Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.10.2 (focal adhesion kinase)
44,029 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The binding of CD40 ligand on activated T cells to CD40 on resting B cells induces the expression of costimulatory molecules B7-1 (CD80) and B7-2 (CD86). The induction of B7 molecules by CD40 ligand-CD40 interaction represents a critical step in rendering B cells competent for antigen presentation. The CBA/N mouse has a defect in CD40 signalling which has been attributed to a mutation in Bruton's tyrosine kinase. We have compared the ability of murine CD40 ligand to induce B7-1 and B7-2 expression on B cells isolated from CBA/N and wild-type CBA/J mice. We find that the CBA/N defect partially impairs both B7-1 and B7-2 induction via CD40. Subsequent experiments investigated the roles of different second messenger systems in B7-1 and B7-2 induction in normal B cells. In M12 B lymphomas either CD40 cross-linking or cAMP treatment can induce B7 molecules. Here we report that treatment with dibutyryl-cAMP also induces B7 molecules in normal B cells provided that they have been preactivated by CD40 cross-linking. We also find that PMA and ionomycin treatment of B cells induces B7-2 but not B7-1 expression. Our data therefore show roles for BTK, cAMP and PMA/ionomycin in B7 induction, as well as providing further evidence for differential regulation of B7-1 and B7-2 induction in B cells.
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PMID:Induction of costimulatory molecules B7-1 and B7-2 in murine B cells. the CBA/N mouse reveals a role for Bruton's tyrosine kinase in CD40-mediated B7 induction. 870 Jan 70

Gap1(IP4BP), one of a member of Ras GTPase-activating proteins, has been identified as a specific inositol 1,3,4,5-tetrakisphosphate (IP4)-binding protein (Cullen, P. J., Hsuan, J. J., Truong, O., Letcher, A. J., Jackson, T. R., Dawson, A. P., and Irvine, R. F. (1995) Nature 386, 527-530). In this paper we describe Gap1(m), which is closely related to Gap1(IP4BP), to also be an IP4-binding protein and show that the pleckstrin homology domain (PH) is the central IP4-binding domain by expressing fragments of the mouse Gap1(m) in Escherichia coli as fusion proteins and examining their activities. However, in addition to the PH domain, an adjacent GAP-related domain and carboxyl terminus are required for high affinity specific IP4 binding. The PH domain is highly conserved in the Gap1 family and also has striking homology to the amino-terminal region of Bruton's tyrosine kinase. Substitution of Cys for Arg at position 628 in the PH domain corresponding to the mutation of Bruton's tyrosine kinase observed in X-linked immunodeficiency mice results in a dramatic reduction of IP4 binding activity as well as phospholipid binding capacity of Gap1(m). This mutant also showed the GAP activity against Ha-Ras to be similar to that of the wild type Gap1(m). Our results suggest that the PH domain of Gap1(m) functions as a modulatory domain of GAP activity by binding IP4 and phospholipids.
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PMID:Structure-function relationships of the mouse Gap1m. Determination of the inositol 1,3,4,5-tetrakisphosphate-binding domain. 870 43

X-linked agammaglobulinemia (XLA) is an inherited immunodeficiency disease with a block in differentiation from pre-B to B cells resulting in a selective defect in the humoral immune response. Affected males have very low concentrations of serum immunoglobulins leading predominantly to recurrent bacterial infections beginning at age 6 to 18 months. The gene responsible for XLA was identified recently to encode a cytoplasmatic tyrosine kinase (Bruton's tyrosine kinase, BTK). We have analyzed the BTK gene in a large family in which two brothers presented with the severe phenotype of XLA. Genomic DNA of affected boys and from healthy relatives was amplified by PCR with primers specific for the putative promoter region and for all 19 exons, including flanking intron boundaries, and subsequently screened for mutations using single strand conformation polymorphism (SSCP) analysis. Altered single strand band patterns were found using primers specific for exon 10, 15, and 18. Direct cycle-sequencing of these BTK segments detected two known polymorphisms in intron 14 and in exon 18. Sequencing of exon 10 from two boys with XLA demonstrated a novel point mutation in the SH2 domain of BTK. Direct identification of healthy female carriers in three generations was performed by amplification mutagenesis using PCR with a modified first primer. This method can easily be applied also to prenatal diagnosis.
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PMID:Detection of a novel mutation in the SRC homology domain 2 (SH2) of Bruton's tyrosine kinase and direct female carrier evaluation in a family with X-linked agammaglobulinemia. 872 28

