Showing papers on "Src family kinase published in 2020"
TL;DR: The findings supported the critical role of Fyn in HSC activation and development of liver fibrosis and suggested that Fyn could serve as a promising drug target for Liver fibrosis treatment.
Abstract: Recent studies suggest that Src family kinase (SFK) plays important roles in systemic sclerosis and pulmonary fibrosis. However, how SFKs contributed to the pathogenesis of liver fibrosis remains largely unknown. Here, we investigated the role of Fyn, a member of SFK, in hepatic stellate cell (HSC) activation and liver fibrosis, and evaluated the anti-fibrotic effects of Saracatinib, a clinically proven safe Fyn inhibitor. Fyn activation was examined in human normal and fibrotic liver tissues. The roles of Fyn in HSC activation and liver fibrosis were evaluated in HSC cell lines by using Fyn siRNA and in Fyn knockout mice. The effects of Saracatinib on HSC activation and liver fibrosis were determined in primary HSCs and CCl4 induced liver fibrosis model. We showed that the Fyn was activated in the liver of human fibrosis patients. TGF-β induced the activation of Fyn in HSC cell lines. Knockdown of Fyn significantly blocked HSC activation, proliferation, and migration. Fyn deficient mice were resistant to CCl4 induced liver fibrosis. Saracatinib treatment abolished the activation of Fyn, downregulated the Fyn/FAK/N-WASP signaling in HSCs, and subsequently prevented the activation of HSCs. Saracatinib treatment significantly reduced the severity liver fibrosis induced by CCl4 in mice. In conclusions, our findings supported the critical role of Fyn in HSC activation and development of liver fibrosis. Fyn could serve as a promising drug target for liver fibrosis treatment. Fyn inhibitor Saracatinib significantly inhibited HSC activation and attenuated liver fibrosis in mouse model.
29 citations
TL;DR: Significance Lck is critical for T cell development and activation, as it is the first kinase transducing TCR signaling, and evidence is presented that the two pools of Lck have different molecular properties.
Abstract: Src family kinase Lck plays critical roles during T cell development and activation, as it phosphorylates the TCR/CD3 complex to initiate TCR signaling. Lck is present either in coreceptor-bound or coreceptor-unbound (free) forms, and we here present evidence that the two pools of Lck have different molecular properties. We discovered that the free Lck fraction exhibited higher mobility than CD8α-bound Lck in OT-I T hybridoma cells. The free Lck pool showed more activating Y394 phosphorylation than the coreceptor-bound Lck pool. Consistent with this, free Lck also had higher kinase activity, and free Lck mediated higher T cell activation as compared to coreceptor-bound Lck. Furthermore, the coreceptor-Lck coupling was independent of TCR activation. These findings give insights into the initiation of TCR signaling, suggesting that changes in coreceptor-Lck coupling constitute a mechanism for regulation of T cell sensitivity.
28 citations
TL;DR: This review addressed the Src involvement in physiological as well as in pathological conditions, especially in tumors, by considering its function as both tyrosine kinase and adapting molecule.
Abstract: Src is the representative member of the Src-family kinases (SFKs), a group of tyrosine kinases involved in several cellular processes. Its main function has been for long confined to the plasma membrane/cytoplasm compartment, being a myristoylated protein anchored to the cell membrane and functioning downstream to receptors, most of them lacking intrinsic kinase activity. In the last decades, new roles for some SFKs have been described in the nuclear compartment, suggesting that these proteins can also be involved in directly regulating gene transcription or nucleoskeleton architecture. In this review, we focused on those nuclear functions specifically attributable to Src, by considering its function as both tyrosine kinase and adapting molecule. In particular, we addressed the Src involvement in physiological as well as in pathological conditions, especially in tumors.
27 citations
TL;DR: It is demonstrated that SFK play a role in neutrophil post-arrest modifications and extravasation during acute inflammation, which may support the current efforts to use SFK-inhibitors in inflammatory diseases with unwanted neutrophils.
