Binding Specificity of Multiprotein Signaling Complexes Is Determined by Both Cooperative Interactions and Affinity Preferences

Houtman, Jon C. D.; Higashimoto, Yuichiro; Dimasi, Nazzareno; Cho, Sangwoo; Yamaguchi, Hiroshi; Bowden, Brent; Regan, Carole K.; Malchiodi, Emilio L.; Mariuzza, Roy A.; Schuck, Peter; Appella, Ettore; Samelson, Lawrence E.

doi:10.1021/bi0357311

Cited by 106 publications

(162 citation statements)

References 32 publications

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“…However, the proteins had a 50 -100-fold weaker affinity for pY132, which explained the lack of binding to that site. The PLC-g1 SH2 domain bound the pY132 motif with stronger affinity than the pY171, pY191, or pY226 motif; however, it was thought that the affinity difference alone would not account for its selectivity (Houtman et al 2004). Overall, these studies indicated that the binding affinities alone were not sufficient to drive the apparent specificity of LAT motifs for PLC-g1 and Gads.…”

Section: Lat Binding Preferences Are Not Solely Driven By Affinitiesmentioning

confidence: 96%

“…Interaction with LAT was shown to be required for both PLC-g1 activation and localization near its substrate PIP 2 at the plasma membrane (Zhang et al 1999a;Zhang et al 2000;Lin and Weiss 2001). Subsequently, the amino-terminal SH2 domain of PLC-g1 was shown to bind phosphorylated LAT Y132 with high affinity (Paz et al 2001;Houtman et al 2004). Furthermore, a single Y132F mutation abolished the association between LAT and PLC-g1 in Jurkat T cells (Zhang et al 2000).…”

Section: Plc-g1 Binds Y132 Of Lat and Mediates Transcriptional Activamentioning

confidence: 99%

“…The three phosphotyrosines of SLP-76 bind PI3K, Vav, Itk, and Nck. The central prolinerich region binds to Gads with high affinity and to PLC-g1 with low affinity Houtman et al 2004). However, SLP-76 only associates weakly with LAT in the absence of the LAT residue Y132, suggesting that SLP-76 binding to LAT is stabilized by the presence of PLC-g1 (Zhang et al 2000).…”

Section: Molecules Recruited To Lat Via Slp-76 Promote T-cell Activationmentioning

confidence: 99%

“…For example, the PLC-g1 PH domain would position PLC-g1 near the membrane, likely favoring interactions with pY132, as it is the most membrane proximal of the four motifs (Houtman et al 2004). However, SLP-76 may also contribute to the stabilization of PLC-g1 binding to LAT.…”

Section: Lat Binding Preferences Are Not Solely Driven By Affinitiesmentioning

confidence: 99%

“…They showed that the high affinity interaction between Gads and SLP-76 stabilized the PLC-g1-binding conformation of SLP-76. Therefore, positive cooperativity mediated by the Gads/SLP-76 pair allowed PLC-g1 to bind SLP-76 (Houtman et al 2004). Interestingly, LAT residue Y191 was phosphorylated substantially earlier than Y132 in T cells.…”

Section: Lat Binding Preferences Are Not Solely Driven By Affinitiesmentioning

confidence: 99%

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The LAT Story: A Tale of Cooperativity, Coordination, and Choreography

Balagopalan¹,

Coussens²,

Sherman³

et al. 2010

Cold Spring Harbor Perspectives in Biology

153

184

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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.

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Section: Lat Binding Preferences Are Not Solely Driven By Affinitiesmentioning

confidence: 96%

Section: Plc-g1 Binds Y132 Of Lat and Mediates Transcriptional Activamentioning

confidence: 99%

Section: Molecules Recruited To Lat Via Slp-76 Promote T-cell Activationmentioning

confidence: 99%

Section: Lat Binding Preferences Are Not Solely Driven By Affinitiesmentioning

confidence: 99%

Section: Lat Binding Preferences Are Not Solely Driven By Affinitiesmentioning

confidence: 99%

See 3 more Smart Citations

The LAT Story: A Tale of Cooperativity, Coordination, and Choreography

Balagopalan¹,

Coussens²,

Sherman³

et al. 2010

Cold Spring Harbor Perspectives in Biology

153

184

View full text Add to dashboard Cite

show abstract

Characterizing Protein‐Protein Interactions by Sedimentation Velocity Analytical Ultracentrifugation

