PI3 kinase function is vital for the function but not formation of LAT-mediated signaling complexes
ABSTRACT The induction of the T cell receptor (TCR) is necessary for the activation and function of human T cells. TCR activation results in the tyrosine phosphorylation of LAT, leading to the direct interaction with several proteins, including PLC-gamma 1, Grb2 and Gads. These direct ligands then mediate the indirect interaction of LAT with proteins, such as SLP-76, Vav1 and Itk. PLC-gamma 1, Vav1 and Itk contain pleckstrin homology (PH) domains that interact with the enzymatic product of phosphoinositide-3-kinase (PI3K), suggesting the function of PI3K may modulate LAT-mediated complexes. Therefore, we characterized the poorly understood role of PI3K activity in the formation and function of multiprotein signaling complexes that form at LAT. Inhibition of PI3K catalytic function had little effect on the phosphorylation of LAT, SLP-76, Vav1 or PLC-gamma 1 or on the ability of PLC-gamma 1 to interact with LAT or SLP-76. However, PI3K activity appeared to be required for the induction of downstream signaling events. These data indicate that the formation of LAT-mediated complexes do not appear to depend on PI3K activity, whereas the optimal downstream function of these complexes requires the catalytic function of PI3K.
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ABSTRACT: The Tec family tyrosine kinases regulate lymphocyte development, activation, and differentiation. In T cells, the predominant Tec kinase is Itk, which functions downstream of the T-cell receptor to regulate phospholipase C-gamma. This review highlights recent advances in our understanding of Itk kinase structure and enzymatic regulation, focusing on Itk protein domain interactions and mechanisms of substrate recognition. We also discuss the role of Itk in the development of conventional versus innate T-cell lineages, including both alphabeta and gammadelta T-cell subsets. Finally, we describe the complex role of Itk signaling in effector T-cell differentiation and the regulation of cytokine gene expression. Together, these data implicate Itk as an important modulator of T-cell signaling and function.Cold Spring Harbor perspectives in biology 07/2010; 2(7):a002287. DOI:10.1101/cshperspect.a002287 · 8.23 Impact Factor
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ABSTRACT: The adapter molecule LAT is a nucleating site for multiprotein signaling complexes that are vital for the function and differentiation of T cells. Extensive investigation of LAT in multiple experimental systems has led to an integrated understanding of the formation, composition, regulation, dynamic movement, and function of LAT-nucleated signaling complexes. This review discusses interactions of signaling molecules that bind directly or indirectly to LAT and the role of cooperativity in stabilizing LAT-nucleated signaling complexes. In addition, it focuses on how imaging studies visualize signaling assemblies as signaling clusters and demonstrate their dynamic nature and cellular fate. Finally, this review explores the function of LAT based on the interpretation of mouse models using various LAT mutants.Cold Spring Harbor perspectives in biology 08/2010; 2(8):a005512. DOI:10.1101/cshperspect.a005512 · 8.23 Impact Factor
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ABSTRACT: IL-2 inducible T-cell kinase (Itk) is a Tec family non-receptor tyrosine kinase involved in signaling downstream of the T-cell receptor. Itk contains an amino-terminal Pleckstrin Homology (PH) domain that binds phosphatidylinositol (3,4,5)-trisphosphate, recruiting Itk to the plasma membrane upon T-cell receptor activation. In addition to phosphoinositide binding, accumulating data suggest that the Itk PH domain likely mediates additional interactions outside of the phosphoinositide ligand binding pocket. The structural basis for additional PH domain functions remains elusive because of the poor recombinant expression and in vitro solution behavior of the Itk PH domain. Here, we determine that the lone α-helix in the Itk PH domain is responsible for the poor solution properties and that mutation of just two residues in the Itk α-helix to the corresponding amino acids in Btk or Tec dramatically improves the soluble recombinant expression and solution behavior of the Itk PH domain. We present this double mutant as a valuable tool to characterize the structure and function of the Itk PH domain. It is also interesting to note that the precise sites of mutation identified in this study appear as somatic mutations associated with cancerous tissue. Collectively, the findings suggest that the two helical residues in the Itk PH domain may serve an important and unique structural role in wild-type Itk that differentiates this tyrosine kinase from its related family members.Protein Science 09/2012; 21(9):1288-97. DOI:10.1002/pro.2114 · 2.86 Impact Factor