Abstract: Leukocyte recruitment into inflamed tissue is highly dependent on the activation and binding of integrins to their respective ligands, followed by the induction of various signaling events within the cell referred to as outside-in signaling. Src family kinases (SFK) are the central players in the outside-in signaling process, assigning them a critical role for proper immune cell function. Our study investigated the role of SFK on neutrophil recruitment in vivo using Hck-/- Fgr-/- Lyn-/- mice, which lack SFK expressed in neutrophils. We show that loss of SFK strongly reduces neutrophil adhesion and post-arrest modifications in a shear force dependent manner. Additionally, we found that in the absence of SFK, neutrophils display impaired Rab27a-dependent surface mobilization of neutrophil elastase, VLA3 and VLA6 containing vesicles. This results in a defect in neutrophil vascular basement membrane penetration and thus strongly impaired extravasation. Taken together, we demonstrate that SFK play a role in neutrophil post-arrest modifications and extravasation during acute inflammation. These findings may support the current efforts to use SFK-inhibitors in inflammatory diseases with unwanted neutrophil recruitment.
15 citations
TL;DR: How leptin regulates surface expression of NR2B-containing NMDA receptors in hippocampal neurons is examined and it is shown that LepRb forms a complex with NR2 B and Fyn, a member of the Src family kinases, is required for leptin stimulated NR2BY1472 phosphorylation.
Abstract: Activation of the leptin receptor, LepRb, by the adipocytokine/neurotrophic factor leptin in the central nervous system has procognitive and antidepressive effects. Leptin has been shown to increase glutamatergic synaptogenesis in multiple brain regions. In contrast, mice that have a mutation in the LepRb gene show abnormal synapse development in the hippocampus as well as deficits in cognition and increased depressive-like symptoms. Leptin increases glutamatergic synaptogenesis, in part, through enhancement of N-methyl-D-aspartic acid (NMDA) receptor function; yet the underlying signaling pathway is not known. In this study, we examine how leptin regulates surface expression of NR2B-containing NMDA receptors in hippocampal neurons. Leptin stimulation increases NR2BY1472 phosphorylation, which is inhibited by the Src family kinase inhibitor, PP1. Moreover, we show that Fyn, a member of the Src family kinases, is required for leptin-stimulated NR2BY1472 phosphorylation. Furthermore, inhibiting Y1472 phosphorylation with either a dominant negative Fyn mutant or an NR2B mutant that lacks the phosphorylation site (NR2BY1472F) blocks leptin-stimulated synaptogenesis. Additionally, we show that LepRb forms a complex with NR2B and Fyn. Taken together, these findings expand our knowledge of the LepRb interactome and the mechanisms by which leptin stimulates glutamatergic synaptogenesis in the developing hippocampus. Comprehending these mechanisms is key for understanding dendritic spine development and synaptogenesis, alterations of which are associated with many neurological disorders.
14 citations
TL;DR: Analysis of the model revealed that the CLEC-2 receptor clustering leading to the membrane-based signalosome formation is a critical element required for the accurate description of the experimental data, and suggests a refined picture of the platelet signal transduction network associated with CLEc-2.
14 citations
TL;DR: It is reported here that SCIMP is phosphorylated and activated in response to agonists of multiple TLRs, including TLR2,TLR3, TLR4, and TLR9, and is thus a universal TLR adaptor for scaffolding the Lyn tyrosine kinase and its effectors to enable responses against a wide range of danger signals.
