Identification and functional analyses of disease-associated P4-ATPase phospholipid flippase variants in red blood cells

Research output: Contribution to journal/Conference contribution in journal/Contribution to newspaperJournal articleResearchpeer-review

  • Angela Y Liou, Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, British Columbia, V6T 1Z3, Canada
  • ,
  • Laurie L Molday, Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, British Columbia, V6T 1Z3, Canada, Canada
  • Jiao Wang, Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, British Columbia, V6T 1Z3, Canada
  • ,
  • Jens Peter Andersen
  • Robert S Molday, Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, British Columbia, V6T 1Z3, Canada, Canada

ATP-dependent phospholipid flippase activity crucial for generating lipid asymmetry was first detected in red blood cell (RBC) membranes, but the P4-ATPases responsible have not been directly determined. Using affinity-based MS, we show that ATP11C is the only abundant P4-ATPase phospholipid flippase in human RBCs, whereas ATP11C and ATP8A1 are the major P4-ATPasesinmouse RBCs. Wealso found that ATP11A and ATP11B are present at low levels. Mutations in the gene encoding ATP11C are responsible for blood and liver disorders, but the disease mechanisms are not known. Using heterologous expression, we show that the T415N substitution in the phosphorylation motif of ATP11C, responsible for congenital hemolytic anemia, reduces ATP11C expression, increases retention in the endoplasmic reticulum, and decreases ATPase activity by 61% relative to WT ATP11C. The I355K substitution in the transmembrane domain associated with cholestasis and anemia in mice was expressed at WT levels and trafficked to the plasma membrane but was devoid of activity. We conclude that the T415N variant causes significant protein misfolding, resulting in low protein expression, cellular mislocalization, and reduced functional activity. In contrast, the I355K variant folds and traffics normally but lacks key contacts required for activity. We propose that the loss in ATP11C phospholipid flippase activity coupled with phospholipid scramblase activity results in the exposure of phosphatidylserine on the surface of RBCs, decreasing RBC survival and resulting in anemia.

Original languageEnglish
JournalJournal of Biological Chemistry
Volume294
Issue17
Pages (from-to)6809-6821
Number of pages13
ISSN0021-9258
DOIs
Publication statusPublished - Apr 2019

    Research areas

  • ATP11C, ATPase, BINDING, CDC50 PROTEINS, CDC50A, ERYTHROCYTE-MEMBRANE, EXPRESSION, P-TYPE ATPASES, P4-ATPases, PHOSPHATIDYLSERINE, PUTATIVE AMINOPHOSPHOLIPID TRANSLOCASE, SUBCELLULAR-LOCALIZATION, TRANSPORT, cell biology, disease mechanisms, erythrocyte, lipid flippases, lipid transport, phosphatidylserine

See relations at Aarhus University Citationformats

ID: 146530111