| Aim: to identify the specific binding of Dok-PTB and RTK, andtoexplain it's exact biologic function. The Dok proteins (downstream oftyrosine kinases) are a newly identified family of adaPtor proteinsphosphorylated by a wde range of protein tyrosine kinases (PTKs). Thefamily includes the prototyPe fAnily member, Dokl, which wasoriginally identified as a tyrosine-phosphorylated 62-kD proteinassociated with pl20RasGAP, a negative regulator of Ras. Dok1 isphosphorylated upon activation of receptor tyrOsine kinases (RTKs)(including the epidermal growth factor receptor, platelet-derived growthfactor receptor, and insulin receptor), antigen receptors (such asFcAlIB), and nonreceptor tyrosine kinases (including v-Src, v-Abl, andthe p2l 0 bcr-abl fusion oncoprotein). While the sequence of Doklsuggested that the protCin mny serve as a docking protein, recent studieshave shed light on its function as a negative regulator of cellprolitbration and the RaS/mitogen-activated protein kinase (MAPK)signaling pathway. Using Dokl-/' derived cells, such as bone marrowm 7 Nn:J1I.k7,2002 '+6R j[,l:[F}tF i2 X [^J (,i l'J4} + llekl-PTB t5t1dt&M Ris. bL{L H x)JtiL{n5'cells, thymocytes, splenic B cel1s and lnouse embryo fibroblasts (MEFs),these studies suggest that the l1egative effect of Dokl on cellproliferation is at least il1 part due to negatively intluencing theRas/MAPK pathway. As a docki11g 111olecule, Dokl harbors severalcharacteristic JnotifS al1d dolnains. At its N terminus, Dokl contains apleckstrin homology (I'H) domain (MI -P l20 ) followed by aI)hospllotyrosine-binLling (P1'B) do111ain (Nl3l-Q260 ), tvhile theCOOH-terminal tail of Dok1 harbors l0 potential tyrosinephosphorylation sitcs and 7 PxxP lnotifS, which may serve as dockingsites lbr SH3 donlaills. 1'lle PTB ilol11ain of Dokl was recently fOu11d tobc a functional PTB module al1d is involved in its association with Srcllomology 2 (SH2) domail1--containing inositol 5-phosphatase(SHIP l ).Pllosphorylation of tl1e muItiple tyrosine residues at tlle COOH terminusof Dok l have been sllown to create docking sites fOr proteins containingSll2 dOlnains, includil1g Nck, RasGAP, and Csk. Despite theidentification of the variot1s protein--protei11 interactions mentionedabove, alld tyrosine phosphorylatiol1 of Dok in response to EGFstimulation has previously been reported, the surely mechanism of thisspccific docking protcin remains ul1clear. How can Dokl affects theinteraction EGFR signal pathway in rcsponse to growth factors, and inturn influencek its biological t\lnctions. Furthermore, the structuralrclcvancc of the association of Dokl with EGFR is ul1clear. Althoughthese discrepancies l11ight be caused by differences between Dok lhmi1ylllen1bers, stilnu1i, or cell types, the specification of the EGFR/Doklcomplex requires further elucidation. As a n1olecular recognitor, a PTBdomain directs association witll an NPxY autophosphrylation site on aspecific receptor. Although aIl Dok members can associate with RTK,tlleir associated properties have many difference each other. Dokl andDok2 are likely sequenced, expressed, even function. The specificityof receptor tyrosine kinase signa1ing has been investigated in greatdetail.'A variety of Substrates are shared by several receptors. Toidentify the particular sites of association between Dokl and EGFR,we carried out GST-pull down and mini peptide inhibition studies byrecombinant GST-Doks-PTB protein. Here, we explore the potentialrole of the Dok family member Dokl in signaling pathWays mediated bytlle epidermal.growtl1 factor (EGF) receptor. PTB domain in Dok1 wascritical fOr its association witl1 two PTB-binding consensus sites on theEGF receptor. Thcsc results suggest that Dokl has an importantm 8 9{I A"/- 2U02 frjfffl'l- .'?lktfe A ftfi'i '?#-/?Dokl-PTB tf'substrate of epidermal growth factor receptors, and in several cell types Dokl mayb... |
PDF Full Text Request