Palladin interacts with SH3 domains of SPIN90 and Src and is required for Src-induced cytoskeletal remodeling

Abstract

Palladin and SPIN90 are widely expressed proteins, which participate in modulation of actin cytoskeleton by binding to a variety of scaffold and signaling molecules. Cytoskeletal reorganization can be induced by activation of signaling pathways, including the PDGF receptor and Src tyrosine kinase pathways. In this study we have analyzed the interplay between palladin, SPIN90 and Src and characterized the role of palladin and SPIN90 in PDGF and Src-induced cytoskeletal remodeling. We show that the SH3 domains of SPIN90 and Src directly bind palladin's poly-proline sequence and the interaction controls intracellular targeting of SPIN90. In PDGF-treated cells, palladin and SPIN90 co-localize in actin-rich membrane ruffles and lamellipodia. The effect of PDGF on the cytoskeleton is at least partly mediated by the Src kinase since PP2, a selective Src kinase family inhibitor, blocked PDGF-induced changes. Furthermore, expression of active Src kinase resulted in coordinated translocation of both palladin and SPIN90 to membrane protrusions. Knock-down of endogenous SPIN90 did not inhibit Src-induced cytoskeletal rearrangement, whereas knock-down of palladin resulted in cytoskeletal disorganization and inhibition of remodeling. Further studies showed that palladin is tyrosine phosphorylated in cells expressing active Src indicating bidirectional interplay between palladin and Src. These results may have implications in understanding the invasive and metastatic phenotype of neoplastic cells induced by Src.

Cell-permeable ceramides act as novel regulators of U937 cell–cell adhesion mediated by CD29, CD98, and CD147

“…4B). As assessed by immunoprecipitation analysis, the molecular association of Src and cytoplasmic actin, which is important for actin cytoskeleton rearrangement (Ronty et al 2007), was not diminished by C6-ceramide (Fig. 4C).…”

A Class III PDZ Binding Motif in the Myotilin and FATZ Families Binds Enigma Family Proteins: a Common Link for Z-Disc Myopathies

“…Extended activation of both kinases is thought to be important in the events that lead to myocyte dysfunction and to heart failure. Except for the finding that Src phosphorylates palladin (39); no information on the interplay between kinases and the myotilin or FATZ (calsarcin/myozenin) families has been available. We identify here CaMKII and PKA as kinases that are able to phosphorylate the C termini of myotilin and FATZ (calsarcin/ myozenin) family members.…”

“…Full-length myotilin (residues 1 to 498) and its variants encoding amino acids (aa) 1 to 270, 347 to 498, 441 to 498, and 214 to 250 or palladin (aa 715 to 772) were PCR amplified from human myotilin or palladin cDNA with the appropriate primer and subcloned into pAHP (38,39,41) for the expression of hemagglutinin (HA)-tagged proteins in eukaryotic cells or in vitro translation, into pEGFP-C2 (Clontech) for the expression of green fluorescent protein (GFP)-tagged proteins in eukaryotic cells, and into pGEX-4T1 (Pharmacia) glutathione S-transferase (GST) fusion vector for protein production in bacterial cells. Myotilin encoding aa 1 to 498 was cloned into the eukaryotic GST expression vector pDEST 27 vector (Gateway System; Invitrogen).…”

“…Yeast transformation, mating, and detection of protein interactions by determining the ␤-galactosidase activity were as previously described (16,39). The bait, myotilin cDNA encoding aa 102 to 498 in pGBKT7 vector, was used in screening a human skeletal muscle library in pACT2 (Clontech).…”

EMT, the cytoskeleton, and cancer cell invasion

“…Palladin binds F-actin and crosslinks actin filaments into bundles. It also interacts with Src, thereby affecting Src-mediated actin remodeling [165].…”