Src-family kinases: rheostats of immune cell signaling
Introduction
One of the premier, and original, investigators in the field of Src-family kinases, Dr. Joan Brugge, once made a comment at a meeting she was chairing that there were so many people now working on Src kinases that there was probably one group for each codon of the gene. She made that comment in 1994 at the Oncogene Meeting in Fredrick, Maryland. In a decade since then, the field has grown dramatically. Both in immune cells and non-hematopoietic cells Src-family kinases have been implicated in a multitude intracellular signaling pathways—everything from responses to UV light to regulation of β-adrenergic signaling to sensitivity to ethanol consumption (Cowen et al., 2003, Kabuyama et al., 2002, Ma and Huang, 2002). In blood cells and platelets, the list is no less extensive. Src-family kinases have been implicated in innate immune signaling, responses to cytokines, growth factors, regulators of apoptosis, regulators of antigen signaling, stimulation of cells by immune complexes, phagocytosis, responses to adhesive stimulation—integrin signaling, controllers of K/Cl channels, and key players in G-protein coupled (GPCR) signaling pathways (as examples see Chan et al., 1998, De Franceschi et al., 1997, Fitzer-Attas et al., 2000, Gardai et al., 2002, Lowell and Berton, 1999, Lowell et al., 1996, Nijhuis et al., 2002, Rane and Reddy, 2002). In most of these cases, the loss of Src-family tyrosine kinase activity, either by pharmacologic inhibition, use of dominant-negative inhibitors or more recently using genetically modified cells or mice, results in diminution of the signaling pathway being studied. More convincingly, the use of knockout mouse models to study the overall physiologic outcome of the loss of Src-family kinase-dependent signaling has pointed the field in the direction of the most significant functions for these kinases. In this regard, the requirement for Src-family kinase activity for T- and B-lymphocyte development (Saijo et al., 2003, van Oers et al., 1996) or for osteoclast function, as demonstrated in the original Src mutant made by Soriano et al. (1991), is absolutely clear and correlates with a large number of cell-based experiments. Nevertheless, it is important to realize that the multitude of signaling reactions these kinases have been implicated in using cell based assays or other approaches does not seem to manifest as severe physiologic phenotypes in the various knockout models; an issue that makes interpretation of both types of data complicated. Obviously, there are many explanations for this difference (compensation between kinases or even between different signaling pathways), but the idea that Src-family kinase activity is critical and central to so many different intracellular signaling pathways may be overstating the argument. The concept that these kinases may be contributory to multiple pathways may be a better reference frame—hence the title of this review that Src-family kinases could be viewed as rheostats of intracellular signaling and not as on-off switches.
Even more amazing is the growing realization that these kinases also play critical roles in downmodulating or inhibiting signaling in immune cells; a concept that was only revealed through study of the mutant models. This inhibitory activity of the kinases was first realized in studies attempting to explain the autoimmune phenotype of lyn−/− mice (DeFranco et al., 1998, Harder et al., 2001, Hibbs et al., 1995), which will be discussed below, but has now been observed with other Src-family kinases in other immune cell types (Gresham et al., 2000, Vines et al., 2001). Indeed, the inhibitory function of these kinases may be one of the more active and surprising areas of investigation. The dual specificity of various members of this kinase family contributes to the general idea that they are overall modulators of signaling, like rheostats, and not absolutely critical, at least as single enzymes alone, in any one pathway.
To address this general theme that Src-family kinases are more like rheostats and not switches, this mini-review will focus on several major pathways in immune cells for which there is clear evidence that Src-family kinases play a significant regulatory role. These pathways are:
- 1.
The immunoreceptor tyrosine-based activation motif (ITAM)-dependent pathways, used by all immunoreceptors (lymphocyte antigen receptors, Fc receptors, NK activating receptors).
- 2.
The immunoreceptor tyrosine-based inhibitory motif (ITIM)-dependent pathways, which negatively regulate both immunoreceptor function as well as a host of other signaling responses, such as cytokines and integrin signaling.
- 3.
The integrin pathways, which regulate leukocyte activation by adhesion.
