Monovalent and Multivalent Ligation of the B Cell Receptor Exhibit Differential Dependence upon Syk and Src Family Kinases

Mukherjee, Sayak; Zhu, Jing; Zikherman, Julie; Parameswaran, Ramya; Kadlecek, Theresa A.; Wang, Qi; Au-Yeung, Byron B; Ploegh, Hidde L.; Kuriyan, John; Das, Jayajit; Weiss, Arthur

doi:10.1126/scisignal.2003220

Cited by 76 publications

(91 citation statements)

References 67 publications

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“…1D). Unexpectedly, we found that SHIP-EGFP is significantly recruited to the plasma membrane when the BCR is ligated alone, and there is in fact no measurable difference in the magnitude of recruitment induced by F(ab9) 2 or intact anti-IgM (Fig. 1E).…”

Section: Fcgriib Engagement Does Not Significantly Impact Recruitmentmentioning

confidence: 76%

“…Each group . Two-way ANOVA did not reveal any significant difference between F(ab9) 2 and intact anti-Ig. of recovery curves was then fit to the following equation, with the constraint that…”

Section: Frap Experimentsmentioning

confidence: 99%

“…Live cell imaging of FcgRIIB + Ramos coexpressing SHIP-EGFP and a red fluorescent membrane marker indicated that recruitment occurs within the first minute of stimulation with either F(ab9) 2 or intact anti-IgM and did not reveal any significant differences between these stimuli (Fig. 2A).…”

Section: Fcgriib Engagement Does Not Significantly Impact Recruitmentmentioning

confidence: 99%

“…5B). Interestingly, the C-terminal region of SHIP is absolutely required for recruitment of SHIP-EGFP upon stimulation with F(ab9) 2 Ab, but it has a less significant role for recruitment after intact Ab stimulation (Fig. 5C).…”

Section: Dependence Of Ship Membrane Recruitment On Upstream Kinases mentioning

confidence: 99%

“…The BCR signaling chains Iga and Igb each contain two ITAMs that can be phosphorylated upon receptor cross-linking by the Src family kinase Lyn and, in some contexts, Syk (1,2). Doubly phosphorylated Iga/b ITAMs provide binding sites for Syk, thereby initiating its activation (3).…”

mentioning

confidence: 99%

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FcγRIIB-Independent Mechanisms Controlling Membrane Localization of the Inhibitory Phosphatase SHIP in Human B Cells

Pauls

Ray

Hou

et al. 2016

The Journal of Immunology

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SHIP is an important regulator of immune cell signaling that functions to dephosphorylate the phosphoinositide phosphatidylinositol 3,4,5-trisphosphate at the plasma membrane and mediate protein–protein interactions. One established paradigm for SHIP activation involves its recruitment to the phospho-ITIM motif of the inhibitory receptor FcγRIIB. Although SHIP is essential for the inhibitory function of FcγRIIB, it also has critical modulating functions in signaling initiated from activating immunoreceptors such as B cell Ag receptor. In this study, we found that SHIP is indistinguishably recruited to the plasma membrane after BCR stimulation with or without FcγRIIB coligation in human cell lines and primary cells. Interestingly, fluorescence recovery after photobleaching analysis reveals differential mobility of SHIP–enhanced GFP depending on the mode of stimulation, suggesting that although BCR and FcγRIIB can both recruit SHIP, this occurs via distinct molecular complexes. Mutagenesis of a SHIP–enhanced GFP fusion protein reveals that the SHIP–Src homology 2 domain is essential in both cases whereas the C terminus is required for recruitment via BCR stimulation, but is less important with FcγRIIB coligation. Experiments with pharmacological inhibitors reveal that Syk activity is required for optimal stimulation-induced membrane localization of SHIP, whereas neither PI3K or Src kinase activity is essential. BCR-induced association of SHIP with binding partner Shc1 is dependent on Syk, as is tyrosine phosphorylation of both partners. Our results indicate that FcγRIIB is not uniquely able to promote membrane recruitment of SHIP, but rather modulates its function via formation of distinct signaling complexes. Membrane recruitment of SHIP via Syk-dependent mechanisms may be an important factor modulating immunoreceptor signaling.

