FGFR3 Associates with and Tyrosine-Phosphorylates p90RSK2, Leading to RSK2 Activation That Mediates Hematopoietic Transformation

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3722-3722
Author(s):  
Sumin Kang ◽  
Shannon Elf ◽  
Shaozhong Dong ◽  
Taro Hitosugi ◽  
Ailan Guo ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Tyrosine Phosphorylation ◽  
Tyrosine Kinases ◽  
Mapk Pathway ◽  
Critical Role ◽  
Ectopic Expression ◽  
Alpha Helix ◽  
Tail Region ◽  
Murine Bone Marrow

Abstract Dysregulation of receptor tyrosine kinase FGFR3 has been implicated to play a pathogenic role in a number of human hematopoietic malignancies and solid tumors. These include t(4;14) multiple myeloma associated with ectopic expression of FGFR3 and t(4;12)(p16;p13) acute myeloid leukemia associated with expression of a constitutively activated fusion tyrosine kinase TEL-FGFR3. We recently reported that FGFR3 directly tyrosine phosphorylates p90 Ribosomal S6 Kinase2(RSK2) at Y529, which consequently regulates RSK2 activation [Kang et al, Cancer Cell 2007 Sep;12(3):201–14]. Here we identified Y707 as an additional tyrosine site of RSK2 that is phosphorylated by FGFR3. Phosphorylation at Y707 contributes to RSK2 activation, through a putative disruption of the autoinhibitory αL-helix on the C-terminus of RSK2, unlike Y529 phosphorylation that facilitates ERK binding. To elucidate the role of tyrosine phosphorylation at Y707 induced by FGFR3 in RSK2 activation, we characterized the RSK2 mutants with single Y→A and Y→F substitutions at Y707. RSK2 Y707F demonstrated decreased kinase activity, suggesting substitution of Y707 attenuates activation of RSK2 induced by FGFR3. Tyrosine phosphorylation at Y529 by FGFR3 regulates RSK2 activation by facilitating inactive ERK binding, whereas substitution of Y707 in RSK2 does not similarly attenuate inactive ERK binding to RSK2. Phosphorylation at Y707 may regulate RSK2 activation by affecting the structure of the autoinhibitory C-terminal domain of RSK2 since the Y707 is localized at the C-terminal tail region which represents a conserved putative auto-inhibitory alpha helix. Since other tyrosine kinases including FGFR1 and Src also phosphorylate RSK2 at Y529 and Y707, tyrosine phosphorylation may be a general requirement for RSK2 activation through the ERK/MAPK pathway. Together, our current and previous findings represent a paradigm for tyrosine phosphorylation-dependent regulation of serine-threonine kinases. Moreover, we found that FGFR3 interacts with RSK2 through residue W332 in the linker region of RSK2, and that this association is required for FGFR3-dependent phosphorylation of RSK2 at Y529 and Y707, and subsequent RSK2 activation. Furthermore, in a murine bone marrow transplant assay, genetic deficiency in RSK2 resulted in a significantly delayed and attenuated myeloproliferative syndrome induced by TEL-FGFR3 as compared with wild type cells, suggesting a critical role of RSK2 in FGFR3-induced hematopoietic transformation.

scholarly journals Fibroblast Growth Factor Receptor 3 Associates with and Tyrosine Phosphorylates p90 RSK2, Leading to RSK2 Activation That Mediates Hematopoietic Transformation

2009 ◽  
Vol 29 (8) ◽  
pp. 2105-2117 ◽  
Author(s):  
Sumin Kang ◽  
Shannon Elf ◽  
Shaozhong Dong ◽  
Taro Hitosugi ◽  
Katherine Lythgoe ◽  
...  
Keyword(s):  
Growth Factor ◽  
Tyrosine Kinase ◽  
Fibroblast Growth Factor ◽  
Fibroblast Growth Factor Receptor ◽  
Critical Role ◽  
Ectopic Expression ◽  
Growth Factor Receptor ◽  
Marrow Transplant ◽  
Fibroblast Growth ◽  
Murine Bone Marrow

