CD2AP/CIN85 Balance Determines Receptor Tyrosine Kinase Signaling Response in Podocytes

Abstract

Defects in podocyte signaling are the basis of many inherited glomerular diseases leading to glomerulosclerosis. CD2-associated protein (CD2AP) is highly expressed in podocytes and is considered to play an important role in the maintenance of the glomerular slit diaphragm. Mice deficient for CD2AP (CD2AP ؊/؊ ) appear normal at birth but develop a rapid onset nephrotic syndrome at 3 weeks of age. We demonstrate that impaired intracellular signaling with subsequent podocyte damage is the reason for this delayed podocyte injury in CD2AP ؊/؊ mice. We document that CD2AP deficiency in podocytes leads to diminished signal initiation and termination of signaling pathways mediated by receptor tyrosine kinases (RTKs). In addition, we demonstrate that CIN85, a paralog of CD2AP, is involved in termination of RTK signaling in podocytes. CIN85 protein expression is increased in CD2AP ؊/؊ podocytes in vitro. Stimulation of CD2AP؊/؊ podocytes with various growth factors, including insulin-like growth factor 1, vascular endothelial growth factor, and fibroblast growth factor, resulted in a significantly decreased phosphatidylinositol 3-kinase/AKT and ERK signaling response. Moreover, increased CIN85 protein is detectable in podocytes in diseased CD2AP ؊/؊ mice, leading to decreased base-line activation of ERK and decreased phosphorylation after growth factor stimulation in vivo. Because repression of CIN85 protein leads to a restored RTK signaling response, our results support an important role of CD2AP/ CIN85 protein balance in the normal signaling response of podocytes. Phosphatidylinositol 3-kinase (PI3K)2 and Ras/ERK mitogen-activated protein kinase signaling pathways are key factors for determining the specificity of cellular responses, including cell proliferation, cell differentiation, and cell survival (1, 2). We and others have previously demonstrated that the PI3K/AKT signaling response plays an important role for podocyte survival in particular in the presence of active transforming growth factor ␤ (3, 4). We recently demonstrated that the PI3K/AKT response is directly targeted by cytokine cross-talk and subsequently influences the cellular outcome (5). The adaptor molecules CD2-associated protein (CD2AP) and CIN85 belong to a family of adaptor molecules that selectively control the spatial and temporal assembly of multiprotein complexes that transmit intracellular signals. For both molecules various interaction partners have been described placing them at the center of regulatory pathways involving signaling (6, 7), cytoskeletal arrangement (8, 9), vesicular trafficking (10), and endocytosis (11,12). In this study we demonstrate that CD2AP and CIN85 contribute to the balance of RTK signaling in podocytes. The anatomical localization of the podocyte exposes this highly specialized cell type to a variety of cellular stressors like stretch force, reactive oxygen species, osmotic milieu changes, cytokines and chemokines, filtrated toxins, and waste products. Therefore, this location requires the cells to have a highly e...

CD2-associated Protein (CD2AP) Enhances Casitas B Lineage Lymphoma-3/c (Cbl-3/c)-mediated Ret Isoform-specific Ubiquitination and Degradation via Its Amino-terminal Src Homology 3 Domains

“…These data suggest that Cbl-3/c, most likely as part of a protein complex, regulates Ret51 turnover in multiple cell types and not just in sympathetic neurons (8). Silencing of CD2AP in podocytes did not alter Ret51 degradation (data not shown), which is consistent with the observation that CD2AP deficiency in podocytes leads to a compensatory up-regulation of its paralog, CIN85 (18).…”

“…CD2AP, along with its paralog CIN85, regulate the association of Cbl proteins with receptors, thereby linking these receptor complexes to the internalization machinery (13)(14)(15)(16)(17). Interestingly, in glomerular podocytes, CD2AP deficiency induces the up-regulation of CIN85, leading to an excessive inhibition of receptor tyrosine kinase signaling (18). In the case of Ret51, it is not known how CD2AP augments the degradative function of Cbl-3/c or whether CD2AP has any effect on the E3 ligase activity of Cbl-3/c.…”

Comparative study of expression of adaptor proteins Ruk/CIN85 and CD2AP/CMS in normal andtumor human uterus tissues

“…Some proteins have been demonstrated to interact with both Ruk/CIN85 and CD2AP/CMS, whereas others bind only one of them. Experimental data obtained by different groups have shown similar functions [8,14] as well as antagonizing functions [34] for Ruk/CIN85 and CD2AP/CMS. It was reported that (Ruk/CIN85)/(CD2AP/CMS) balance is involved in the orchestrated signal transduction response [34].…”

“…Different combinations of promoter utilization and splicing events create multiple ruk/cin85 transcripts in various tissues and cell lines, and expression of some of these transcripts is regulated during development [3].In contrast, CD2AP/CMS is expressed as a single transcript that corresponds to mRNA encoding full-length form of Ruk/CIN85 with molecular mass of 85 kDa [17,34].…”

“…Both molecules are composed of three N-terminal SH3 domains, followed by a proline-rich region (providing binding sites for SH3 domain-containing proteins), an unstructured region of approximately 160 residues, enriched in Ser/Thr and C-terminal coil-coiled domain mediating homotypic and heterotypic interactions [17,34]. The SH3 domains share similarities among themselves and between family members, and their overlapping functions were identified [8,14,33].…”