Identification of phosphorylation sites for Bruton's tyrosine kinase within the transcriptional regulator BAP/TFII-I.
Bruton's tyrosine kinase (Btk), a member of the Tec family of cytosolic kinases, is essential for B cell development and function. BAP/TFII-I, a protein implicated in transcriptional regulation, is associated with Btk in B cells and is transiently phosphorylated on tyrosine following B cell receptor engagement. BAP/TFII-I is a substrate for Btk in vitro and is hyperphosphorylated on tyrosine upon coexpression with Btk in mammalian cells. In an effort to understand the physiologic consequences of BAP/TFII-I tyrosine phosphorylation following B cell receptor stimulation, site-directed mutagenesis and phosphopeptide mapping were used to locate the predominant sites of BAP/TFII-I phosphorylation by Btk in vitro. These residues, Tyr248, Tyr357, and Tyr462, were also found to be the major sites for Btk-dependent phosphorylation of BAP/TFII-I in vivo. Residues Tyr357 and Tyr462 are contained within the loop regions of adjacent helix-loop-helix-like repeats within BAP/TFII-I. Mutation of either Tyr248, Tyr357, or Tyr462 to phenylalanine reduced transcription from a c-fos promoter relative to wild-type BAP/TFII-I in transfected COS-7 cells, consistent with the interpretation that phosphorylation at these sites contributes to transcriptional activation. Phosphorylation of BAP/TFII-I by Btk may link engagement of receptors such as surface immunoglobulin to modulation of gene expression.
Egloff
A M
AM
Department of Molecular Biology and Genetics and Howard Hughes Medical Institute, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
Desiderio
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D000077329:Agammaglobulinaemia Tyrosine Kinase; D000595:Amino Acid Sequence; D000818:Animals; D001402:B-Lymphocytes / Q000378:metabolism; D001665:Binding Sites; D019556:COS Cells; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D000939:Epitopes / Q000737:chemistry; D004926:Escherichia coli / Q000378:metabolism; D017930:Genes, Reporter; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D010649:Phenylalanine / Q000737:chemistry; D010766:Phosphorylation; D010957:Plasmids / Q000378:metabolism; D017354:Point Mutation; D011233:Precipitin Tests; D017434:Protein Structure, Tertiary; D011505:Protein-Tyrosine Kinases / Q000737:chemistry* / Q000378:metabolism*; D016760:Proto-Oncogene Proteins c-fos / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D014157:Transcription Factors / Q000737:chemistry* / Q000378:metabolism*; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014162:Transfection; D014357:Trypsin / Q000378:metabolism; D014443:Tyrosine / Q000737:chemistry
D004268:DNA-Binding Proteins; D000939:Epitopes; D016760:Proto-Oncogene Proteins c-fos; D011994:Recombinant Proteins; D014157:Transcription Factors; D014443:Tyrosine; D010649:Phenylalanine; D011505:Protein-Tyrosine Kinases; D000077329:Agammaglobulinaemia Tyrosine Kinase; C000625949:BTK protein, human; C000625951:Btk protein, mouse; D014357:Trypsin