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Sustained activation of Lyn tyrosine kinase in vivo leads to autoimmunity.

Hibbs ML, Harder KW, Armes J, Kountouri N, Quilici C, Casagranda F, Dunn AR, Tarlinton DM - J. Exp. Med. (2002)

Bottom Line: However, exaggerated positive signaling is also apparent as evidenced by the constitutive phosphorylation of Syk and phospholipase Cgamma2 in resting Lyn(up/up) B cells.Similarly, Lyn(up/up) B cells show a heightened calcium flux in response to BCR stimulation.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.

View Article: PubMed Central - PubMed

Affiliation: Ludwig Institute for Cancer Research, Melbourne Tumour Biology Branch. Walter and Eliza Hall Institute of Medical Research, Royal Melbourne Hospital, Victoria 3050, Australia.

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 (Lyn(up/up) mice). Lyn(up/up) mice have reduced numbers of conventional B lymphocytes, down-regulated surface immunoglobulin M and costimulatory molecules, and elevated numbers of B1a B cells. Lyn(up/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 Cgamma2 in resting Lyn(up/up) B cells. Similarly, Lyn(up/up) B cells show a heightened calcium flux in response to BCR stimulation. Surprisingly, Lyn(up/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.

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Cellularity and composition of lymphoid tissue in 8-wk-old Lyn+/+, Lyn+/up, and Lynup/up mice. (a) Nucleated cells in lymphoid tissue in Lyn+/+, Lyn+/up, and Lynup/up mice. Numbers represent per ml blood, per whole spleen, per mesenteric lymph nodes, and per femur. (b) Numbers of conventional B cells, B1 cells, T cells, granulocytes (Gr), and monocytes (Mo) in blood and (c) spleen of Lyn+/+, Lyn+/up, and Lynup/up mice. (d) Numbers of B and T cells in mesenteric lymph nodes of Lyn+/+, Lyn+/up, and Lynup/up mice. (e) Numbers of recirculating B cells (Rec B), immature B cells (Imm B), and Pre-B cells in the BM of Lyn+/+, Lyn+/up, and Lynup/up mice. Data in a–e is compiled from nine mice in three experiments, except B cell and B1 cell numbers in blood and spleen, which were compiled from six mice in two experiments. Data is mean ± standard error. (f) BM B cell progenitors in Lyn+/+ and Lynup/up mice. Numbers are the mean ± standard deviation for five mice of each genotype. For a–f, white bars, Lyn+/+; gray bars, Lyn+/up; black bars, Lynup/up.
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fig2: Cellularity and composition of lymphoid tissue in 8-wk-old Lyn+/+, Lyn+/up, and Lynup/up mice. (a) Nucleated cells in lymphoid tissue in Lyn+/+, Lyn+/up, and Lynup/up mice. Numbers represent per ml blood, per whole spleen, per mesenteric lymph nodes, and per femur. (b) Numbers of conventional B cells, B1 cells, T cells, granulocytes (Gr), and monocytes (Mo) in blood and (c) spleen of Lyn+/+, Lyn+/up, and Lynup/up mice. (d) Numbers of B and T cells in mesenteric lymph nodes of Lyn+/+, Lyn+/up, and Lynup/up mice. (e) Numbers of recirculating B cells (Rec B), immature B cells (Imm B), and Pre-B cells in the BM of Lyn+/+, Lyn+/up, and Lynup/up mice. Data in a–e is compiled from nine mice in three experiments, except B cell and B1 cell numbers in blood and spleen, which were compiled from six mice in two experiments. Data is mean ± standard error. (f) BM B cell progenitors in Lyn+/+ and Lynup/up mice. Numbers are the mean ± standard deviation for five mice of each genotype. For a–f, white bars, Lyn+/+; gray bars, Lyn+/up; black bars, Lynup/up.

Mentions: To determine the effect of the Lyn gain-of-function mutation on the B cell compartment, we performed analyses of primary and secondary lymphoid tissue of 8-wk-old Lyn+/+, Lyn+/up, and Lynup/up mice (Figs. 2–4) . Spleen cellularity was reduced in both Lyn+/up and Lynup/up mice whereas white blood cell counts were elevated in Lynup/up mice (Fig. 2 a), presumably due to an increase in neutrophils (27). Lynup/up mice showed an overall reduction in B cells: twofold in peripheral blood, fivefold in spleen, 10-fold in lymph node, and 20-fold in the recirculating B cell compartment of the BM (Fig. 2, b–e, and Fig. 3 a). The B cell deficiency in heterozygous Lyn+/up mice was less severe than in homozygous mice, being approximately twofold in all lymphoid tissues. Numbers of pro-B, pre-B, and immature B cells in the BM of Lynup/up mice were not significantly different from controls (Fig. 2 e and Fig. 3 a) and B cell colony assays demonstrated no significant change in the numbers of B cell progenitors (Fig. 2 f). Although the BM results suggest that B cell development is not blocked in Lynup/up mice, levels of IgM on the surface of immature Lynup/up B cells are reduced by ∼35% (not depicted). Peripheral B cells from Lynup/up and Lyn+/up mice also express lower surface IgM (Fig. 3, a and b), Igβ, CD19, CD21, and CD22 (Fig. 3 b), whereas IgD (Fig. 3 b) and MHC class II (not depicted) are unaffected.