Abstract: In innate immune cells, pathogens and danger signals activate TLRs, unleashing potent and tailored inflammatory responses. Previously, we reported that an immune-specific transmembrane adaptor, SLP adaptor and CSK interacting membrane protein (SCIMP), interacts with TLR4 via direct binding to its cytoplasmic TIR domain. SCIMP scaffolds a Src family kinase, Lyn, for TLR4 phosphorylation and activation. Consequently, SCIMP is able to direct selective production of the proinflammatory cytokines IL-6 and IL-12p40 downstream of TLR4 in macrophages. Here, we set out to investigate whether SCIMP also acts as an adaptor for other TLR family members. We report here that SCIMP is phosphorylated and activated in response to agonists of multiple TLRs, including TLR2, TLR3, TLR4, and TLR9. SCIMP also interacts with TLRs that are known to signal from both the cell surface and endosomal compartments. In so doing, this transmembrane adaptor presents Lyn, along with other effectors such as Grb2, Csk, and SLP65, to multiple TLRs during cellular activation. CRISPR-mediated knockout or silencing of SCIMP in macrophages alters TLR signaling outputs and the production of IL-6 and IL-12p40 downstream of multiple TLRs, and upon challenge with live bacteria. Furthermore, the selectivity in cytokine responses is preserved downstream of TLR3, with inducible expression of Il-12p40 and IL-6, but not IFNβ, being SCIMP dependent. SCIMP is thus a universal TLR adaptor for scaffolding the Lyn tyrosine kinase and its effectors to enable responses against a wide range of danger signals.
14 citations
TL;DR: The key nongenetic mechanisms underlying SFK tumour activities in colorectal cancer are described and how these mechanisms could be exploited in therapeutic strategies to target SFK signalling in metastatic colon cancer are discussed.
Abstract: Src, originally identified as an oncogene, is a membrane-anchored tyrosine kinase and the Src family kinase (SFK) prototype. SFKs regulate the signalling induced by a wide range of cell surface receptors leading to epithelial cell growth and adhesion. In the intestine, the SFK members Src, Fyn and Yes regulate epithelial cell proliferation and migration during tissue regeneration and transformation, thus implicating conserved and specific functions. In patients with colon cancer, SFK activity is a marker of poor clinical prognosis and a potent driver of metastasis formation. These tumorigenic activities are linked to SFK capacity to promote the dissemination and tumour-initiating capacities of epithelial tumour cells. However, it is unclear how SFKs promote colon tumour formation and metastatic progression because SFK-encoding genes are unfrequently mutated in human cancer. Here, we review recent findings on SFK signalling during intestinal homeostasis, regeneration and tumorigenesis. We also describe the key nongenetic mechanisms underlying SFK tumour activities in colorectal cancer, and discuss how these mechanisms could be exploited in therapeutic strategies to target SFK signalling in metastatic colon cancer.
13 citations
TL;DR: The data indicate that STAT3 differentially regulates early- and late-phase TLR4-mediated inflammatory responses, and iNOS expression was downregulated after prolonged treatment with LPS.
Abstract: Toll-like receptor 4 (TLR4) signaling is an important therapeutic target to manage lipopolysaccharide (LPS)-induced inflammation. The transcription factor signal transducer and activator of transcription 3 (STAT3) has been identified as an important regulator of various immune-related diseases and has generated interest as a therapeutic target. Here, we investigated the time-dependent roles of STAT3 in LPS-stimulated RAW264.7 macrophages. STAT3 inhibition induced expression of the pro-inflammatory genes iNOS and COX-2 at early time points. STAT3 depletion resulted in regulation of nuclear translocation of nuclear factor (NF)-κB subunits p50 and p65 and IκBα/Akt/PI3K signaling. Moreover, we found that one Src family kinase, Lyn kinase, was phosphorylated in STAT3 knockout macrophages. In addition to using pharmacological inhibition of NF-κB, we found out that STAT3KO activation of NF-κB subunit p50 and p65 and expression of iNOS was significantly inhibited; furthermore, Akt tyrosine kinase inhibitors also inhibited iNOS and COX-2 gene expression during early time points of LPS stimulation, demonstrating an NF-κB- Akt-dependent mechanism. On the other hand, iNOS expression was downregulated after prolonged treatment with LPS. Activation of NF-κB signaling was also suppressed, and consequently, nitric oxide (NO) production and cell invasion were repressed. Overall, our data indicate that STAT3 differentially regulates early- and late-phase TLR4-mediated inflammatory responses.
13 citations
TL;DR: For the first time, it is demonstrated that TWEAK promoted HSC migration through the activation of EGFR/Src and PI3K/AKT pathways, and showed a novel potential mechanism of H SC migration regulated by TWE AK.