ABSTRACT Dysregulation of the receptor tyrosine kinase fibroblast growth factor receptor 3 (FGFR3) plays a pathogenic role in a number of human hematopoietic malignancies and solid tumors. These include t(4;14) multiple myeloma associated with ectopic expression of FGFR3 and t(4;12)(p16;p13) acute myeloid leukemia associated with expression of a constitutively activated fusion tyrosine kinase, TEL-FGFR3. We recently reported that FGFR3 directly tyrosine phosphorylates RSK2 at Y529, which consequently regulates RSK2 activation. Here we identified Y707 as an additional tyrosine in RSK2 that is phosphorylated by FGFR3. Phosphorylation at Y707 contributes to RSK2 activation, through a putative disruption of the autoinhibitory αL-helix on the C terminus of RSK2, unlike Y529 phosphorylation, which facilitates ERK binding. Moreover, we found that FGFR3 interacts with RSK2 through residue W332 in the linker region of RSK2 and that this association is required for FGFR3-dependent phosphorylation of RSK2 at Y529 and Y707, as well as the subsequent RSK2 activation. Furthermore, in a murine bone marrow transplant assay, genetic deficiency in RSK2 resulted in a significantly delayed and attenuated myeloproliferative syndrome induced by TEL-FGFR3 as compared with wild-type cells, suggesting a critical role of RSK2 in FGFR3-induced hematopoietic transformation. Our current and previous findings represent a paradigm for tyrosine phosphorylation-dependent regulation of serine-threonine kinases.

Critical Role of Dok-1 and Dok-2 in Leukemia Suppression.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2951-2951
Author(s):  
Masaru Niki ◽  
Antonio Di Cristofano ◽  
Mingming Zhao ◽  
Hisamaru Hirai ◽  
Linda Van Aelst ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Tyrosine Kinases ◽  
Protein Tyrosine Kinase ◽  
Cellular Proliferation ◽  
Critical Role ◽  
Myelogenous Leukemia ◽  
Kinase Activation ◽  
Abl Tyrosine Kinase ◽  
Long Latency

Abstract Chronic myelogenous leukemia (CML) is characterized by the presence of the chimeric p210bcr/abl oncoprotein which shows elevated and constitutive protein tyrosine kinase activity relative to the normal c-abl tyrosine kinase. While several p210bcr/abl substrates have been identified, their relevance in the pathogenesis of the disease is unclear. We have identified a family of proteins, Dok (downstream of tyrosine kinase), coexpressed in hematopoietic progenitor cells. Members of this family such as p62dok (Dok-1) and p56dok-2 (Dok-2) associate with the p120 rasGTPase-activating protein (rasGAP) upon phosphorylation by p210bcr/abl as well as receptor and non-receptor tyrosine kinases. Here we report the generation and characterization of single and double Dok-2 or Dok-1/Dok-2 KO mutants. Single KO mice displayed normal steady state hematopoiesis. By contrast, concomitant Dok-1 and Dok-2 inactivation resulted in aberrant hemopoiesis and Ras/MAP kinase activation. Strikingly, all Dok-1/Dok-2 double KO mutants spontaneously developed transplantable CML-like leukemia due to increased cellular proliferation and reduced apoptosis. Furthermore, Dok-1 or Dok-2 inactivation markedly accelerated leukemia and blastic crisis onset in Tec-p210bcr/abl transgenic mice known to develop, after long latency, a myeloproliferative disorder resembling human CML. These findings unravel the critical and unexpected role of Dok-1 and 2 in tumor suppression and control of the hematopoietic compartment homeostasis.

scholarly journals Role of Protein Tyrosine Kinase p53/56lyn in Diminished Lipopolysaccharide Priming of Formylmethionylleucyl- phenylalanine-Induced Superoxide Production in Human Newborn Neutrophils

2004 ◽  
Vol 72 (11) ◽  
pp. 6455-6462 ◽  
Author(s):  
Sen Rong Yan ◽  
David M. Byers ◽  
Robert Bortolussi
Keyword(s):  
Tyrosine Kinase ◽  
Tyrosine Kinases ◽  
Protein Tyrosine Kinase ◽  
Kinase Inhibitor ◽  
Critical Role ◽  
Polymorphonuclear Neutrophils ◽  
Protein Tyrosine Kinases ◽  
Protein Tyrosine ◽  
Adult Cells

