title: Differential interaction of Cbl with Grb2 and CrkL in CD2-mediated NK cell activation.
abstract: Natural killer (NK) cells participate in both innate and adoptive immunity by their prompt secretion of cytokines and by their ability to lyse virally infected cells or tumor cells. CD2 is surface glycoprotein receptors and crucial for NK cell activation. However, molecular events involved in CD2-mediated NK cell activation have not been fully elucidated. Cbl-Grb2 and Cbl-CrkL interactions have been implicated in T cell and B cell receptor, and cytokine receptor signaling. Here we analyzed tyrosine phosphorylation and interactions of Cbl with adapter proteins, Grb2 and CrkL, in NK3.3 cells. CD2 crosslinking results in the marked tyrosine phosphorylation of Cbl in an antibody concentration- and time-dependent manner. Immunodepletion studies reveal that Grb2-associated tyrosine phosphorylated p120 kDa protein is Cbl. In vitro binding studies using GST-fusion proteins demonstrate that Cbl constitutively associates with the SH3 domains of Grb2, with a preference for the amino-terminal domain. In addition, we demonstrate that CrkL associates with a large portion of tyrosine phosphorylated Cbl after CD2 stimulation of NK3.3 cells. In contrast to constitutive Cbl association with Grb2, tyrosine phosphorylated Cbl interacts with CrkL via its SH2 domain only after CD2 stimulation. Although the precise roles of interactions of Cbl with Grb2 and CrkL in NK cell activation remains to be elucidated, their tyrosine phosphorylation, in addition to the multiple protein interactions described here, strongly suggest that interactions of Cbl with Grb2 and CrkL may play pivotal roles in CD2-mediated NK cell activation.
authors.affiliation: Department of Internal Medicine, Osaka Dental University, 8-1 Kuzuha Hanazono-cho, Hirakata-shi, 573-1121, Osaka, Japan.
authors.forename: J Y
authors.initials: JY
authors.lastname: Huang
authors.forename: H
authors.initials: H
authors.lastname: Umehara
authors.forename: H
authors.initials: H
authors.lastname: Inoue
authors.forename: F H
authors.initials: FH
authors.lastname: Tabassam
authors.forename: T
authors.initials: T
authors.lastname: Okazaki
authors.forename: T
authors.initials: T
authors.lastname: Kono
authors.forename: Y
authors.initials: Y
authors.lastname: Minami
authors.forename: Y
authors.initials: Y
authors.lastname: Tanaka
authors.forename: N
authors.initials: N
authors.lastname: Domae
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D018801:CD2 Antigens / Q000276:immunology*; D002352:Carrier Proteins / Q000276:immunology; D002460:Cell Line; D006801:Humans; D007694:Killer Cells, Natural / Q000276:immunology*; D008213:Lymphocyte Activation / Q000276:immunology; D009687:Nuclear Proteins / Q000276:immunology; D050722:Oncogene Protein v-cbl; D010766:Phosphorylation; D015682:Retroviridae Proteins, Oncogenic / Q000276:immunology; D015398:Signal Transduction / Q000276:immunology*; D014443:Tyrosine / Q000276:immunology
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D018801:CD2 Antigens; C082869:CRKL protein; D002352:Carrier Proteins; C409587:GRAP2 protein, human; D009687:Nuclear Proteins; D050722:Oncogene Protein v-cbl; D015682:Retroviridae Proteins, Oncogenic; D014443:Tyrosine