title: The ML1Nx2 Phosphatidylinositol 3,5-Bisphosphate Probe Shows Poor Selectivity in Cells.
abstract: Phosphatidylinositol (3,5)-bisphosphate (PtdIns(3,5)P2) is a quantitatively minor phospholipid in eukaryotic cells that plays a fundamental role in regulating endocytic membrane traffic. Despite its clear importance for cellular function and organism physiology, mechanistic details of its biology have so far not been fully elucidated. In part, this is due to a lack of experimental tools that specifically probe for PtdIns(3,5)P2 in cells to unambiguously identify its dynamics and site(s) of action. In this study, we have evaluated a recently reported PtdIns(3,5)P2 biosensor, GFP-ML1Nx2, for its veracity as such a probe. We report that, in live cells, the localization of this biosensor to sub-cellular compartments is largely independent of PtdIns(3,5)P2, as assessed after pharmacological, chemical genetic or genomic interventions that block the lipid's synthesis. We therefore conclude that it is unwise to interpret the localization of ML1Nx2 as a true and unbiased biosensor for PtdIns(3,5)P2.
authors.affiliation: Department of Cell Biology, University of Pittsburgh School of Medicine, Pittsburgh, PA, United States of America; Program in Developmental Neuroscience, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD, 20892, United States of America.
authors.forename: Gerald R V
authors.initials: GR
authors.lastname: Hammond
authors.affiliation: Department of Medical Biology, Graduate School of Medicine, Akita University, Akita, Japan.
authors.forename: Shunsuke
authors.initials: S
authors.lastname: Takasuga
authors.affiliation: Department of Medical Biology, Graduate School of Medicine, Akita University, Akita, Japan; Research Center for Biosignal, Akita University, Akita, 010-8543, Japan.
authors.forename: Takehiko
authors.initials: T
authors.lastname: Sasaki
authors.affiliation: Program in Developmental Neuroscience, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD, 20892, United States of America.
authors.forename: Tamas
authors.initials: T
authors.lastname: Balla
mesh_terms: D000818:Animals; D019556:COS Cells; D002462:Cell Membrane / Q000378:metabolism; D002477:Cells / Q000378:metabolism*; D002522:Chlorocebus aethiops; D011992:Endosomes / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000378:metabolism; D051379:Mice; D015335:Molecular Probes / Q000378:metabolism*; D000066450:Mouse Embryonic Stem Cells / Q000378:metabolism; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism*; D020696:rab5 GTP-Binding Proteins / Q000378:metabolism
chemical_list: D015335:Molecular Probes; D018129:Phosphatidylinositol Phosphates; C106336:phosphatidylinositol 3,5-diphosphate; D049452:Green Fluorescent Proteins; D020696:rab5 GTP-Binding Proteins