title: The linker region of breast cancer resistance protein ABCG2 is critical for coupling of ATP-dependent drug transport.
abstract: The ATP-binding cassette (ABC) transporters of class G display a different domain organisation than P-glycoprotein/ABCB1 and bacterial homologues with a nucleotide-binding domain preceding the transmembrane domain. The linker region connecting these domains is unique and its function and structure cannot be predicted. Sequence analysis revealed that the human ABCG2 linker contains a LSGGE sequence, homologous to the canonical C-motif/ABC signature present in all ABC nucleotide-binding domains. Predictions of disorder and of secondary structures indicated that this C2-sequence was highly mobile and located between an α-helix and a loop similarly to the C-motif. Point mutations of the two first residues of the C2-sequence fully abolished the transport-coupled ATPase activity, and led to the complete loss of cell resistance to mitoxantrone. The interaction with potent, selective and non-competitive, ABCG2 inhibitors was also significantly altered upon mutation. These results suggest an important mechanistic role for the C2-sequence of the ABCG2 linker region in ATP binding and/or hydrolysis coupled to drug efflux.
authors.affiliation: Equipe Labellisée Ligue 2014, BMSSI, UMR 5086 CNRS, Université Lyon 1, IBCP, 69007, Lyon, France.
authors.forename: S
authors.initials: S
authors.lastname: Macalou
authors.affiliation: Medical Oncology Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD, 20892, USA.
authors.forename: R W
authors.initials: RW
authors.lastname: Robey
authors.affiliation: Equipe Labellisée Ligue 2014, BMSSI, UMR 5086 CNRS, Université Lyon 1, IBCP, 69007, Lyon, France.
authors.forename: G
authors.initials: G
authors.lastname: Jabor Gozzi
authors.affiliation: Laboratory of Cell Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD, 20892, USA.
authors.forename: S
authors.initials: S
authors.lastname: Shukla
authors.affiliation: CelluloNet Biobank BB-0033-00072 Facility of UMS3444/US8/SFR Biosciences, IBCP, 69007, Lyon, France.
authors.forename: I
authors.initials: I
authors.lastname: Grosjean
authors.affiliation: MTA-SE Molecular Biophysics Research Group, Hungarian Academy of Sciences and Department of Biophysics and Radiation Biology, Semmelweis University, Budapest, Hungary.
authors.forename: T
authors.initials: T
authors.lastname: Hegedus
authors.affiliation: Laboratory of Cell Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD, 20892, USA.
authors.forename: S V
authors.initials: SV
authors.lastname: Ambudkar
authors.affiliation: Medical Oncology Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD, 20892, USA.
authors.forename: S E
authors.initials: SE
authors.lastname: Bates
authors.affiliation: Equipe Labellisée Ligue 2014, BMSSI, UMR 5086 CNRS, Université Lyon 1, IBCP, 69007, Lyon, France. a.dipietro@ibcp.fr.
authors.forename: A
authors.initials: A
authors.lastname: Di Pietro
mesh_terms: D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000241:Adenosine / Q000031:analogs & derivatives / Q000494:pharmacology; D000251:Adenosine Triphosphatases / Q000037:antagonists & inhibitors / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D000970:Antineoplastic Agents / Q000378:metabolism* / Q000494:pharmacology; D001943:Breast Neoplasms / Q000378:metabolism / Q000473:pathology; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D054659:Diketopiperazines; D019008:Drug Resistance, Neoplasm; D005260:Female; D057809:HEK293 Cells; D006576:Heterocyclic Compounds, 4 or More Rings; D006801:Humans; D008942:Mitoxantrone / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009363:Neoplasm Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment
chemical_list: C541506:3-(6-isobutyl-9-methoxy-1,4-dioxo-1,2,3,4,6,7,12,12a-octahydropyrazino(1',2'-1,6)pyrido(3,4-b)indol-3-yl)propionic acid tert-butyl ester; C116614:ABCG2 protein, human; D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters; D000970:Antineoplastic Agents; D054659:Diketopiperazines; D006576:Heterocyclic Compounds, 4 or More Rings; D009363:Neoplasm Proteins; D000255:Adenosine Triphosphate; D008942:Mitoxantrone; D000251:Adenosine Triphosphatases; D000241:Adenosine
keywords: ABC transporter; ATP hydrolysis; Breast cancer resistance protein/ABCG2; C motif/ABC signature; Drug efflux coupling; Specific sequence