title: Lysosomal interaction of Akt with Phafin2: a critical step in the induction of autophagy.
abstract: Autophagy is an evolutionarily conserved mechanism for the gross disposal of intracellular proteins in mammalian cells and dysfunction in this pathway has been associated with human disease. Although the serine threonine kinase Akt is suggested to play a role in this process, little is known about the molecular mechanisms by which Akt induces autophagy. Using a yeast two-hybrid screen, Phafin2 (EAPF or PLEKHF2), a lysosomal protein with a unique structure of N-terminal PH (pleckstrin homology) domain and C-terminal FYVE (Fab 1, YOTB, Vac 1, and EEA1) domain was found to interact with Akt. A sucrose gradient fractionation experiment revealed that both Akt and Phafin2 co-existed in the same lysosome enriched fraction after autophagy induction. Confocal microscopic analysis and BiFC analysis demonstrated that both Akt and Phafin2 accumulate in the lysosome after induction of autophagy. BiFC analysis using PtdIns (3)P interaction defective mutant of Phafin2 demonstrated that lysosomal accumulation of the Akt-Phafin2 complex and subsequent induction of autophagy were lysosomal PtdIns (3)P dependent events. Furthermore, in murine macrophages, both Akt and Phafin2 were required for digestion of fluorescent bacteria and/or LPS-induced autophagy. Taken together, these findings establish that lysosomal accumulation of Akt and Phafin2 is a critical step in the induction of autophagy via an interaction with PtdIns (3)P.
authors.affiliation: Division of Cancer Biology, Institute for Genetic Medicine, Hokkaido University, Kita-ku, Sapporo, Japan.
authors.forename: Mami
authors.initials: M
authors.lastname: Matsuda-Lennikov
authors.affiliation: Division of Cancer Biology, Institute for Genetic Medicine, Hokkaido University, Kita-ku, Sapporo, Japan.
authors.forename: Futoshi
authors.initials: F
authors.lastname: Suizu
authors.affiliation: Division of Cancer Biology, Institute for Genetic Medicine, Hokkaido University, Kita-ku, Sapporo, Japan.
authors.forename: Noriyuki
authors.initials: N
authors.lastname: Hirata
authors.affiliation: Division of Cancer Biology, Institute for Genetic Medicine, Hokkaido University, Kita-ku, Sapporo, Japan.
authors.forename: Manabu
authors.initials: M
authors.lastname: Hashimoto
authors.affiliation: Division of Cancer Biology, Institute for Genetic Medicine, Hokkaido University, Kita-ku, Sapporo, Japan.
authors.forename: Kohki
authors.initials: K
authors.lastname: Kimura
authors.affiliation: Division of Cancer Biology, Institute for Genetic Medicine, Hokkaido University, Kita-ku, Sapporo, Japan.
authors.forename: Tadashi
authors.initials: T
authors.lastname: Nagamine
authors.affiliation: Laboratory of Pathophysiology and Signal Transduction, Hokkaido University Graduate School of Medicine, Kita-ku, Sapporo, Japan.
authors.forename: Yoichiro
authors.initials: Y
authors.lastname: Fujioka
authors.affiliation: Laboratory of Pathophysiology and Signal Transduction, Hokkaido University Graduate School of Medicine, Kita-ku, Sapporo, Japan.
authors.forename: Yusuke
authors.initials: Y
authors.lastname: Ohba
authors.affiliation: Laboratory of Histology and Cytology, Hokkaido University Graduate School of Medicine, Kita-ku, Sapporo, Japan.
authors.forename: Toshihiko
authors.initials: T
authors.lastname: Iwanaga
authors.affiliation: Division of Cancer Biology, Institute for Genetic Medicine, Hokkaido University, Kita-ku, Sapporo, Japan.
authors.forename: Masayuki
authors.initials: M
authors.lastname: Noguchi
mesh_terms: D000818:Animals; D001343:Autophagy; D006801:Humans; D008247:Lysosomes / Q000378:metabolism* / Q000648:ultrastructure; D051379:Mice; D008954:Models, Biological; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism; D011485:Protein Binding; D021381:Protein Transport; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D033921:Vesicular Transport Proteins / Q000378:metabolism*
chemical_list: D018129:Phosphatidylinositol Phosphates; D051057:Proto-Oncogene Proteins c-akt; D033921:Vesicular Transport Proteins; C547687:PLEKHF2 protein, human; C000591291:PLEKHF2 protein, mouse