title: Methods to Study the BECN1 Interactome in the Course of Autophagic Responses.
abstract: Autophagy is an extremely dynamic process that mediates the rapid degradation of intracellular components in response to different stress conditions. The autophagic response is executed by specific protein complexes, whose function is regulated by posttranslational modifications and interactions with positive and negative regulators. A comprehensive analysis of how autophagy complexes are temporally modified upon stress stimuli is therefore particularly relevant to understand how this pathway is regulated. Here, we describe a method to define the protein-protein interaction network of a central complex involved in autophagy induction, the Beclin 1 complex. This method is based on the quantitative comparison of protein complexes immunopurified at different time points using a stable isotope labeling by amino acids in cell culture approach. Understanding how the Beclin 1 complex dynamically changes in response to different stress stimuli may provide useful insights to disclose novel molecular mechanisms responsible for the dysregulation of autophagy in pathological conditions, such as cancer, neurodegeneration, and infections.
authors.affiliation: Freiburg Institute for Advanced Studies (FRIAS), University of Freiburg, Freiburg, Germany; National Institute for Infectious Diseases I.R.C.C.S. 'Lazzaro Spallanzani', Rome, Italy.
authors.forename: M
authors.initials: M
authors.lastname: Antonioli
authors.affiliation: National Institute for Infectious Diseases I.R.C.C.S. 'Lazzaro Spallanzani', Rome, Italy.
authors.forename: F
authors.initials: F
authors.lastname: Ciccosanti
authors.affiliation: Freiburg Institute for Advanced Studies (FRIAS), University of Freiburg, Freiburg, Germany.
authors.forename: J
authors.initials: J
authors.lastname: Dengjel
authors.affiliation: National Institute for Infectious Diseases I.R.C.C.S. 'Lazzaro Spallanzani', Rome, Italy; University of Salento, Lecce, Italy. Electronic address: gianmaria.fimia@inmi.it.
authors.forename: G M
authors.initials: GM
authors.lastname: Fimia
mesh_terms: D001343:Autophagy / Q000502:physiology*; D000071186:Beclin-1 / Q000032:analysis / Q000378:metabolism*; D002460:Cell Line; D002853:Chromatography, Liquid / Q000379:methods; D006801:Humans; D007553:Isotope Labeling / Q000379:methods; D025941:Protein Interaction Mapping / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: C491997:BECN1 protein, human; D000071186:Beclin-1
keywords: Autophagic response; Beclin 1 complex; Immunoprecipitation; Interactome; Mass spectrometry; Protein complexes; Proteomics; SILAC