title: Residual pyruvate kinase activity in PKLR-deficient erythroid precursors of a patient suffering from severe haemolytic anaemia.
abstract: OBJECTIVE
  Here, we present a 7-year-old patient suffering from severe haemolytic anaemia. The most common cause of chronic hereditary non-spherocytic haemolytic anaemia is red blood cell pyruvate kinase (PK-R) deficiency. Because red blood cells rely solely on glycolysis to generate ATP, PK-R deficiency can severely impact energy supply and cause reduction in red blood cell lifespan. We determined the underlying cause of the anaemia and investigated how erythroid precursors in the patient survive.
  
  
  METHODS
  PK activity assays, Western blot and Sanger sequencing were employed to determine the underlying cause of the anaemia. Patient erythroblasts were cultured and reticulocytes were isolated to determine PK-R and PKM2 contribution to glycolytic activity during erythrocyte development.
  
  
  RESULTS
  We found a novel homozygous mutation (c.583G>A) in the PK-R coding gene (PKLR). Although this mutation did not influence PKLR mRNA production, no PK-R protein could be detected in the red blood cells nor in its precursors. In spite of the absence of PK-R, the reticulocytes of the patient exhibited 20% PK activity compared with control. Western blotting revealed that patient erythroid precursors, like controls, express residual PKM2.
  
  
  CONCLUSIONS
  We conclude that PKM2 rescues glycolysis in PK-R-deficient erythroid precursors.
authors.affiliation: Sanquin Research and Landsteiner Laboratory, Department of Blood Cell Research, University of Amsterdam, Amsterdam, The Netherlands.
authors.forename: Thomas R L
authors.identifier: http://orcid.org/0000-0002-2864-4073
authors.initials: TRL
authors.lastname: Klei
authors.affiliation: Laboratory for Red Blood Cell Diagnostics, Sanquin, Amsterdam, The Netherlands.|Sara Medical Genetics Centre, Tehran, Iran.
authors.forename: Sima
authors.initials: S
authors.lastname: Kheradmand Kia
authors.affiliation: Laboratory for Red Blood Cell Diagnostics, Sanquin, Amsterdam, The Netherlands.
authors.forename: Martijn
authors.initials: M
authors.lastname: Veldthuis
authors.affiliation: Sanquin Research and Landsteiner Laboratory, Department of Blood Cell Research, University of Amsterdam, Amsterdam, The Netherlands.
authors.forename: Boukje M
authors.initials: BM
authors.lastname: Beuger
authors.affiliation: Laboratory for Red Blood Cell Diagnostics, Sanquin, Amsterdam, The Netherlands.
authors.forename: Judy
authors.initials: J
authors.lastname: Geissler
authors.affiliation: PeyvandLab Complex, Shiraz, Iran.
authors.forename: Javad
authors.initials: J
authors.lastname: Dehbozorgian
authors.affiliation: Hematology Research Centre, Shiraz University of Medical Sciences, Shiraz, Iran.
authors.forename: Mehran
authors.initials: M
authors.lastname: Karimi
authors.affiliation: Sanquin Research and Landsteiner Laboratory, Department of Blood Cell Research, University of Amsterdam, Amsterdam, The Netherlands.
authors.forename: Robin
authors.initials: R
authors.lastname: van Bruggen
authors.affiliation: Sanquin Research and Landsteiner Laboratory, Department of Blood Cell Research, University of Amsterdam, Amsterdam, The Netherlands.|Laboratory for Red Blood Cell Diagnostics, Sanquin, Amsterdam, The Netherlands.
authors.forename: Rob
authors.initials: R
authors.lastname: van Zwieten
mesh_terms: D000746:Anemia, Hemolytic, Congenital Nonspherocytic / Q000201:enzymology / Q000235:genetics* / Q000473:pathology; D001483:Base Sequence; D002352:Carrier Proteins / Q000235:genetics*; D002454:Cell Differentiation; D002648:Child; D003241:Consanguinity; D004900:Erythroblasts / Q000201:enzymology* / Q000473:pathology; D015870:Gene Expression; D006019:Glycolysis / Q000235:genetics; D006720:Homozygote; D006801:Humans; D008297:Male; D008565:Membrane Proteins / Q000172:deficiency / Q000235:genetics*; D009154:Mutation; D022423:Myeloid Cells / Q000166:cytology / Q000201:enzymology; D061251:Primary Cell Culture; D011770:Pyruvate Kinase / Q000172:deficiency* / Q000235:genetics*; D015323:Pyruvate Metabolism, Inborn Errors / Q000201:enzymology / Q000235:genetics* / Q000473:pathology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012156:Reticulocytes / Q000201:enzymology* / Q000473:pathology; D013963:Thyroid Hormones / Q000172:deficiency / Q000235:genetics*; D000097810:Thyroid Hormone-Binding Proteins
chemical_list: D002352:Carrier Proteins; D008565:Membrane Proteins; D011770:Pyruvate Kinase; D012333:RNA, Messenger; D013963:Thyroid Hormones; D000097810:Thyroid Hormone-Binding Proteins
keywords: PK-R; PKM2; pyruvate kinase deficiency; red blood cell; reticulocyte