11 citations
TL;DR: It is indicated that genetic knockdown or pharmacological inhibition of Fyn dampens systemic inflammation, demonstrating for the first time that Fyn kinase plays a critical role in mediating the endotoxic inflammatory response.
TL;DR: Findings elucidate important mechanisms of CIC regulation and provide the rationale to target c-Src alongside ERK pathway inhibitors as a way to fully restore CIC tumor suppressor function in neoplasms such as GBM.
Abstract: Capicua (CIC) is a transcriptional repressor that counteracts activation of genes in response to receptor tyrosine kinase (RTK)/Ras/ERK signaling. Following activation of RTK, ERK enters the nucleus and serine-phosphorylates CIC, releasing it from its targets to permit gene expression. We recently showed that ERK triggers ubiquitin-mediated degradation of CIC in glioblastoma (GBM). In this study, we examined whether another important downstream effector of RTK/EGFR, the non-RTK c-Src, affects CIC repressor function in GBM. We found that c-Src binds and tyrosine-phosphorylates CIC on residue 1455 to promote nuclear export of CIC. On the other hand, CIC-mutant allele (CIC-Y1455F), that escapes c-Src-mediated tyrosine phosphorylation, remains localized to the nucleus and retains strong repressor function against CIC targets, the oncogenic transcription factors ETV1 and ETV5. Furthermore, we show that the orally available Src family kinase inhibitor, dasatinib, which prevents EGF-mediated tyrosine phosphorylation of CIC and attenuates elevated ETV1 and ETV5 levels, reduces viability of GBM cells and glioma stem cells (GSC), but not of their control cells with undetectable c-Src activity. In fact, GBM cells and GSC expressing the tyrosine-defective CIC mutant (Y1455F) lose sensitivity to dasatinib, further endorsing the effect of dasatinib on Src-mediated tyrosine phosphorylation of CIC. These findings elucidate important mechanisms of CIC regulation and provide the rationale to target c-Src alongside ERK pathway inhibitors as a way to fully restore CIC tumor suppressor function in neoplasms such as GBM. IMPLICATIONS: c-Src tyrosine-phosphorylates CIC exports to cytoplasm and inactivates its repressor function in GBM.
TL;DR: A first-of-its-kind atomically detailed structural ensemble of a membrane-bound intrinsically disordered protein of the Src family kinase Hck, offering the first estimations of the lengthscale associated with the concept of SFK subcellular localization.
TL;DR: Adulthood social isolation in rats enhances the Fyn-mGlu5 interaction, which appears to be critical for the upregulation of surface mGLU5 receptor expression in striatal neurons.
TL;DR: It is suggested that WHIMP-associated actin assembly plays a direct role in membrane protrusion, but also results in feedback control of tyrosine kinase signaling to modulate the activation of multiple WASP-family members.
Abstract: Cell motility is governed by cooperation between the Arp2/3 complex and nucleation-promoting factors from the Wiskott-Aldrich Syndrome Protein (WASP) family, which together assemble actin filament networks to drive membrane protrusion. Here we identify WHIMP (WAVE Homology In Membrane Protrusions) as a new member of the WASP family. The Whimp gene is encoded on the X chromosome of a subset of mammals, including mice. Murine WHIMP promotes Arp2/3-dependent actin assembly, but is less potent than other nucleation factors. Nevertheless, WHIMP-mediated Arp2/3 activation enhances both plasma membrane ruffling and wound healing migration, whereas WHIMP depletion impairs protrusion and slows motility. WHIMP expression also increases Src-family kinase activity, and WHIMP-induced ruffles contain the additional nucleation-promoting factors WAVE1, WAVE2, and N-WASP, but not JMY or WASH. Perturbing the function of Src-family kinases, WAVE proteins, or Arp2/3 complex inhibits WHIMP-driven ruffling. These results suggest that WHIMP-associated actin assembly plays a direct role in membrane protrusion, but also results in feedback control of tyrosine kinase signaling to modulate the activation of multiple WASP-family members.
