Combinatorial diversity of Syk recruitment driven by its multivalent engagement with FcεRIγ

Abstract

Syk/Zap70 family kinases are essential for signaling via multichain immune-recognition receptors such as tetrameric (αβγ2) FcεRI. Syk activation is generally attributed to cis binding of its tandem SH2 domains to dual phosphotyrosines within FcεRIγ-ITAMs (immunoreceptor tyrosine-based activation motifs). However, the mechanistic details of Syk docking on γ homodimers are unresolved. Here, we estimate that multivalent interactions for WT Syk improve cis-oriented binding by three orders of magnitude. We applied molecular dynamics (MD), hybrid MD/worm-like chain polymer modeling, and live cell imaging to evaluate relative binding and signaling output for all possible cis and trans Syk–FcεRIγ configurations. Syk binding is likely modulated during signaling by autophosphorylation on Y130 in interdomain A, since a Y130E phosphomimetic form of Syk is predicted to lead to reduced helicity of interdomain A and alter Syk’s bias for cis binding. Experiments in reconstituted γ-KO cells, whose γ subunits are linked by disulfide bonds, as well as in cells expressing monomeric ITAM or hemITAM γ-chimeras, support model predictions that short distances between γ ITAM pairs are required for trans docking. We propose that the full range of docking configurations improves signaling efficiency by expanding the combinatorial possibilities for Syk recruitment, particularly under conditions of incomplete ITAM phosphorylation.

Dephosphorylation accelerates the dissociation of ZAP70 from the T cell receptor

“…3 G ). This was within the error of the value determined above and similar to previous estimates for Syk ( 35 , 36 ). Given that k off is estimated with higher accuracy compared with K D in SPR, we proceeded to use the estimate of σ obtained by the kinetic data.…”

“…Thus, a highly kinetic binding mode can take place even if ZAP70 is relatively rigid. Although it is energetically less favorable, we note that ZAP70 may bind across phosphotyrosines from different ITAMs, as has been studied for Syk ( 35 , 51 ). This was not included in our model and would be unlikely in our SPR experiments because we used a low density of immobilized ITAMs.…”

Regulation of Trafficking and Signaling of the High Affinity IgE Receptor by FcεRIβ and the Potential Impact of FcεRIβ Splicing in Allergic Inflammation

“…FcεRIβ facilitates signaling by binding SRC family kinases such as LYN via its own, non-canonical ITAM, located near its C-terminus. This acts as an activation loop [ 29 , 30 , 31 ], which subsequently leads to recruitment and phosphorylation of SYK from the cell cytosol [ 32 , 33 ] The mechanism of SYK recruitment and phosphorylation appears to be modulated by the kinetics of FcεRI aggregation, since the aggregation of larger numbers of IgE-FcεRI complexes provides a greater pool of phosphorylated ITAMs for SYK to bind to [ 34 , 35 ]. The formation of FcεRI aggregates (which can differ in the orientation and distance of the receptors within aggregates) appears to modulate signaling efficiency, as well as negative regulation of FcεRI activation by the preferential recruitment of inhibitory phosphatases over SYK [ 36 , 37 ].…”

Perilla Leaf Extract Attenuates Asthma Airway Inflammation by Blocking the Syk Pathway

“…Spleen tyrosine kinase (Syk) is a 72 kD cytoplasmic nonreceptor tyrosine kinase, which plays key roles in allergic asthma inflammatory responses [ 10 ] through promoting IgE activation, degranulation mediator release, eicosanoid production, and cytokine synthesis [ 11 , 12 ]. Syk activation occurs primarily through SH2 domains binding to Fc ε RI-ITAMs (immunoreceptor tyrosine-based activation motifs) [ 13 ] and then phosphorylating its own activation loop in tyrosine 525/526 to fully activate itself and transduce Fc ε RI signaling in mast cells and basophils [ 14 ]. As an upstream signaling molecule, activated Syk regulates multiple signaling molecules and amplifies inflammatory signals [ 15 ], including nuclear factor- κ B (NF- κ B), protein kinase C (PKC), and cytosolic phospholipase A 2 (cPLA 2 ) [ 16 ].…”