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Section: Fcgriib Engagement Does Not Significantly Impact Recruitmentmentioning

confidence: 76%

Section: Frap Experimentsmentioning

confidence: 99%

Section: Fcgriib Engagement Does Not Significantly Impact Recruitmentmentioning

confidence: 99%

Section: Dependence Of Ship Membrane Recruitment On Upstream Kinases mentioning

confidence: 99%

mentioning

confidence: 99%

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FcγRIIB-Independent Mechanisms Controlling Membrane Localization of the Inhibitory Phosphatase SHIP in Human B Cells

Pauls

Ray

Hou

et al. 2016

The Journal of Immunology

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B Lymphocytes: Receptors

DeFranco

2015

Encyclopedia of Life Sciences

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B lymphocytes can synthesise Ig heavy chains that are either secreted or membrane‐bound, the latter having a transmembrane domain and a short cytoplasmic domain. Membrane‐bound Ig heavy chains and Ig light chains assemble in the ER with each other and with a heterodimer of Igα and Igβ, which mediate signalling and internalisation functions. The complex of these four polypeptides is called the B cell antigen receptor (BCR). BCR signalling occurs via three types of tyrosine kinases. Src‐family kinases phosphorylate the two tyrosine‐containing motif in the cytoplasmic domains of Igα and Igβ (the ITAMs), which recruits a second type of tyrosine kinase Syk. Syk is responsible for most downstream signalling, although calcium and diacylglycerol also require a third tyrosine kinase Btk. BCR signalling pathways are central to virtually all aspects of B cell biology, as they are required for B cell development, tolerance induction, survival and activation. Key Concepts Naïve B cells express their immunoglobulin as a cell surface receptor for antigen, called the BCR (B cell receptor for antigen). In developing precursors of B cells, the successful generation of an immunoglobulin heavy chain is sensed by the incorporation of that heavy chain into pre‐BCR molecules, which exhibit constitutive signalling. Signalling by the BCR controls development, survival and activation of B cells in a manner that is dependent on the developmental stage or activation state of the B cell. This signalling is important both for tolerisation of self‐reactive B cells and for activation of B cells recognising foreign antigen. Co‐receptors that recognise complement components, sialic acid residues, or IgG bound to antigens strongly enhance or inhibit signalling by the BCR to aid in self‐non‐self‐discrimination. The BCR is also an endocytic receptor that delivers antigen to internal compartments where protein antigens are processed into peptides and loaded onto MHC class II molecules to facilitate interactions with T cells, which are essential for production of high‐affinity antibodies. Defects in generation of BCRs or signalling components of the BCR result in B cell immunodeficiencies, the most common of which is due to loss‐of‐function mutations of the intracellular protein tyrosine kinase Btk, referred to as X‐linked agammaglobulinemia. B cell malignancies often have activated BCR signalling as a driver of their proliferation and/or survival, and selective Btk inhibitors are now approved therapeutics for some types of B cell malignancies.

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Differences in the dynamics of the tandem‐SH2 modules of the Syk and ZAP‐70 tyrosine kinases

et al. 2021

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The catalytic activity of Syk‐family tyrosine kinases is regulated by a tandem Src homology 2 module (tSH2 module). In the autoinhibited state, this module adopts a conformation that stabilizes an inactive conformation of the kinase domain. The binding of the tSH2 module to phosphorylated immunoreceptor tyrosine‐based activation motifs necessitates a conformational change, thereby relieving kinase inhibition and promoting activation. We determined the crystal structure of the isolated tSH2 module of Syk and find, in contrast to ZAP‐70, that its conformation more closely resembles that of the peptide‐bound state, rather than the autoinhibited state. Hydrogen–deuterium exchange by mass spectrometry, as well as molecular dynamics simulations, reveal that the dynamics of the tSH2 modules of Syk and ZAP‐70 differ, with most of these differences occurring in the C‐terminal SH2 domain. Our data suggest that the conformational landscapes of the tSH2 modules in Syk and ZAP‐70 have been tuned differently, such that the autoinhibited conformation of the Syk tSH2 module is less stable. This feature of Syk likely contributes to its ability to more readily escape autoinhibition when compared to ZAP‐70, consistent with tighter control of downstream signaling pathways in T cells.

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Monovalent and Multivalent Ligation of the B Cell Receptor Exhibit Differential Dependence upon Syk and Src Family Kinases

Cited by 76 publications

References 67 publications

FcγRIIB-Independent Mechanisms Controlling Membrane Localization of the Inhibitory Phosphatase SHIP in Human B Cells

FcγRIIB-Independent Mechanisms Controlling Membrane Localization of the Inhibitory Phosphatase SHIP in Human B Cells

B Lymphocytes: Receptors

Differences in the dynamics of the tandem‐SH2 modules of the Syk and ZAP‐70 tyrosine kinases

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