Palladin is a novel binding partner for Ena/VASP family members.
Palladin is an actin-associated protein that contains proline-rich motifs within its amino-terminal sequence that are similar to motifs found in zyxin, vinculin, and the Listeria protein ActA. These motifs are known to be potential binding sites for the Vasodilator-Stimulated Phosphoprotein (VASP). Here, we demonstrate that palladin is an additional direct binding partner for VASP, by using co-immunoprecipitation and blot overlay techniques with both endogenous palladin and recombinant myc-tagged palladin. These results show that VASP binds to full-length palladin and also to the amino-terminal half of palladin, where the polyproline motifs are located. Using a synthetic peptide array, two discrete binding sites for VASP were identified within palladin's proline-rich amino-terminal domain. Using double-label immunofluorescence staining of fully-spread and actively-spreading fibroblasts, the extent of co-localization of palladin and VASP was explored. These proteins were found to strongly co-localize along stress fibers, and partially co-localize in focal adhesions, lamellipodia, and focal complexes. These results suggest that the recently described actin-associated protein palladin may play an important role in recruiting VASP to sites of actin filament growth, anchorage, and crosslinking.
Boukhelifa
Malika
M
Department of Cell and Molecular Physiology, University of North Carolina at Chapel Hill, 27599-7545, USA.
Parast
Mana M
MM
Bear
James E
JE
Gertler
Frank B
FB
Otey
Carol A
CA
D000595:Amino Acid Sequence; D000818:Animals; D015815:Cell Adhesion Molecules / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D002642:Chick Embryo; D003598:Cytoskeletal Proteins / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D005347:Fibroblasts / Q000378:metabolism; D006023:Glycoproteins; D006801:Humans; D008667:Metalloproteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008840:Microfilament Proteins; D008969:Molecular Sequence Data; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D017386:Sequence Homology, Amino Acid; D016596:Vinculin / Q000235:genetics / Q000378:metabolism; D060589:Zyxin; D000098662:Vasodilator-Stimulated Phosphoprotein
D015815:Cell Adhesion Molecules; D003598:Cytoskeletal Proteins; D004268:DNA-Binding Proteins; C092744:ENA-VASP proteins; D006023:Glycoproteins; D008667:Metalloproteins; D008840:Microfilament Proteins; C412722:PALLD protein, human; D010750:Phosphoproteins; C092842:ZYX protein, human; C498285:Zyx protein, mouse; D060589:Zyxin; C485873:palladin protein, mouse; D000098662:Vasodilator-Stimulated Phosphoprotein; D016596:Vinculin