Analysis of the subcellular distribution of avian p95-APP2, an ARF-GAP orthologous to mammalian paxillin kinase linker

Simona Paris, Lorena Za, Barbara Sporchia, Ivan De Curtis

Research output: Contribution to journalArticlepeer-review

Abstract

We describe here the identification and characterization of avian p95-APP2, a multi-domain protein of a recently identified family of ADP-ribosylation factor (ARF)-GTPase-activating proteins (GAPs) including mammalian G protein-coupled receptor kinases (GRK)-interactor 1 (GIT1), paxillin kinase linker (PKL), and GIT2, as well as avian p95-APP1. The p95-APP2 is eluted from Rac-GTP-γ-S, but not from Rac-GDP-β-S columns. As other members of the family, p95-APP2 has binding regions for the focal adhesion protein paxillin, and for the Rac exchanging factor PIX. Sequence comparison indicates that p95-APP2 is the avian orthologue of mammalian PKL. Expression studies showed a largely diffuse distribution of the full length p95-APP2, without evident effects on cell morphology. We observed a dramatic difference between the localization of the amino-terminal portion of the protein, including the ARF-GAP domain and the three ankyrin repeats, and the carboxy-terminal portion including the paxillin-binding site. Moreover, the expression of truncated carboxy-terminal polypeptides including both the PIX- and paxillin-binding regions leads to a marked localization of the protein together with paxillin at large vesicles. Comparison of the expression of corresponding ARF-GAP-deficient constructs from p95-APP2 and p95-APP1 shows their distribution at distinct endocytic compartments. Altogether, these data support a role of distinct members of this family of ARF-GAPs in the regulation of different steps of membrane traffic during cell motility, and suggest that p95-APP2 may shuttle between an intracellular compartment and the cell periphery, although, further work will be needed to address this point.

Original languageEnglish
Pages (from-to)826-837
Number of pages12
JournalInternational Journal of Biochemistry and Cell Biology
Volume34
Issue number7
DOIs
Publication statusPublished - 2002

Keywords

  • ARF-GAPs
  • Cell motility
  • Endosomes
  • GTP-binding proteins
  • PIX

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology

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