title: Fluorescent FYVE Chimeras to Quantify PtdIns3P Synthesis During Autophagy.
abstract: Autophagy is the major cellular process of degradation and is modulated by several signaling pathways. Phosphatidylinositol 3-kinase (PtdIns3K) class III (Vps34) and PtdIns3K class I regulate the autophagy pathway positively and negatively, respectively. Both classes of PtdIns3K participate in the synthesis of phosphatidylinositol 3-phosphate (PtdIns3P), which plays a crucial role in autophagosome biogenesis and membrane traffic. PtdIns3P is a membrane phospholipid that is associated with endogenous FYVE domain-containing proteins. Indeed, such interactions facilitate autophagosome fusion with lysosomes and subsequent cargo degradation. During starvation-induced autophagy, the expression of FYVE domain-containing proteins increases, and their binding to PtdIns3P is strengthened. Nonetheless, not all FYVE domain proteins are related to the induction of autophagy. This method report presents the quantification of PtdIns3P synthesis by using cells either transiently transfected with or stably expressing FYVE-dsRed.
authors.affiliation: Centro de Investigación Biomédica en Red en Enfermedades Neurodegenerativas (CIBERNED), Universidad de Extremadura, Cáceres, Spain; Facultad de Enfermería y Terapia Ocupacional, Universidad de Extremadura, Cáceres, Spain.
authors.forename: S M S
authors.initials: SM
authors.lastname: Yakhine-Diop
authors.affiliation: Centro de Investigación Biomédica en Red en Enfermedades Neurodegenerativas (CIBERNED), Universidad de Extremadura, Cáceres, Spain; Facultad de Enfermería y Terapia Ocupacional, Universidad de Extremadura, Cáceres, Spain.
authors.forename: G
authors.initials: G
authors.lastname: Martínez-Chacón
authors.affiliation: Centro de Investigación Biomédica en Red en Enfermedades Neurodegenerativas (CIBERNED), Universidad de Extremadura, Cáceres, Spain; Facultad de Enfermería y Terapia Ocupacional, Universidad de Extremadura, Cáceres, Spain.
authors.forename: R A
authors.initials: RA
authors.lastname: González-Polo
authors.affiliation: Centro de Investigación Biomédica en Red en Enfermedades Neurodegenerativas (CIBERNED), Universidad de Extremadura, Cáceres, Spain; Facultad de Enfermería y Terapia Ocupacional, Universidad de Extremadura, Cáceres, Spain. Electronic address: jfuentes@unex.es.
authors.forename: J M
authors.initials: JM
authors.lastname: Fuentes
authors.affiliation: Centro de Investigación Biomédica en Red en Enfermedades Neurodegenerativas (CIBERNED), Universidad de Extremadura, Cáceres, Spain; Facultad de Enfermería y Terapia Ocupacional, Universidad de Extremadura, Cáceres, Spain. Electronic address: mnisosan@unex.es.
authors.forename: M
authors.initials: M
authors.lastname: Niso-Santano
mesh_terms: D001343:Autophagy; D002460:Cell Line; D005456:Fluorescent Dyes / Q000378:metabolism; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008856:Microscopy, Fluorescence / Q000379:methods*; D008967:Molecular Biology / Q000379:methods*; D018129:Phosphatidylinositol Phosphates / Q000032:analysis / Q000096:biosynthesis*; D011994:Recombinant Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*
chemical_list: C572072:FGD5 protein, human; D005456:Fluorescent Dyes; D020662:Guanine Nucleotide Exchange Factors; D008164:Luminescent Proteins; D018129:Phosphatidylinositol Phosphates; D011994:Recombinant Proteins; C413663:fluorescent protein 583; C055525:phosphatidylinositol 3-phosphate
keywords: 3-Methyladenine; Autophagy; FYVE domain; Fluorescence microscopy; PtdIns3K; PtdIns3P; Starvation