Integrins Influence the Size and Dynamics of Signaling Microclusters in a Pyk2-dependent Manner

Steblyanko, Maria; Anikeeva, Nadia; Campbell, Kerry S.; Keen, James H.; Sykulev, Yuri

doi:10.1074/jbc.m114.614719

Cited by 24 publications

(29 citation statements)

References 50 publications

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“…We suspect that the initial adhesion is due to cytoskeletal changes that occur as the CTL binds to ICAM-1 immobilized on the beads or plastic surface. Consistent with this, a recent study showed that NK adhesion to ICAM-1-expressing cells was reduced upon Pyk2 siRNA knockdown by ∼20-25% when examined at 20 min after conjugate formation (20), which could be due to the decreased mobility of microclusters (18). When the expression of a number of genes encoding proteins known to function upstream and downstream of Pyk2 was downregulated with siRNA, only knockdown of CasL blocked adhesion of NK cells (20).…”

Section: Discussionsupporting

confidence: 65%

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Pyk2 Controls Integrin-Dependent CTL Migration through Regulation of De-Adhesion

Cheung

Ostergaard

2016

The Journal of Immunology

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Protein tyrosine kinase 2 (Pyk2) is required for T cell adhesion to ICAM-1; however, the mechanism by which it regulates adhesion remains unexplored. Pyk2 function in murine CTL clones and activated ex vivo CD8+ T cells was disrupted by pharmacological inhibition, knockdown of expression with small interfering RNA, or expression of the dominant-negative C-terminal domain. We found that Pyk2 is not absolutely required for adhesion of CTL to ICAM-1, but rather delays the initial adhesion. Disruption of Pyk2 function caused cells to display an unusual elongated appearance after 1 h on ICAM-1, consistent with abnormally strong adhesion. Furthermore, the random mobility of CTL on ICAM-1 was severely compromised using all three methods of disrupting Pyk2 function. Live-cell imaging studies revealed that the decreased migration is the result of a defect in the detachment from ICAM-1 at the trailing edge when Pyk2 function is inhibited. Examination of Pyk2 tyrosine phosphorylation in normal polarized cells demonstrated that Pyk2 phosphorylated at Y579 and Y580 preferentially localizes to the leading edge, whereas Y881-phosphorylated Pyk2 is enriched at the trailing edge, suggesting that the tyrosine phosphorylation of Pyk2 is spatially regulated in migrating CTL. Additionally, inhibition of Pyk2 caused cells to form multiple LFA-1–rich tails at the trailing edge, most likely resulting from a defect in LFA-1 release required for forward movement. Our results show that Pyk2 contributes to CTL migration by regulating detachment of CTL at the trailing edge, which could explain why Pyk2 is important for chemotactic and migratory responses.

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Section: Discussionsupporting

confidence: 65%

“…In T cells, Pyk2 is critical for short-lived effector CD8 + T cell generation as well as adhesion on ICAM-1 substrates (17). More recently, it was demonstrated that inhibition of Pyk2 activity in NK-92 cells decreases the integrin-regulated mobility and displacement of the microclusters that form at the contact zone with a target cell (18).…”

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confidence: 99%

Pyk2 Controls Integrin-Dependent CTL Migration through Regulation of De-Adhesion

Cheung

Ostergaard

2016

The Journal of Immunology

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“…We have previously shown that MHC clusters on APC may include heterotypic membrane proteins such as ICAM-1, a ligand for LFA-1 integrin, which augment the sensitivity of the T cell response (1). This is consistent with our recent findings showing that integrinmediated signaling regulates the efficiency of proximal signaling in cytotoxic lymphocytes (46,47). Productive engagement of other receptors leads to recruitment of adaptor and signaling proteins to the TCR-containing clusters and formation of signalosomes (48) that integrate receptor-mediated signaling.…”

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confidence: 90%

Extent of MHC Clustering Regulates Selectivity and Effectiveness of T Cell Responses

Anikeeva

Fischer

Blanchette

et al. 2019

The Journal of Immunology

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“…Prior to the polarization of the microtubule organizing center (MTOC) to the IS, lytic granules converge to the MTOC in a dynein-dependent minusended directed manner that is independent of actin polymerization and microtubule dynamics; as such, ligation of LFA-1 or CD28 alone is sufficient to induce convergence (19,20). LFA-1-mediated outside-in signaling leads to Factin polymerization and reorganization, phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2) generation, and the activation of protein tyrosine kinases, including Src family kinases (21,22). Activating signaling and sustained F-actin remodeling promotes the polarization of the MTOC, with converged lytic granules, to the lytic IS (23)(24)(25).…”

Section: Introductionmentioning

confidence: 99%

CD56 regulates human NK cell cytotoxicity through Pyk2

Gunesch

Dixon

Ebrahim

et al. 2020

Preprint

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Natural killer (NK) cells are innate immune cells that control viral infection and tumorigenic cell growth through targeted cell lysis and cytokine secretion. Human NK cells are classically defined as CD56 + CD3 − in peripheral blood. CD56 is neural cell adhesion molecule (NCAM1), and despite its ubiquitous expression on human NK cells, the role of CD56 in human NK cell cytotoxic function has not been fully explored. In non-immune cells, NCAM can induce signaling, mediate adhesion, and promote exocytosis, in part through interactions with focal adhesion kinase (FAK). Here we describe the generation and use of CD56-deficient human NK cell lines to define a novel requirement for CD56 in target cell lysis. Namely, we demonstrate that deletion of CD56 on the NK92 cell line led to impaired cytotoxic function against multiple susceptible target cell lines. Deletion of CD56 in a second NK cell line, YTS cells, led to a less severe cytotoxicity defect but impairment in cytokine secretion. Confocal microscopy of wild-type and CD56-KO NK92 cells conjugated to susceptible targets revealed that CD56-KO cells failed to polarize during immunological synapse (IS) formation and had severely impaired exocytosis of lytic granules at the IS. Phosphorylation of the FAK family member Pyk2 at tyrosine 402 was decreased in NK92 CD56-KO cells, demonstrating a functional link between CD56 and IS formation and signaling in human NK cells. Cytotoxicity, lytic granule exocytosis, and the phosphorylation of Pyk2 were rescued by the reintroduction of NCAM140 (CD56), into NK92 CD56-KO cells. These data highlight a novel functional role for CD56 in stimulating exocytosis and promoting cytotoxicity in human NK cells. cytotoxicity | natural killer cell | NCAM | CD56 Correspondence: em3375@cumc.columbia.edu 2 | bioRχiv Mace et al. | CD56 and human NK cell cytotoxicity

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Integrins Influence the Size and Dynamics of Signaling Microclusters in a Pyk2-dependent Manner

Cited by 24 publications

References 50 publications

Pyk2 Controls Integrin-Dependent CTL Migration through Regulation of De-Adhesion

Pyk2 Controls Integrin-Dependent CTL Migration through Regulation of De-Adhesion

Extent of MHC Clustering Regulates Selectivity and Effectiveness of T Cell Responses

CD56 regulates human NK cell cytotoxicity through Pyk2

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