TL;DR: It is demonstrated that PAG1 directs SFK intracellular localization to control activity and to mediate signaling by RTKs that induce neuronal differentiation, and disruption of FYN localization led to the observed defects in differentiation.
Abstract: Different receptor tyrosine kinases employ shared downstream cell signaling pathways to induce differentiation or proliferation. The scaffold protein, PAG1 controls SRC-family kinase (SFK) activity...
TL;DR: SYK lowered the threshold for SFK activity necessary to initiate antigen receptor signaling in both T and B cells, however, neither SYK nor ZAP-70 were able to initiate signaling independently of SFK.
TL;DR: Asa may serve as a potential novel Src family kinase inhibitor to inhibit IgE-dependent andIgE-independent allergic reaction and treat anaphylactic disease.
TL;DR: The collected findings provide the first comprehensive overview of retinal tyrosine kinome changes in the Akimba model of diabetic retinopathy and for the first time highlight Src family kinase inhibition using highly specific inhibitors as an attractive therapeutic intervention for retinal vascular pathology.
TL;DR: Investigation of the role of autocrine LIF in human esophageal squamous cell carcinoma confirmed that SFK activation and LIF expression were correlated with YAP activation in human ESCC clinical samples, and simultaneous inhibition of the SFK–YAP and JAK–STAT3 pathways inhuman ESCC cells was more effective at suppressing cell proliferation than single inhibition.
Abstract: The IL6 family of cytokines, including IL6 and leukemia-inhibitory factor (LIF), are induced during inflammation and are also expressed in many types of cancer where they play an important role in tumor development. IL6 family cytokines mainly activate the JAK-STAT3 pathway via the coreceptor, gp130, and IL6 is known to activate the Src family kinase (SFK)-Yes-associated protein (YAP) pathway. The current study investigated the role of autocrine LIF in human esophageal squamous cell carcinoma (ESCC) that highly expresses LIF. LIF knockdown had various effects on cancer cells, including profound changes in gene expression, suppression of cell proliferation, migration/invasion and sphere formation, and induction of apoptosis. Similar to IL6, LIF activated the SFK-YAP pathway as well as the JAK-STAT3 pathway. LIF-induced YAP activation was more important for cancer cell proliferation than LIF-induced STAT3 activation, and concomitant YAP and STAT3 activation completely compensated for the role of LIF in human ESCC growth. We also confirmed that SFK activation and LIF expression were correlated with YAP activation in human ESCC clinical samples. Furthermore, simultaneous inhibition of the SFK-YAP and JAK-STAT3 pathways in human ESCC cells was more effective at suppressing cell proliferation than single inhibition, and autocrine LIF signaling promoted human ESCC growth in vivo. Therefore, the LIF-SFK-YAP axis may represent a new therapeutic target for human ESCC. IMPLICATIONS: Autocrine LIF signaling promotes human ESCC progression via SFK-dependent YAP activation and is a new potential target of treatment for human ESCC.
TL;DR: The results suggest that there is limited transport or diffusion of Fyn kinase between the cytosol and nucleus in the cells, which is important for the maintenance of different equilibrium states of F Cyn in situ.
TL;DR: Deep sequencing confirmed representation of nearly all other missense mutations across the entire HCK open reading frame, suggesting that A-419259 and other C-helix-out Src-family kinase inhibitors may have a narrow path to acquired resistance in the context of AML cases where Hck is an oncogenic driver.