ABSTRACT Human newborns are more susceptible than adults to bacterial infection. With gram-negative bacteria, this may be due to a diminished response of newborn leukocytes to lipopolysaccharide (LPS). Since protein tyrosine kinase inhibition abolishes LPS priming in adult cells, we hypothesized that protein tyrosine kinases may have a critical role in LPS priming of polymorphonuclear neutrophils (PMNs) and that newborn PMNs may have altered protein tyrosine kinase activities. In the present study, we investigated the role of src family protein tyrosine kinases in the LPS response of newborn PMNs compared to adult cells. In a respiratory assay, the LPS-primed increase in formylmethionylleucylphenylalanine (fMLP)-triggered O2 − release by adult PMNs was greatly decreased by PP1 [4-amino-5-(4-methyphenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine], a src kinase inhibitor, to the level of untreated newborn PMNs, in which LPS failed to prime. LPS activated the src-like kinases p59hck (HCK) and p58fgr (FGR) in both adult and newborn PMNs but increased the activation of p53/56 lyn (LYN) only in adult cells. In newborn PMNs, LYN was highly phosphorylated independent of LPS. We evaluated subcellular fractions of PMNs and found that the phosphorylated form of LYN was mainly in the Triton-extractable, cytosolic fraction in adult PMNs, while in newborn cells it was located mainly in Triton-insoluble, granule- and membrane-associated fractions. In contrast, the phosphorylated mitogen-activated protein kinases ERK1/2 and p38 were mainly detected in the cytosol in both adult and newborn PMNs. These data indicate a role for LYN in the regulation of LPS priming. The trapping of phosphorylated LYN in the membrane-granule fraction in newborn PMNs may contribute to the deficiency of newborn cells in responding to LPS stimulation.

scholarly journals A role of tyrosine phosphorylation in the formation of acetylcholine receptor clusters induced by electric fields in cultured Xenopus muscle cells.

1993 ◽  
Vol 120 (1) ◽  
pp. 197-204 ◽  
Author(s):  
H B Peng ◽  
L P Baker ◽  
Z Dai
Keyword(s):  
Tyrosine Kinase ◽  
Tyrosine Phosphorylation ◽  
Electric Fields ◽  
Tyrosine Kinases ◽  
Kinase Inhibitor ◽  
Muscle Cells ◽  
Postsynaptic Membrane ◽  
Achr Clustering

During the development of the neuromuscular junction, acetylcholine receptors (AChRs) become clustered in the postsynaptic membrane in response to innervation. In vitro, several non-neuronal stimuli can also induce the formation of AChR clusters. DC electric field (E field) is one of them. When cultured Xenopus muscle cells are exposed to an E field of 5-10 V/cm, AChRs become clustered along the cathode-facing edge of the cells within 2 h. Recent studies have suggested the involvement of tyrosine kinase activation in the action of several AChR clustering stimuli, including nerve, polymer beads, and agrin. We thus examined the role of tyrosine phosphorylation in E field-induced AChR clustering. An antibody against phosphotyrosine (PY) was used to examine the localization of PY-containing proteins in E field-treated muscle cells. We found that anti-PY staining was colocalized with AChR clusters along the cathodal edge of the cells. In fact, cathodal PY staining could be detected before the first appearance of AChR clusters. When cultures were subjected to E fields in the presence of a tyrosine kinase inhibitor, tyrphostin RG-50864, cathodal AChR clustering was abolished with a half maximal inhibitory dosage of 50 microM. An inactive form of tyrphostin (RG-50862) had no effect on the field-induced clustering. These data suggest that the activation of tyrosine kinases is an essential step in E field-induced AChR clustering. Thus, the actions of several disparate stimuli for AChR clustering seem to converge to a common signal transduction mechanism based on tyrosine phosphorylation at the molecular level.

Protein-tyrosine Kinase p72 syk Is Activated by Platelet Activating Factor in Platelets

1994 ◽  
Vol 72 (06) ◽  
pp. 937-941 ◽  
Author(s):  
Karim Rezaul ◽  
Shigeru Yanagi ◽  
Kiyonao Sada ◽  
Takanobu Taniguchi ◽  
Hirohei Yamamura
Keyword(s):  
Tyrosine Kinase ◽  
Tyrosine Phosphorylation ◽  
Protein Tyrosine Kinase ◽  
Kinase Inhibitor ◽  
Critical Role ◽  
Platelet Activating Factor ◽  
Kinase Assay ◽  
Potential Candidate ◽  
Protein Tyrosine ◽  
Induced Aggregation

