Cloning and characterization of a novel adaptor protein, CIN85, that interacts with c-Cbl
ABSTRACT The c-Cbl protooncogene product is a prominent substrate of protein tyrosine kinases and is rapidly tyrosine-phosphorylated upon stimulation of a wide variety of cell-surface receptors. We have identified a novel c-Cbl-interacting protein termed CIN85 with a molecular mass of 85 kDa which shows similarity to adaptor proteins, CMS and CD2AP. CIN85 mRNA is expressed ubiquitously in normal human tissues and cancer cell lines analyzed. CIN85 was basally associated with c-Cbl. For interaction of CIN85 with c-Cbl, the second SH3 domain of CIN85 was shown to serve as a central player. The CIN85-c-Cbl association was enhanced shortly after stimulation of 293 cells with epidermal growth factor (EGF) and gradually diminished to a basal level, which correlated with a tyrosine phosphorylation level of c-Cbl. Our results suggest that CIN85 may play a specific role in the EGF receptor-mediated signaling cascade via its interaction with c-Cbl.
SourceAvailable from: Mehmet M Altintas[Show abstract] [Hide abstract]
ABSTRACT: CD2-associated protein (CD2AP) is a multidomain scaffolding protein that has a critical role in renal function. CD2AP is expressed in glomerular podocytes at the slit diaphragm, a modified adherens junction that comprises the protein filtration barrier of the kidney, and interacts with a number of protein ligands involved in cytoskeletal remodeling, membrane trafficking, cell motility, and cell survival. The structure of CD2AP is unknown. We used electron microscopy and single particle image analysis to determine the three-dimensional structure of recombinant full-length CD2AP and found that the protein is a tetramer in solution. Image reconstruction of negatively stained protein particles generated a structure at 21 Å resolution. The protein assumed a roughly spherical, very loosely packed structure. Analysis of the electron density map revealed that CD2AP consists of a central coiled-coil domain, which forms the tetramer interface, surrounded by four symmetry-related motifs, each containing three globular domains corresponding to the three SH3 domains. The spatial organization exposes the binding sites of all 12 SH3 domains in the tetramer, allowing simultaneous binding to multiple targets. Determination of the structure of CD2AP provides novel insights into the biology of this slit diaphragm protein and lays the groundwork for characterizing the interactions between key molecules of the slit diaphragm that control glomerular filtration.Journal of the American Society of Nephrology 02/2014; 25(7). DOI:10.1681/ASN.2013090949 · 9.47 Impact Factor
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ABSTRACT: Capping protein (CP) binds the fast growing barbed end of the actin filament and regulates actin assembly by blocking the addition and loss of actin subunits. Recent studies provide new insights into how CP and barbed-end capping are regulated. Filament elongation factors, such as formins and ENA/VASP (enabled/vasodilator-stimulated phosphoprotein), indirectly regulate CP by competing with CP for binding to the barbed end, whereas other molecules, including V-1 and phospholipids, directly bind to CP and sterically block its interaction with the filament. In addition, a diverse and unrelated group of proteins interact with CP through a conserved 'capping protein interaction' (CPI) motif. These proteins, including CARMIL (capping protein, ARP2/3 and myosin I linker), CD2AP (CD2-associated protein) and the WASH (WASP and SCAR homologue) complex subunit FAM21, recruit CP to specific subcellular locations and modulate its actin-capping activity via allosteric effects.Nature Reviews Molecular Cell Biology 09/2014; 15(10). DOI:10.1038/nrm3869 · 37.16 Impact Factor
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ABSTRACT: During EGFR internalization CIN85 bridges EGFR-Cbl complex, endocytic machinery and fusible membrane through the interactions of CIN85 with c-Cbl, endophilins and phosphatidic acid. These protein-protein and protein-lipid interactions are mediated or regulated by the positively charged C-terminal coiled-coil domain of CIN85. However, the details of CIN85-lipid interaction remain unknown. The present study suggested a possible electric interaction between the negative charge of phosphatidic acid and the positive charge of basic amino acids in coiled-coil domain.BMC Biochemistry 07/2014; 15(1):13. DOI:10.1186/1471-2091-15-13 · 1.94 Impact Factor