lymphocytes rendered tolerant by constitutive engagement of the antigen receptor. However, exaggerated positive signaling is also apparent as evidenced by the constitutive phosphorylation of Syk and phospholipase Cγ2 in resting Lynup/up B cells. Similarly, Lynup/up B cells show a heightened calcium flux in response to BCR stimulation. Surprisingly, Lynup/up mice develop circulating autoreactive antibodies and lethal autoimmune glomerulonephritis, suggesting that enhanced positive signaling eventually overrides constitutive negative signaling. These studies highlight the difficulty in maintaining tolerance in the face of chronic stimulation and emphasize the pivotal role of Lyn in B cell signaling.

Constitutive activation of Bruton's tyrosine kinase induces the formation of autoreactive IgM plasma cells

“…Immature B cells were arrested at the progression from IgM low into IgM high cells, reflecting the first immune tolerance checkpoint at which autoreactive B cells become susceptible to apoptosis and the peripheral mature B-cell pool was reduced to o1% of its normal size. This phenotype is in marked contrast with that of other mouse models with increased BCR signaling [12][13][14][15][16][17][18][19], which are mainly characterized by B-cell hyperresponsiveness, enhanced B-1 cell differentiation and autoimmunity. In our Tg mice the expression levels of mutated E41K-Btk were in the same range as the endogenous, unmutated Btk.…”

“…Conversely, a complex B-cell phenotype characterized by reduced numbers of follicular B cells, elevated numbers of B-1 B cells and to some extent MZ B cells, B-cell hyper-responsiveness and auto-antibody formation is found in genetic changes that increase BCR signaling. These include gain-of-function mutants of Lyn or Plcg2, deficiency for PTEN, a lipid phosphatase that antagonizes PI3K activity, overexpression of CD19 or mutations that disable inhibitory signaling of membrane receptors such as CD22, Pir-B, Siglec-G and FcgRIIB or their downstream signaling molecules Shp1 or Ship [12][13][14][15][16][17][18][19][20][21].Btk is a member of the Tec protein tyrosine kinase family that mediates many aspects of B-cell development, survival and function [8,22]. Whereas in humans Btk mutations cause a severe arrest of B-cell development at the pre-B-cell stage leading to X-linked agammaglobulinemia, in the mouse there is only a mild pre-B-cell defect, differentiation of transitional into mature peripheral B cells is impaired and B-1 cells are lacking [23][24][25].…”

“…2C) and micro-architecture of the spleen (data not shown). Moreover, we have previously found that Y223 phosphorylation is not essential for Btk function in vivo: Y223F-Btk can fully correct the features of the Btk-deficient phenotype, including pre-B-cell B-1 cell development, serum IgM levels and TI-II responses [27].The complex phenotype of mice with constitutively activated Btk largely resembles that of other Tg or knock-out mice with increased BCR signaling [12][13][14][15][16][17][18][19]. These mice also contain fewer follicular B cells, increased numbers of B-1 B cells, together with B-cell hyperresponsiveness and autoimmunity.…”

“…Conversely, a complex B-cell phenotype characterized by reduced numbers of follicular B cells, elevated numbers of B-1 B cells and to some extent MZ B cells, B-cell hyper-responsiveness and auto-antibody formation is found in genetic changes that increase BCR signaling. These include gain-of-function mutants of Lyn or Plcg2, deficiency for PTEN, a lipid phosphatase that antagonizes PI3K activity, overexpression of CD19 or mutations that disable inhibitory signaling of membrane receptors such as CD22, Pir-B, Siglec-G and FcgRIIB or their downstream signaling molecules Shp1 or Ship [12][13][14][15][16][17][18][19][20][21].…”

Engagement of the B-cell antigen receptor activates STAT through Lyn in a Jak-independent pathway

“…However, the phosphorylation of STATs mediated by Lyn through BCR is transient. We ascribe this to Lyn's dual role in both positive and inhibitory regulation of BCR signaling (Hibbs et al, 2002) that results in balance between PTKs and protein tyrosine phosphatases (Ishikawa et al, 2002;Pereira and Lowell, 2003), so the functional consequences of Lyn-mediated STATs activation remain undetermined and needs further study. Transient activation by Lyn may result in a different set of genes transcribed than more sustained activation by the JAKs.…”