2008

Self Cite

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This unit introduces the basic principles and practice of sedimentation velocity analytical ultracentrifugation for the study of reversible protein interactions, such as the characterization of self-association, heterogeneous association, multi-protein complexes, binding stoichiometry, and the determination of association constants. The analytical tools described include sedimentation coefficient and molar mass distributions, multi-signal sedimentation coefficient distributions, Gilbert-Jenkins theory, different forms of isotherms, and global Lamm equation modeling. Concepts for the experimental design are discussed, and a detailed step-by-step protocol guiding the reader through the experiment and the data analysis is available as an Internet resource.

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Competition between SLP76 and LAT for PLCγ1 binding in resting T cells

et al. 2010

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The constitutive interaction between the P1 domain (a 67-amino-acid functional domain within the proline-rich region) of SLP76 and the SH3 domain of phospholipase Cc1 (PLCc1) has been shown. To determine the significance of the interaction between SLP76 and PLCc1 in resting T cells, we examined molecules associated with PLCc1 in the absence of both SLP76 and, more specifically, the P1 domain of SLP76. Using a mutant Jurkat T-cell line, we showed that PLCc1 associated with LAT when the constitutive association with SLP76 was blocked. We also found that the PLCc1 association with LAT occurred in the membranes of resting T cells. Further experiments demonstrated that LAT competed with SLP76 for PLCc1 binding and that the LAT interaction with PLCc1 was mediated by the SH3 domain of PLCc1. Collectively, these results suggest that the constitutive association of SLP76 with PLCc1 is required to prevent the association with LAT as well as the premature recruitment of PLCc1 to the cell membrane.Key words: LAT . Phospholipase Cc1 . SSLP76 . Signalling . T cell IntroductionEngagement of the TCR triggers a complex cascade of signaling events leading to T-cell activation. A key initial event in the T-cell signaling pathway is tyrosine phosphorylation and activation of PLCg1 (phospholipase Cg1) [1,2]. TCR signaling pathways activate Src-, Syk-, and Tek-family protein tyrosine kinases that induce phosphorylation of many proteins including PLCg1 [3,4]. The adaptor proteins LAT linker for activation of T cells, SLP76 (Src homology 2-domain-containing leukocyte protein of 76 kDa), and Gads (Grb2-related adaptor downstream of Shc) appear to constitute an integrated signaling unit that couples TCR-mediated activation of cytoplasmic protein tyrosine kinases to PLCg1 enzyme activation [5,6].LAT is a transmembrane adaptor protein containing two palmitoylated cysteine residues proximal to the transmembrane domain that is constitutively localized to specialized membrane microdomains known as glycosphingolipid-enriched membrane domains (GEM), lipid rafts, or detergent-insoluble membrane domains [7,8]. LAT is heavily tyrosine-phosphorylated upon TCR stimulation, creating binding motifs for proteins containing Src homology (SH) 2 domains [8,9]. The N-terminal SH2 domain of PLCg1 binds to LAT phospho-tyrosine 132 (pY 132 ) and this interaction recruits PLCg1 to the GEM, leading to TCR-induced tyrosine phosphorylation and enzyme activation [10][11][12]. However, LAT is unable to activate PLCg1 in the absence of SLP76, which is indirectly recruited to LAT via Gads [13,14].SLP76 consists of three domains that mediate interactions with many proteins, leading to activation of TCR-mediated signaling pathways. An N-terminal acidic domain containing three tyrosine phosphorylation sites and a central proline-rich region containing the Gads-binding sites are indispensable for PLCg1 activation [15]. A functionally important 67 amino acid segment within the proline-rich region of SLP76, denoted the P1 region, binds to the SH3 domain of PLCg1 in re...

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Binding Specificity of Multiprotein Signaling Complexes Is Determined by Both Cooperative Interactions and Affinity Preferences

Cited by 106 publications

References 32 publications

The LAT Story: A Tale of Cooperativity, Coordination, and Choreography

The LAT Story: A Tale of Cooperativity, Coordination, and Choreography

Characterizing Protein‐Protein Interactions by Sedimentation Velocity Analytical Ultracentrifugation

Competition between SLP76 and LAT for PLCγ1 binding in resting T cells

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