A point mutation in the pleckstrin homology domain of the mouse Bruton's tyrosine kinase (btk) gene results in an X-linked immune defect, Xid, characterized by immunologic unresponsiveness to polymeric carbohydrate Ags. In Xid mice, B cells specific for phosphocholine (PC) do not develop in peripheral lymphoid tissues because they either fail to be positively selected from the marrow or they are clonally deleted via an Ag-driven, receptor-mediated process. Overexpression of the bcl-2 gene allows PC-specific B cells to survive and mature in Xid mukappa anti-PC transgenic mice, but PC-specific B cells are not rescued by bcl-2 in Xid mu-only transgenic mice. The failure of bcl-2 to rescue PC-specific B cells, in mu-only transgenic mice suggests that either it does not correct the btk defect in the Ag-driven selection process that occurs in pre-B cells and/or in very immature B cells or that a btk-dependent proliferative phase is required for the selection and amplification of the PC-specific B cells in mu-only transgenic mice. The rescue of PC-specific B cells in mukappa transgenic mice indicates that bcl-2 can alter receptor-mediated B cell selection at late stages in B cell development. The rescued PC-specific B cells in Xid male mice do not exhibit an altered proliferation profile in response to B cell-stimulating agents compared with B cells from unmanipulated Xid mice; thus, they fail to respond to soluble anti-mu, or PC-dextran, but they proliferate in response to PC, anti-mu, or anti-id conjugated to Sepharose.
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PMID:bcl-2 alters the antigen-driven selection of B cells in mukappa but not in mu-only Xid transgenic mice. 875 9

Alterations of the Bruton's tyrosine kinase (Btk) gene are responsible for X-linked agammaglobulinemia (XLA). Although mutations in various regions were reported mainly in the Caucasian population, correlation between the locations of mutation and the clinical phenotypes remains unclear. We report 12 abnormalities of the Btk gene found in 12 unrelated families out of 14 XLA families in Japan and their clinical features. We utilized Southern blotting and single-strand conformation polymorphism (SSCP) analysis. Gene rearrangement in the kinase domain was identified in two patients by Southern blotting. Seven point mutations, two small deletions, and one small insertion were detected by SSCP and sequencing. The SSCP analysis also provided information about the carriers in these families. We found some clinical heterogeneity in the affected family members with the same gene mutation. Moreover, there is considerable inconsistency between the locations of gene aberrations and the immunological phenotypes. Some patients with a nonsense mutation, which may result in the lack of kinase domain, have detectable B cells and immunoglobulins. These identified alterations will provide valuable clues to the Btk protein function and the pathogenesis of XLA.
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PMID:Mutations of the Btk gene in 12 unrelated families with X-linked agammaglobulinemia in Japan. 883 36

Mutations of the Bruton's tyrosine kinase (btk) gene cause X-linked agammaglobulinemia (XLA) in humans and X-linked immune deficiency (Xid) in mice. To establish the BTK role in B-cell activation we examined the responses of wild-type and Xid B cells to stimulation through surface IgM and CD40, the transducers of thymus independent-type 2 and thymus-dependent activation, respectively. Wild-type BTK was necessary for proliferation induced by soluble anti-IgM (a prototype for thymus independent-type 2 antigen), but not for responses to soluble CD40 ligand (CD40L, the B-cell activating ligand expressed on T-helper cells). In the absence of wild-type BTK, B cells underwent apoptotic death after stimulation with anti-IgM. In the presence of wild-type but not mutated BTK, anti-IgM stimulation reduced apoptotic cell death. In contrast, CD40L increased viability of both wild-type and Xid B cells. Importantly, viability after stimulation correlated with the induced expression of bcl-XL. In fresh ex vivo small resting B cells from wild-type mice there was only barely detectable bcl-XL protein, but there was more in the larger, low-density ("activated") splenic B cells and peritoneal B cells. In vitro Bcl-XL induction following ligation of sIgM-required BTK, was cyclosporin A (CsA)-sensitive and dependent on extracellular Ca2+. CD40-mediated induction of bcl-x required neither wild-type BTK nor extracellular Ca2+ and was insensitive to CsA. These results indicate that BTK lies upstream of bcl-XL in the sIgM but not the CD40 activation pathway. bcl-XL is the first induced protein to be placed downstream of BTK.
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PMID:An essential role for Bruton's [corrected] tyrosine kinase in the regulation of B-cell apoptosis. 885 92

Bruton's tyrosine kinase (Btk) is a cytoplasmic protein kinase that is defective in X-linked agammaglobulinaemia in man and in X-linked immunodeficiency in the mouse. There is controversy regarding the stages of B cell development that are dependent on Btk function. To determine the point in B cell differentiation at which defects in Btk become apparent, we generated a mouse model by inactivating the Btk gene through an in-frame insertion of a lacZ reporter by homologous recombination in embryonic stem cells. The phenomenon of X-chromosome inactivation in Btk+/- heterozygous female mice enabled us to evaluate the competition between B cell progenitors expressing wild-type Btk and those expressing the Btk-/lacZ allele in each successive step of development. Although Btk was already expressed in pro-B cells, the first selective disadvantage only became apparent at the transition from small pre-B cells to immature B cells in the bone marrow. A second differentiation arrest was found during the maturation from IgD(low)IgM(high) to IgD(high)IgM(low) stages in the periphery. Our results show that Btk expression is essential at two distinct differentiation steps, both past the pre-B cell stage.
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PMID:Inactivation of Btk by insertion of lacZ reveals defects in B cell development only past the pre-B cell stage. 889 Jan 60

