Abstract #94A

Pex13p and the actin cytoskeleton. Galina Kaneva¹, Alexandre Soulard¹, Jan Havlis², Sylvie Friant¹, Anna Shevchenko², Barbara Winsor¹. 1) UMR7156 CNRS - ULP, Génétique moléculaire Génomique et Microbiologie, Dépt. Génétique moléculaire et cellulaire, 21 rue Decartes, Strasbourg; 2) Max Planck Institute for Molecular Cell Biology and Genetics, Pfotenhauerstrasse 108, 01307 Dresden.
   SH3 domains most frequently interact with proline-rich regions containing a PXXP core motif in protein-protein interactions that are important in signal transduction, actin cytoskeleton organization, endocytosis, cytokinesis and membrane trafficking in a wide range of organisms. In a proteomic screen of all SH3 domains of Saccharomyces cerevisiae we found that the SH3 (Src homology-3 domain) domain of peroxin-13 (Pex13), expressed as a GST fusion, was able to trigger actin polymerization in whole cell extracts. This activity depends on the actin nucleation complex Arp2/3 and on the activator, Las17p, a homologue of WASP (Wiskott-Aldrich Syndrome Protein). Pex13 is a transmembrane protein of the peroxisomal membrane. Peroxisomes, like lysosomes, are bound by a single membrane and are filled with enzymes. Peroxisomes contain oxidatitive enzymes for degradation of toxic substances such as H2O2 ,the product of fatty-acid oxidation, and other metabolites. After translation peroxisomal proteins have to be imported into the peroxisomal compartment. The peroxisomal targeting sequences PTS1 (C-terminal position in the protein) and PTS2 (N-terminal position) are recognized by the receptor proteins Pex5 and Pex7, respectively. Pex13 interactions with Pex14 and Pex5 are essential for import into the peroxisome (Douangamath et al., 2002). Thus, we questioned whether the in vitro link between Pex13 and the actin machinery is important in vivo. Pex13-SH3 domain interacting proteins were pulled down from whole cell wild-type extracts, separated on SDS-PAGE and identified based on mass spectrometry analyses of gel bands. Peroxisomal import and biogenesis in actin cytoskeleton mutants was investigated.

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