A-kinase anchor protein 84/121 are targeted to mitochondria and mitotic spindles by overlapping amino-terminal motifs

Luca Cardone, Tiziana De Cristofaro, Adelina Affaitati, Corrado Garbi, Michael D. Ginsberg, Michele Saviano, Stelio Varrone, Charles S. Rubin, Max E. Gottesman, Enrico V. Avvedimento, Antonio Feliciello

Research output: Contribution to journalArticlepeer-review

Abstract

A-kinase anchor proteins (AKAPs) assemble multi-enzyme signaling complexes in proximity to substrate/effector proteins, thus directing and amplifying membrane-generated signals. S-AKAP84 and AKAP121 are alternative splicing products with identical NH2 termini. These AKAPs bind and target protein kinase A (PKA) to the outer mitochondrial membrane. Tubulin was identified as a binding partner of S-AKAP84 in a yeast two-hybrid screen. Immunoprecipitation and co-sedimentation experiments in rat testis extracts confirmed the interaction between microtubules and S-AKAP84. In situ immunostaining of testicular germ cells (GC2) shows that AKAP121 concentrates on mitochondria in interphase and on mitotic spindles during M phase. Purified tubulin binds directly to S-AKAP84 but not to a deletion mutant lacking the mitochondrial targeting domain (MT) at residues 1-30. The MT is predicted to form a highly hydrophobic α-helical wheel that might also mediate interaction with tubulin. Disruption of the wheel by site-directed mutagenesis abolished tubulin binding and reduced mitochondrial attachment of an MT-GFP fusion protein. Some MT mutants retain tubulin binding but do not localize to mitochondria. Thus, the tubulin-binding motif lies within the mitochondrial attachment motif. Our findings indicate that S-AKAP84/AKAP121 use overlapping targeting motifs to localize signaling enzymes to mitochondrial and cytoskeletal compartments.

Original languageEnglish
Pages (from-to)663-675
Number of pages13
JournalJournal of Molecular Biology
Volume320
Issue number3
DOIs
Publication statusPublished - 2002

Keywords

  • A-kinase anchor proteins
  • Mitochondria
  • Mitotic spindles
  • S-AKAP84/AKAP121

ASJC Scopus subject areas

  • Virology

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