Section snippets
Structure and expression of immune cell Src-family kinases
The members of the Src-family kinases consist of Src, Fyn, Yes, Lck, Hck, Fgr, Lyn, Blk, and Yrk (for an excellent prior review of Src-family kinases in leukocytes see Korade-Mirnics and Corey, 2000). Each of these proteins are about 60 kD in molecular weight and have a common structure consisting of an N-terminal unique domain, followed by SH3, SH2 and tyrosine kinases domains (see Fig. 1). Several of the Src-family proteins are expressed as two proteins due to alternative splicing patterns or
ITAM-dependent signaling pathways
Immunoreceptors such as the T- and B-lymphocyte antigen receptors, Fcγ and Fcε receptors, NK activating receptors and the platelet receptor GPVI, all depend on Src-family kinases for appropriate signaling. The vast majority of immunoreceptors are physically associated with ITAM-containing subunit proteins, each of which have paired YXXL motifs (Fig. 2). Examples of these ITAM subunits are the TCRζ, CD3γ/δ/ε chains in T-cells, the Igα/β chains in the B-cell receptor, the FcRγ chain in Fc
ITIM-mediated inhibitory signaling
The realization that Src-family kinases play an important physiologic role in downmodulating or inhibiting signaling responses first came to light in studies examining lyn−/− mutant mice. The initial phenotype described for lyn−/− animals was quite surprising: the mice had hyperresponsive B-cells, increased Ig levels and autoimmunity (immune complex nephritis) (reviewed in DeFranco et al., 1998). The mechanism for this surprising result was demonstrated by examination of cell surface receptors
Src-family kinases in leukocyte integrin signaling pathways
There is a large body of evidence implicating Src-family tyrosine kinases in integrin signaling pathways, both in blood cells and non-hematopoietic cells. Integrins are major cellular receptors that mediate cell–cell and cell–substratum interactions. These receptors signal in two fashions, which are commonly referred to as “inside-out” versus “outside-in” (integrin signaling pathways in T-lymphocytes have recently been reviewed in Hogg et al., 2003, Shimizu, 2003). The “inside-out” pathway
Conclusions
This review has focused on only a limited number of the signaling pathways in which Src-family kinases have been implicated. In immune cells, this list could be extended to include responses to chemokines/chemoattractants (Barlic et al., 2000, Mocsai et al., 2000), cytokines (Rane and Reddy, 2002), innate immune stimuli, such as LPS (Yazawa et al., 2003) as well as the myriad of non-immune cell specific stimuli, such as responses to UV irradiation, heat, osmotic shock and others. In many of
References (105)
- et al.
Differential effects of filipin and methyl-beta-cyclodextrin on B cell receptor signaling
Biochem. Biophys. Res. Commun.
(2001) - et al.
Positive and negative roles of the tyrosine kinase Lyn in B cell function
Semin. Immunol.
(1998) - et al.
Cbl-b, a RING-type E3 ubiquitin ligase, targets phosphatidylinositol 3-kinase for ubiquitination in T cells
J. Biol. Chem.
(2001) - et al.
The Src kinase p56lck up-regulates VLA-4 integrin affinity. Implications for rapid spontaneous and chemokine-triggered T cell adhesion to VCAM-1 and fibronectin
J. Biol. Chem.
(2001) - et al.
Activation of SHIP by NADPH oxidase-stimulated Lyn leads to enhanced apoptosis in neutrophils
J. Biol. Chem.
(2002) - et al.
Protein-tyrosine phosphatase alpha regulates Src family kinases and alters cell-substratum adhesion
J. Biol. Chem.
(1998) - et al.
Gain- and loss-of-function Lyn mutant mice define a critical inhibitory role for Lyn in the myeloid lineage
Immunity
(2001) - et al.
Multiple defects in the immune system of Lyn-deficient mice, culminating in autoimmune disease
Cell
(1995) - et al.
The role of lipid rafts in T cell antigen receptor (TCR) signalling
Semin. Immunol.
(2000) - et al.
Differential intrinsic enzymatic activity of Syk and Zap-70 protein-tyrosine kinases
J. Biol. Chem.
(1996)