Defective lymphocyte chemotaxis in beta-arrestin2- and GRK6-deficient mice.

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2002-05-28

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Abstract

Lymphocyte chemotaxis is a complex process by which cells move within tissues and across barriers such as vascular endothelium and is usually stimulated by chemokines such as stromal cell-derived factor-1 (CXCL12) acting via G protein-coupled receptors. Because members of this receptor family are regulated ("desensitized") by G protein-coupled receptor kinase (GRK)-mediated receptor phosphorylation and beta-arrestin binding, we examined signaling and chemotactic responses in splenocytes derived from knockout mice deficient in various beta-arrestins and GRKs, with the expectation that these responses might be enhanced. Knockouts of beta-arrestin2, GRK5, and GRK6 were examined because all three proteins are expressed at high levels in purified mouse CD3+ T and B220+ B splenocytes. CXCL12 stimulation of membrane GTPase activity was unaffected in splenocytes derived from GRK5-deficient mice but was increased in splenocytes from the beta-arrestin2- and GRK6-deficient animals. Surprisingly, however, both T and B cells from beta-arrestin2-deficient animals and T cells from GRK6-deficient animals were strikingly impaired in their ability to respond to CXCL12 both in transwell migration assays and in transendothelial migration assays. Chemotactic responses of lymphocytes from GRK5-deficient mice were unaffected. Thus, these results indicate that beta-arrestin2 and GRK6 actually play positive regulatory roles in mediating the chemotactic responses of T and B lymphocytes to CXCL12.

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10.1073/pnas.112198299

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Fong, Alan M, Richard T Premont, Ricardo M Richardson, Yen-Rei A Yu, Robert J Lefkowitz and Dhavalkumar D Patel (2002). Defective lymphocyte chemotaxis in beta-arrestin2- and GRK6-deficient mice. Proc Natl Acad Sci U S A, 99(11). pp. 7478–7483. 10.1073/pnas.112198299 Retrieved from https://hdl.handle.net/10161/7804.

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Scholars@Duke

Yu

Yen-Rei Andrea Yu

Adjunct Assistant Professor in the Department of Medicine

Dhavalkumar Dhirajlal Patel

Associate Consulting Professor in the Department of Medicine

The overall goals of the Patel laboratory are two-fold: 1) to define the mechanisms of inflammation, focusing on signaling through G protein coupled receptors, for the purpose of identifying novel therapeutic targets for immunologic diseases; and 2) to define the role that T cell education in the thymus plays in diseases of disordered immunity such as autoimmune diseases and primary immunodeficiency syndromes.

1. Roles of Membrane-Tethered Chemokines in Inflammation. Based on one of our primary hypotheses that the critical step that regulates leukocyte migration from the circulation into tissues is at the stage of firm adhesion of a rolling leukocyte to the vascular endothelium, and that membrane-bound but not soluble chemokines are important mediators of this critical event, we identified a new pathway by which leukocytes can migrate into sites of inflammation. Fractalkine (FKN, CX3CL1), a unique endothelial cell surface molecule with chemokine and mucin domains that is expressed on IL-1 or TNF activated vascular endothelium, mediates the rapid capture, firm adhesion, and activation of circulating monocytes, CD8+ T cells and NK cells under physiologic shear stresses. The co-receptor for FKN is CX3CR1, a G-protein coupled receptor (GPCR). These findings have identified that both chemokines and GPCR can function as cell adhesion molecules. We are studying the structure-activity relationships of FKN and CX3CR1 and also testing the physiologic role of the FKN pathway of leukocyte migration to sites of inflammation. Using animal models (CX3CR1 and FKN-deficient mice), we are determining the roles of FKN and CX3CR1 in monocyte and natural killer cell function. Using FKN as a model system, we are also studying the functional roles of tethered vs. soluble chemokines in leukocyte trafficking and in inflammation.

2. Role of Thymic Education in Diseases of Disordered Immunity. The thymus is essential for developing a normal immune system. Abnormal T cell education in thymus can result in the production of autoreactive T cells that lead to autoimmune diseases such as diabetes mellitus and multiple sclerosis. The lack of a thymus can result in immunodeficiency. We are studying the roles of the thymus in various human diseases.

Role of the Thymus in Immune Reconstitution. T cell reconstitution after hematopoietic cell transplantation can occur either by peripheral expansion of passively transferred, mature T cells or by T cell education in the thymus and production of naïve T cells. Whether naïve T cells develop or T cell reconstitution occurs via expansion of mature T cells may predict whether the recipient can respond well to infections or the recipient's T cell repertoire is limited. We have, in collaboration with Dr. Rebecca Buckley, been studying T cell reconstitution in severe combined immunodeficiency (SCID) patients who have received haploidentical T cell-depleted stem cell transplants to answer basic questions about the human immune system.

Role of the Thymus in Autoimmunity. Escape from central tolerance in the thymus can result in autoimmunity, and we are interested in defining whether T cell education plays a role in the development of autoimmunity. Thymic dendritic cells (DC) are primarily responsible for deletion of autoreactive T cells, and defective negative selection by thymic DCs could result in a propensity to develop autoimmune disease. We have begun to study the biology of human thymic DCs and their roles in autoimmune processes.


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