scholarly journals The interaction between the pleckstrin homology domain of ceramide kinase and phosphatidylinositol 4,5-bisphosphate regulates the plasma membrane targeting and ceramide 1-phosphate levels

2006 ◽  
Vol 342 (2) ◽  
pp. 611-617 ◽  
Author(s):  
Tack-Joong Kim ◽  
Susumu Mitsutake ◽  
Yasuyuki Igarashi
Keyword(s):  
Plasma Membrane ◽  
Pleckstrin Homology Domain ◽  
Membrane Targeting ◽  
Pleckstrin Homology ◽  
Ceramide Kinase ◽  
Ceramide 1

scholarly journals A critical β6–β7 loop in the pleckstrin homology domain of ceramide kinase

2006 ◽  
Vol 400 (2) ◽  
pp. 255-265 ◽  
Author(s):  
Philipp Rovina ◽  
Markus Jaritz ◽  
Siegfried Höfinger ◽  
Christine Graf ◽  
Piroska Dévay ◽  
...  
Keyword(s):  
Amino Acid ◽  
Pleckstrin Homology Domain ◽  
Membrane Association ◽  
Amino Acid Residues ◽  
Ph Domain ◽  
Pleckstrin Homology ◽  
Ceramide Kinase ◽  
Mutagenesis Study ◽  
Ceramide 1 ◽  
Positively Charged

CerK (ceramide kinase) produces ceramide 1-phosphate, a sphingophospholipid with recognized signalling properties. It localizes to the Golgi complex and fractionates essentially between detergent-soluble and -insoluble fractions; however, the determinants are unknown. Here, we made a detailed mutagenesis study of the N-terminal PH domain (pleckstrin homology domain) of CerK, based on modelling, and identified key positively charged amino acid residues within an unusual motif in the loop interconnecting β-strands 6 and 7. These residues are critical for CerK membrane association and polyphosphoinositide binding and activity. Their mutagenesis results in increased thermolability, sensitivity to proteolysis, reduced apparent molecular mass as well as propensity of the recombinant mutant protein to aggregate, indicating that this loop impacts the overall conformation of the CerK protein. This is in contrast with most PH domains whose function strongly relies on charges located in the β1–β2 loop.

scholarly journals Role for the Pleckstrin Homology Domain-Containing Protein CKIP-1 in Phosphatidylinositol 3-Kinase-Regulated Muscle Differentiation

2004 ◽  
Vol 24 (3) ◽  
pp. 1245-1255 ◽  
Author(s):  
Alexias Safi ◽  
Marie Vandromme ◽  
Sabine Caussanel ◽  
Laure Valdacci ◽  
Dominique Baas ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Active Form ◽  
Muscle Differentiation ◽  
C2c12 Cells ◽  
Pleckstrin Homology Domain ◽  
Dependent Manner ◽  
Ph Domain ◽  
Pleckstrin Homology ◽  
Phosphatidylinositol 3 Kinase ◽  
Phosphatidylinositol 3

ABSTRACT In this work, we report the implication of the pleckstrin homology (PH) domain-containing protein CKIP-1 in phosphatidylinositol 3-kinase (PI3-K)-regulated muscle differentiation. CKIP-1 is upregulated during muscle differentiation in C2C12 cells. We show that CKIP-1 binds to phosphatidylinositol 3-phosphate through its PH domain and localizes to the plasma membrane in a PI3-K-dependent manner. Activation of PI3-K by insulin or expression of an active form of PI3-K p110 induces a rapid translocation of CKIP-1 to the plasma membrane. Conversely, expression of the 3-phosphoinositide phosphatase myotubularin or PI3-K inhibition by LY294002, wortmannin, or mutant p85 abolishes CKIP-1 binding to the membrane. Upon induction of differentiation in low-serum medium, CKIP-1 overexpression in C2C12 myoblasts first promotes proliferation and then stimulates the expression of myogenin and cell fusion in a manner reminiscent of the dual positive effect of insulin-like growth factors on muscle cells. Interference with the PI3-K pathway impedes the effect of CKIP-1 on C2C12 cell differentiation. Finally, silencing of CKIP-1 by RNA interference abolishes proliferation and delays myogenin expression. Altogether, these data strongly implicate CKIP-1 as a new component of PI3-K signaling in muscle differentiation.

scholarly journals Membrane Targeting of Grb2-associated Binder-1 (Gab1) Scaffolding Protein through Src Myristoylation Sequence Substitutes for Gab1 Pleckstrin Homology Domain and Switches an Epidermal Growth Factor Response to an Invasive Morphogenic Program

2003 ◽  
Vol 14 (4) ◽  
pp. 1691-1708 ◽  
Author(s):  
Christiane R. Maroun ◽  
Monica A. Naujokas ◽  
Morag Park
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Epithelial Cells ◽  
Subcellular Localization ◽  
Epithelial Morphogenesis ◽  
Pleckstrin Homology Domain ◽  
Membrane Targeting ◽  
Ph Domain ◽  
Pleckstrin Homology ◽  
Epidermal Growth

The hepatocyte growth factor receptor tyrosine kinase Met promotes cell dissociation and the inherent morphogenic program of epithelial cells. In a search for substrates downstream from Met, we have previously identified the Grb2-associated binder-1 (Gab1) as critical for the morphogenic program. Gab1 is a scaffold protein that acts to diversify the signal downstream from the Met receptor through its ability to couple with multiple signal transduction pathways. Gab1 contains a pleckstrin homology (PH) domain with specificity for phosphatidylinositol 3,4,5-trisphosphate. The phospholipid binding capacity of the Gab1 PH domain is required for the localization of Gab1 at sites of cell-cell contact in colonies of epithelial cells and for epithelial morphogenesis, suggesting that PH domain-dependent subcellular localization of Gab1 is a prerequisite for function. We have investigated the requirement for membrane localization of Gab1 for biological activity. We show that substitution of the Gab1 PH domain with the myristoylation signal from the c-Src protein is sufficient to replace the Gab1 PH domain for epithelial morphogenesis. The membrane targeting of Gab1 enhances Rac activity in the absence of stimulation and switches a nonmorphogenic noninvasive response to epidermal growth factor to a morphogenic invasive program. These results suggest that the subcellular localization of Gab1 is a critical determinant for epithelial morphogenesis and invasiveness.

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