Article

Regulation of EGF-induced phospholipase C-gamma1 translocation and activation by its SH2 and PH domains.

Department of Cell Biology, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, Alberta T6G 2H7, Canada.
Traffic (Impact Factor: 4.71). 10/2003; 4(9):618-30. DOI: 10.1034/j.1600-0854.2003.00121.x
Source: PubMed

ABSTRACT Translocation of phospholipase C-gamma1 is essential for its function in response to growth factors. However, in spite of recent progress, the phospholipase C-gamma1 translocation pattern and the molecular mechanism of the translocation are far from fully understood. Contradictory results were reported as to which domain, PH or SH2, controls the epidermal growth factor-induced translocation of phospholipase C-gamma1. In this communication, we studied epidermal growth factor-induced translocation of phospholipase C-gamma1 by using comprehensive approaches including biochemistry, indirect fluorescence and live fluorescence imaging. We provided original evidence demonstrating that: (i) endogenous phospholipase C-gamma1, similar to YFP-tagged phospholipase C-gamma1, translocated to endosomes following its initial translocation from cytosol to the plasma membrane in response to epidermal growth factor; (ii) phospholipase C-gamma1 remained phosphorylated in endosomes, but phospholipase C-gamma1 activity is not required for its translocation, which suggests a signaling role for phospholipase C-gamma1 in endosomes; (iii) the PH domain was not required for the initial translocation of phospholipase C-gamma1 from cytosol to the plasma membrane, but it stabilizes phospholipase C-gamma1 in the membrane at a later time; (iv) the function of the phospholipase C-gamma1 PH domain in stabilizing phospholipase C-gamma1 membrane association is very important in maintaining the activity of phospholipase C-gamma1; and (v) the role of the PH domain in phospholipase C-gamma1 membrane association and activation is dependent on PI3K activity. We conclude that the phospholipase C-gamma1 SH2 and PH domains coordinate to determine epidermal growth factor-induced translocation and activation of phospholipase C-gamma1.

0 Followers
 · 
73 Views
  • [Show abstract] [Hide abstract]
    ABSTRACT: It is well established that epidermal growth factor (EGF) induces the cytoskeleton reorganization and cell migration through two major signaling cascades: phospholipase C-gamma1 (PLC-gamma1) and Rho GTPases. However, little is known about the cross talk between PLC-gamma1 and Rho GTPases. Here we showed that PLC-gamma1 forms a complex with Rac1 in response to EGF. This interaction is direct and mediated by PLC-gamma1 Src homology 3 (SH3) domain and Rac1 (106)PNTP(109) motif. This interaction is critical for EGF-induced Rac1 activation in vivo, and PLC-gamma1 SH3 domain is actually a potent and specific Rac1 guanine nucleotide exchange factor in vitro. We have also demonstrated that the interaction between PLC-gamma1 SH3 domain and Rac1 play a significant role in EGF-induced F-actin formation and cell migration. We conclude that PLC-gamma1 and Rac1 coregulate EGF-induced cell cytoskeleton remodeling and cell migration by a direct functional interaction.
    Molecular Endocrinology 04/2009; 23(6):901-13. DOI:10.1210/me.2008-0368 · 4.20 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: ABSTRACT Knock-down of Grb2 by RNA interference (RNAi) strongly inhibitsclathrin-mediat ed endocytosis of the EGF receptor (EGFR). To gaininsights into the function of Grb2 in EGFR endocytosis, we have generated cell lines inwhich endogenous Grb2 was replaced by YFP- tagged Grb2 expressed at the physiological level. In these cellsGrb2-YFP fully reversed the inhibitory effect of Grb2 knock-down on EGFR endocytosis and moreover, trafficked together with EGFR during endocytosis. Overexpression of Grb2-binding protein, c-Cbl did not restore endocytosis in Grb2-depletedcel ls. However, EGFR endocytosis was rescued in Grb2-depleted cells by chimeric proteins consisting of the Src Homology (SH) 2 domain of Grb2 fused to c- Cbl. The “knock-down and rescue” analysis revealed that the expression of Cbl-Grb2/SH2 fusions containing RING finger domain,of Cbl restores normal ubiquitylation and internalization
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Although endocrine therapy has dramatically improved the treatment of breast cancer therapeutic resistance and tumour recurrence occurs, even in estrogen receptor (ER) positive cases. Identifying and understanding the molecular mechanisms which underpin endocrine resistance is therefore important if future therapeutic strategies are to be developed. Members of the fibroblast growth factor (FGF) and fibroblast growth factor receptor (FGFR) families have been implicated in breast cancer development and progression. Our results demonstrate that culture of michigan cancer foundation - 1 (MCF)7 cells with FGF1 results in reduced sensitivity to tamoxifen in vitro. Furthermore, our tissue microarray expression data demonstrates that FGFR3 expression is increased in tamoxifen resistant breast tumours. To confirm that activation of FGFR3 reduced sensitivity to tamoxifen we used an inducible activation system and a constitutively active mutant of FGFR3 expressed in MCF7 cells. Activation of FGFR3 reduced sensitivity to tamoxifen and Fulvestrant but did not lead to phosphorylation of ER demonstrating that FGFR3 does not feedback to modulate ER activity. FGFR3 activation in MCF7 cells stimulated activation of the mitogen-activated protein kinase (MAPK) and phosphoinositide 3-kinase (PI3K) signalling pathways, both of which have been implicated in tamoxifen resistance in breast cancer. Furthermore, our data indicates that activation of phospholipase C gamma is a key-signalling event regulating MAPK and PI3K activation and that its activation reduces sensitivity to tamoxifen. Therefore, we hypothesise that FGFRs could play an integral part, not only in breast cancer development but also in resistance to endocrine-therapy.
    International Journal of Cancer 06/2012; 130(12):2857-66. DOI:10.1002/ijc.26304 · 5.01 Impact Factor

Preview

Download
0 Downloads