SummaryIt has been demonstrated that activation of platelets by platelet-activating factor (PAF) results in a dramatic increase in tyrosine phosphorylation of several cellular proteins. We report here that p72 syk is a potential candidate for the protein-tyrosine phosphorylation following PAF stimulation in porcine platelets. Immunoprecipitation kinase assay revealed that PAF stimulation resulted in a rapid activation of p72 syk which peaked at 10 s. The level of activation was found to be dose dependent and could be completely inhibited by the PAF receptor antagonist, CV3988. Phosphorylation at the tyrosine residues of p72 syk coincided with activation of yllsyk. Pretreatment of platelets with aspirin and apyrase did not affect PAF induced activation of p72 syk .Furthermore, genistein, a potent protein-tyrosine-kinase inhibitor, diminished PAF-induced p72 syk activation and Ca2+ mobilization as well as platelet aggregation. These results suggest that p72 syk may play a critical role in PAF-induced aggregation, possibly through regulation of Ca2+ mobilization.

Differential regulation of granulopoiesis by the basic helix-loop-helix transcriptional inhibitors Id1 and Id2

Blood ◽  
2005 ◽  
Vol 105 (11) ◽  
pp. 4272-4281 ◽  
Author(s):  
Miranda Buitenhuis ◽  
Hanneke W. M. van Deutekom ◽  
Liesbeth P. Verhagen ◽  
Anders Castor ◽  
Sten Eirik W. Jacobsen ◽  
...  
Keyword(s):  
Critical Role ◽  
Ectopic Expression ◽  
Constitutive Expression ◽  
Retroviral Transduction ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix ◽  
Cd34 Cells ◽  
Final Maturation ◽  
Inhibitor Of Dna Binding

Abstract Inhibitor of DNA binding (Id) proteins function as inhibitors of members of the basic helix-loop-helix family of transcription factors and have been demonstrated to play an important role in regulating lymphopoiesis. However, the role of these proteins in regulation of myelopoiesis is currently unclear. In this study, we have investigated the role of Id1 and Id2 in the regulation of granulopoiesis. Id1 expression was initially up-regulated during early granulopoiesis, which was then followed by a decrease in expression during final maturation. In contrast, Id2 expression was up-regulated in terminally differentiated granulocytes. In order to determine whether Id expression plays a critical role in regulating granulopoiesis, Id1 and Id2 were ectopically expressed in CD34+ cells by retroviral transduction. Our experiments demonstrate that constitutive expression of Id1 inhibits eosinophil development, whereas in contrast neutrophil differentiation was modestly enhanced. Constitutive Id2 expression accelerates final maturation of both eosinophils and neutrophils, whereas inhibition of Id2 expression blocks differentiation of both lineages. Transplantation of β2-microglobulin-/- nonobese diabetic severe combined immunodeficient (NOD/SCID) mice with CD34+ cells ectopically expressing Id1 resulted in enhanced neutrophil development, whereas ectopic expression of Id2 induced both eosinophil and neutrophil development. These data demonstrate that both Id1 and Id2 play a critical, although differential role in granulopoiesis.

scholarly journals Activating mutations in BRAF disrupt the hypothalamo-pituitary axis leading to hypopituitarism in mice and humans

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Angelica Gualtieri ◽  
Nikolina Kyprianou ◽  
Louise C. Gregory ◽  
Maria Lillina Vignola ◽  
James G. Nicholson ◽  
...  
Keyword(s):  
Mapk Pathway ◽  
Critical Role ◽  
Cell Lineage ◽  
Activating Mutations ◽  
Pathway Activation ◽  
Tumour Formation ◽  
Conditional Expression ◽  
Cell Cycle Inhibitors ◽  
Mutation Braf

AbstractGermline mutations in BRAF and other components of the MAPK pathway are associated with the congenital syndromes collectively known as RASopathies. Here, we report the association of Septo-Optic Dysplasia (SOD) including hypopituitarism and Cardio-Facio-Cutaneous (CFC) syndrome in patients harbouring mutations in BRAF. Phosphoproteomic analyses demonstrate that these genetic variants are gain-of-function mutations leading to activation of the MAPK pathway. Activation of the MAPK pathway by conditional expression of the BrafV600E/+ allele, or the knock-in BrafQ241R/+ allele (corresponding to the most frequent human CFC-causing mutation, BRAF p.Q257R), leads to abnormal cell lineage determination and terminal differentiation of hormone-producing cells, causing hypopituitarism. Expression of the BrafV600E/+ allele in embryonic pituitary progenitors leads to an increased expression of cell cycle inhibitors, cell growth arrest and apoptosis, but not tumour formation. Our findings show a critical role of BRAF in hypothalamo-pituitary-axis development both in mouse and human and implicate mutations found in RASopathies as a cause of endocrine deficiencies in humans.