Abstract: Understanding cancer cell drug resistance to protein-tyrosine kinase inhibitors, which often arises from acquired mutations in the target kinase, is central to the development of more durable therapies. Experimental systems that reveal potential paths to resistance for a given inhibitor and kinase target have an important role in preclinical development of kinase inhibitor drugs. Here, we employed a codon mutagenesis strategy to define the mutational landscape of acquired resistance in HCK, a member of the SRC tyrosine kinase family and therapeutic target in acute myeloid leukemia (AML). Using PCR-based saturation mutagenesis, we created a cDNA library designed to replace each codon in the HCK open reading frame with all possible codons. This HCK mutant library was used to transform Rat-2 fibroblasts, followed by selection for resistant colonies with A-419259, a pyrrolopyrimidine HCK inhibitor and drug lead for AML. X-ray crystallography has shown that A-419259 binding induces outward rotation of the kinase domain αC-helix, a conformation incompatible with phosphotransfer. Remarkably, only a single resistance mutation evolved during A-419259 selection: histidine substitution for threonine at the gatekeeper position in the kinase domain. Deep sequencing confirmed representation of nearly all other missense mutations across the entire HCK open reading frame. This observation suggests that A-419259 and other C-helix-out Src-family kinase inhibitors may have a narrow path to acquired resistance in the context of AML cases where Hck is an oncogenic driver.
TL;DR: It is demonstrated that PAG1 directs SFK intracellular localization to control activity and to mediate signaling by RTKs that induce neuronal differentiation, and disruption of FYN localization led to the observed defects in differentiation.
Abstract: All receptor tyrosine kinases (RTKs) activate similar downstream signaling pathways through a common set of effectors, yet it is not fully understood how different receptors elicit distinct cellular responses to cause cell proliferation, differentiation, or other cell fates. We tested the hypothesis that regulation of SRC Family Kinase (SFK) signaling by the scaffold protein, PAG1, influences cell fate decisions following RTK activation. We generated a neuroblastoma cell line expressing a PAG1 fragment that lacks the membrane spanning domain (PAG1™-) and localized to the cytoplasm. PAG1™- cells exhibited higher amounts of active SFKs and increased growth rate. PAG1™- cells were unresponsive to TRKA and RET signaling, two RTKs that induce neuronal differentiation, but retained responses to EGFR and KIT. Under differentiation conditions, PAG1™- cells continued to proliferate and did not extend neurites or increase β-III tubulin expression. FYN and LYN were sequestered in multivesicular bodies (MVBs), and dramatically more FYN and LYN were in the lumen of MVBs in PAG1™- cells. In particular, activated FYN was sequestered in PAG1™- cells, suggesting that disruption of FYN localization led to the observed defects in differentiation. The results demonstrate that PAG1 directs SFK intracellular localization to control activity and to mediate signaling by RTKs that induce neuronal differentiation.
TL;DR: In vitro mast cell models found that saracatinib inhibited the degranulation response and cytokine production in RBL2H3 cells that were stimulated with IgE and antigen without affecting cell viability, indicating the valuable suggestion that sarcatinib and the Src family kinase inhibitors are good candidates for anti-anaphylaxis and anti-allergic drugs.
Abstract: The aim of this study was to evaluate the anti-anaphylactic and anti-allergic potentials of saracatinib, a Src family kinase inhibitor that was already shown to be safe in clinical trials when it was used as an anti-cancer drug. Using in vitro mast cell models, we found that saracatinib inhibited the degranulation response and cytokine production in RBL2H3 cells that were stimulated with IgE and antigen without affecting cell viability. Phosphorylation of Lyn, Akt, a PI3K substrate, and MAPKs including ERK, JNK, and p38, as well as the intracellular Ca2+ increase induced by this stimulation were also suppressed by saracatinib. This drug also inhibited symptoms in our established anaphylaxis mouse model, anaphylaxis-dependent spotted distribution of immune complex in skin (ASDIS). The intravenous injection of the mixture of IgE and antigen induced acute spotted distribution of immune complex in skin in hairless HR-1 mice, and its inhibition by intradermal injection of saracatinib was observed. Moreover, toluidine blue-stained skin sections indicated that the degranulation ratio of dermal mast cells was reduced in saracatinib-treated skin compared with vehicle-treated skin. Because only a few signaling inhibitors are used as anti-anaphylaxis and anti-allergic drugs, these results indicated the valuable suggestion that saracatinib and the Src family kinase inhibitors are good candidates for anti-anaphylaxis and anti-allergic drugs.