Bruton's tyrosine kinase (Btk), a cytoplasmic protein-tyrosine kinase, plays a pivotal role in B cell activation and development. Mutations in the pleckstrin homology (PH) domain of the Btk gene cause human X-linked agammaglobulinemia (XLA) and murine X-linked immunodeficiency (Xid). In this paper, we report that the PH domain of Btk functions as an inositol 1,3,4,5-tetrakisphosphate (IP4), inositol 1,3,4,5,6-pentakisphosphate, and inositol 1,2,3,4,5,6-hexakisphosphate (IP6) binding domain (Kd of approximately 40 nM for IP4), and that all of the XLA (Phe replaced by Ser at position 25 (F25S), R28H, T33P, V64F, and V113D) and Xid mutations (R28C) found in the PH domain result in a dramatic reduction of IP4 binding activity. Furthermore, the rare alternative splicing variant, with 33 amino acids deleted in the PH domain, corresponding to exon 3 of the Btk gene, also impaired IP4 binding capacity. In contrast, a gain-of-function mutant called Btk*, which carries a E41K mutation in the PH domain, binds IP6 with two times higher affinity than the wild type. Our data suggest that B cell differentiation is closely correlated with the IP4 binding capacity of the PH domain of Btk.
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PMID:Mutation of the pleckstrin homology domain of Bruton's tyrosine kinase in immunodeficiency impaired inositol 1,3,4,5-tetrakisphosphate binding capacity. 893 85

Pleckstrin homology (PH) domains may act as membrane localization modules through specific interactions with phosphoinositide phospholipids. These interactions could represent responses to second messengers, with scope for regulation by soluble inositol polyphosphates. A biosensor-based assay was used here to probe interactions between PH domains and unilamellar liposomes containing different phospholipids and to demonstrate specificity for distinct phosphoinositides. The dynamin PH domain specifically interacted with liposomes containing phosphatidylinositol-4,5-bisphosphate [PI(4,5)P2] and, more weakly, with liposomes containing phosphatidylinositol-4-phosphate [PI(4)P]. This correlates with phosphoinositide activation of the dynamin GTPase. The functional GTPase of a dynamin mutant lacking the PH domain, however, cannot be activated by PI(4,5)P2. The phosphoinositide-PH domain interaction can be abolished selectively by point mutations in the putative binding pocket predicted by molecular modelling and NMR spectroscopy. In contrast, the Bruton's tyrosine kinase (Btk)PH domain specifically bound liposomes containing phosphatidylinositol-3,4,5-trisphosphate [PI(3,4,5)P3]: an interaction requiring Arg28, a residue found to be mutated in some X-linked agammaglobulinaemia patients. A rational explanation for these different specificities is proposed through modelling of candidate binding pockets and is supported by NMR spectroscopy.
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PMID:Distinct specificity in the recognition of phosphoinositides by the pleckstrin homology domains of dynamin and Bruton's tyrosine kinase. 894 47

Bruton's tyrosine kinase (BTK) plays an important role in B cell development. Deletion of C-terminal 14 amino acids of the SH3 domain of BTK results in X-linked agammaglobulinemia (XLA), an inherited disease. We report here on the stability and folding of SH3 domain of BTK. Peptides corresponding to residues 216-273 (58 residues) and 216-259 (44 residues) of BTK SH3 domain were synthesized by solid phase methods; the first peptide constitutes the entire SH3 domain of BTK while the latter peptide lacks 14 amino acid residues of the C-terminal. The 58 amino acid peptide forms mainly a beta-barrel type folding unit. Although small and lacking disulfide bonds, this peptide is extremely stable to thermal denaturation. Based on circular dichroism measurements, its melting temperature was found to be high, 82 degrees C at pH 6.0. However, the Gibbs free energy (delta GH2O) of the intrinsic stability and thermodynamic spontaneity of unfolding were found to be low, 2.6 kcal/mol by Gdn.HCl denaturation experiments, as compared to 12 kcal/mol obtained for larger single domain proteins, indicating poor stability of SH3 domain. Addition of 500 mM of Na2SO4 increased the free energy change delta GH2O to 4.0 kcal/mol, suggesting an ionic strength effect. The truncated peptide fails to fold correctly and adopts random coil conformation in contrast to 58 amino acid beta-barrel peptide, which exhibits high thermal stability but normal or low stability at ambient temperature. These results, to our knowledge the first to delineate the importance of C-terminal in structural integrity of SH3 domains, indicate also that improper folding and/or poor stability of mutant SH3 domain in BTK likely causes XLA.
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PMID:Stability and folding of the SH3 domain of Bruton's tyrosine kinase. 899 Apr 99


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