IL-36R ligands are potent regulators of dendritic and T cells

Blood ◽  
2011 ◽  
Vol 118 (22) ◽  
pp. 5813-5823 ◽  
Author(s):  
Solenne Vigne ◽  
Gaby Palmer ◽  
Céline Lamacchia ◽  
Praxedis Martin ◽  
Dominique Talabot-Ayer ◽  
...  
Keyword(s):  
T Cells ◽  
Mhc Class Ii ◽  
Critical Role ◽  
T Helper ◽  
Innate And Adaptive Immunity ◽  
Murine Bone Marrow ◽  
Intracellular Signals ◽  
Tnf Α ◽  
Ifn Γ

Abstract IL-36α (IL-1F6), IL-36β (IL-1F8), and IL-36γ (IL-1F9) are members of the IL-1 family of cytokines. These cytokines bind to IL-36R (IL-1Rrp2) and IL-1RAcP, activating similar intracellular signals as IL-1, whereas IL-36Ra (IL-1F5) acts as an IL-36R antagonist (IL-36Ra). In this study, we show that both murine bone marrow-derived dendritic cells (BMDCs) and CD4+ T lymphocytes constitutively express IL-36R and respond to IL-36α, IL-36β, and IL-36γ. IL-36 induced the production of proinflammatory cytokines, including IL-12, IL-1β, IL-6, TNF-α, and IL-23 by BMDCs with a more potent stimulatory effect than that of other IL-1 cytokines. In addition, IL-36β enhanced the expression of CD80, CD86, and MHC class II by BMDCs. IL-36 also induced the production of IFN-γ, IL-4, and IL-17 by CD4+ T cells and cultured splenocytes. These stimulatory effects were antagonized by IL-36Ra when used in 100- to 1000-fold molar excess. The immunization of mice with IL-36β significantly and specifically promoted Th1 responses. Our data thus indicate a critical role of IL-36R ligands in the interface between innate and adaptive immunity, leading to the stimulation of T helper responses.

scholarly journals 436. Investigation of the role of SRC in capacitation associated tyrosine phosphorylation of human spermatozoa

2008 ◽  
Vol 20 (9) ◽  
pp. 116
Author(s):  
L. A. Mitchell ◽  
B. Nixon ◽  
M. A. Baker ◽  
R. J. Aitken
Keyword(s):  
Tyrosine Kinase ◽  
Tyrosine Phosphorylation ◽  
Signal Transduction Pathway ◽  
Human Sperm ◽  
Complex Signal ◽  
Phosphorylation Cascade ◽  
A Site ◽  
Signalling Cascade ◽  
Mammalian Spermatozoa

Capacitation is a pre-requisite for mammalian spermatozoa allowing them to gain the ability to fertilise an oocyte. A fundamental part of this mechanism is a dramatic increase in tyrosine phosphorylation. Implicated in this process in the mouse is a unique cAMP/PKA-mediated pathway involving a PKA-activated tyrosine kinase suggested to be pp60c-src (SRC). The Src kinases examined were predominantly expressed in the human sperm tail, a site compatible with a role in mediating the capacitation-associated tyrosine phosphorylation cascade. Co-immunoprecipitation revealed that PKA-c could be isolated from sperm and this interaction was restricted to capacitated cells, suggesting PKA-mediated activation of SRC forms an integral part of the signalling cascade assembled during capacitation. Upon activation, SRC undergoes autophosphorylation of Y416 and thus phosphorylation of this residue indicates the presence of active SRC kinase. The phosphorylation status of SRC was compared using both 2D-immunoblotting and immunocytochemical studies, both revealing a significant increase in SRC activation during capacitation. Furthermore, suppression of PKA and SRC through application of SU6656, or H89, a PKA inhibitor, led to a dramatic decrease in tyrosine phosphorylation and SRC activity. In conclusion, this study has provided evidence for the involvement of non-receptor tyrosine kinase, SRC, in regulating tyrosine phosphorylation associated with capacitation. Inhibition of SRC did not completely suppress tyrosine phosphorylation suggesting this complex signal transduction pathway exhibits a degree of functional redundancy.

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