Sustained activation of Lyn tyrosine kinase in vivo leads to autoimmunity.

Hibbs ML, Harder KW, Armes J, Kountouri N, Quilici C, Casagranda F, Dunn AR, Tarlinton DM - J. Exp. Med. (2002)

Cellularity and composition of lymphoid tissue in 8-wk-old Lyn+/+, Lyn+/up, and Lynup/up mice. (a) Nucleated cells in lymphoid tissue in Lyn+/+, Lyn+/up, and Lynup/up mice. Numbers represent per ml blood, per whole spleen, per mesenteric lymph nodes, and per femur. (b) Numbers of conventional B cells, B1 cells, T cells, granulocytes (Gr), and monocytes (Mo) in blood and (c) spleen of Lyn+/+, Lyn+/up, and Lynup/up mice. (d) Numbers of B and T cells in mesenteric lymph nodes of Lyn+/+, Lyn+/up, and Lynup/up mice. (e) Numbers of recirculating B cells (Rec B), immature B cells (Imm B), and Pre-B cells in the BM of Lyn+/+, Lyn+/up, and Lynup/up mice. Data in a–e is compiled from nine mice in three experiments, except B cell and B1 cell numbers in blood and spleen, which were compiled from six mice in two experiments. Data is mean ± standard error. (f) BM B cell progenitors in Lyn+/+ and Lynup/up mice. Numbers are the mean ± standard deviation for five mice of each genotype. For a–f, white bars, Lyn+/+; gray bars, Lyn+/up; black bars, Lynup/up.
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fig2: Cellularity and composition of lymphoid tissue in 8-wk-old Lyn+/+, Lyn+/up, and Lynup/up mice. (a) Nucleated cells in lymphoid tissue in Lyn+/+, Lyn+/up, and Lynup/up mice. Numbers represent per ml blood, per whole spleen, per mesenteric lymph nodes, and per femur. (b) Numbers of conventional B cells, B1 cells, T cells, granulocytes (Gr), and monocytes (Mo) in blood and (c) spleen of Lyn+/+, Lyn+/up, and Lynup/up mice. (d) Numbers of B and T cells in mesenteric lymph nodes of Lyn+/+, Lyn+/up, and Lynup/up mice. (e) Numbers of recirculating B cells (Rec B), immature B cells (Imm B), and Pre-B cells in the BM of Lyn+/+, Lyn+/up, and Lynup/up mice. Data in a–e is compiled from nine mice in three experiments, except B cell and B1 cell numbers in blood and spleen, which were compiled from six mice in two experiments. Data is mean ± standard error. (f) BM B cell progenitors in Lyn+/+ and Lynup/up mice. Numbers are the mean ± standard deviation for five mice of each genotype. For a–f, white bars, Lyn+/+; gray bars, Lyn+/up; black bars, Lynup/up.
Mentions: To determine the effect of the Lyn gain-of-function mutation on the B cell compartment, we performed analyses of primary and secondary lymphoid tissue of 8-wk-old Lyn+/+, Lyn+/up, and Lynup/up mice (Figs. 2–4) . Spleen cellularity was reduced in both Lyn+/up and Lynup/up mice whereas white blood cell counts were elevated in Lynup/up mice (Fig. 2 a), presumably due to an increase in neutrophils (27). Lynup/up mice showed an overall reduction in B cells: twofold in peripheral blood, fivefold in spleen, 10-fold in lymph node, and 20-fold in the recirculating B cell compartment of the BM (Fig. 2, b–e, and Fig. 3 a). The B cell deficiency in heterozygous Lyn+/up mice was less severe than in homozygous mice, being approximately twofold in all lymphoid tissues. Numbers of pro-B, pre-B, and immature B cells in the BM of Lynup/up mice were not significantly different from controls (Fig. 2 e and Fig. 3 a) and B cell colony assays demonstrated no significant change in the numbers of B cell progenitors (Fig. 2 f). Although the BM results suggest that B cell development is not blocked in Lynup/up mice, levels of IgM on the surface of immature Lynup/up B cells are reduced by ∼35% (not depicted). Peripheral B cells from Lynup/up and Lyn+/up mice also express lower surface IgM (Fig. 3, a and b), Igβ, CD19, CD21, and CD22 (Fig. 3 b), whereas IgD (Fig. 3 b) and MHC class II (not depicted) are unaffected.

Bottom Line: However, exaggerated positive signaling is also apparent as evidenced by the constitutive phosphorylation of Syk and phospholipase Cgamma2 in resting Lyn(up/up) B cells.Similarly, Lyn(up/up) B cells show a heightened calcium flux in response to BCR stimulation.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.

View Article: PubMed Central - PubMed

Affiliation: Ludwig Institute for Cancer Research, Melbourne Tumour Biology Branch. Walter and Eliza Hall Institute of Medical Research, Royal Melbourne Hospital, Victoria 3050, Australia.

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 (Lyn(up/up) mice). Lyn(up/up) mice have reduced numbers of conventional B lymphocytes, down-regulated surface immunoglobulin M and costimulatory molecules, and elevated numbers of B1a B cells. Lyn(up/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 Cgamma2 in resting Lyn(up/up) B cells. Similarly, Lyn(up/up) B cells show a heightened calcium flux in response to BCR stimulation. Surprisingly, Lyn(up/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.

Show MeSH
Related in: MedlinePlus