Satterthwaite, Anne B. and Willis, Fiona and Kanchanastit, Prim and Fruman, David and Cantley, Lewis C. and Helgason, Cheryl D. and Humphries, R. Keith and Lowell, Clifford A. and Simon, Melvin and Leitges, Michael and Tarakhovsky, Alexander and Tedder, Thomas F. and Lesche, Ralf and Wu, Hong and Witte, Owen N. (2000) A sensitized genetic system for the analysis of murine B lymphocyte signal transduction pathways dependent on Bruton's tyrosine kinase. Proceedings of the National Academy of Sciences of the United States of America, 97 (12). pp. 6687-6692. ISSN 0027-8424 http://resolver.caltech.edu/CaltechAUTHORS:SATpnas00
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Modifier screens have been powerful genetic tools to define signaling pathways in lower organisms. The identification of modifier loci in mice has begun to allow a similar dissection of mammalian signaling pathways. Transgenic mice (Btklo) expressing 25% of endogenous levels of Bruton's tyrosine kinase (Btk) have B cell functional responses between those of wild-type and Btk-/- mice. We asked whether reduced dosage or complete deficiency of genes previously implicated as Btk regulators would modify the Btklo phenotype. We used two independent assays of Btk-dependent B cell function. Proliferative response to B cell antigen receptor cross-linking in vitro was chosen as an example of a relatively simple, well-defined signaling system. In vivo response to type II T-independent antigens (TI-II) measures complex interactions among multiple cell types over time and may identify additional Btk pathways. All modifiers identified differentially affected these two assays, indicating that Btk mediates these processes via distinct mechanisms. Loss of Lyn, PTEN (phosphatase and tensin homolog), or SH2-containing inositol phosphatase suppressed the Btklo phenotype in vitro but not in vivo, whereas CD19 and the p85alpha form of phosphoinositide 3-kinase behaved as Btklo enhancers in vivo but not in vitro. Effects of Lyn, PTEN, or p85alpha haploinsufficiency were observed. Haploinsufficiency or complete deficiency of protein kinase C beta , Fyn, CD22, Galpha q, or Galpha 11 had no detectable effect on the function of Btklo B cells. A transgenic system creating a reduction in dosage of Btk can therefore be used to identify modifier loci that affect B cell responses and quantitatively rank their contribution to Btk-mediated processes.
|Additional Information:||Copyright © 2000 by the National Academy of Sciences Contributed by Owen N. Witte, April 3, 2000. Published online before print May 30, 2000, 10.1073/pnas.110146697 We thank Dr. Roger Perlmutter for providing fyn2/2 mice, Drs. Larry Zipursky and Naomi Rosenberg for critical reading of the manuscript and helpful suggestions, and Jamie White for assistance in the preparation of this manuscript. O.N.W. is an Investigator of the Howard Hughes Medical Institute. A.B.S. and D.F. are Special Fellows of the Leukemia and Lymphoma Society of America. This work was partially supported by National Institutes of Health Grants CA-81776 and CA-54464 (to T.F.T.), DK50267 (to C.A.L.), and GM41890 (to L.C.C.), and funds from the National Cancer Institute of Canada (to R.K.H. and C.D.H.). The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. §1734 solely to indicate this fact.|
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|Deposited On:||11 Oct 2005|
|Last Modified:||26 Dec 2012 08:41|
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