title: An ENU-induced recessive mutation in Mpl leads to thrombocytopenia with overdominance.
abstract: OBJECTIVE
  The aim of this study was to identify and characterize the causative mutation in the thrombocytopenic mouse strain HLB219 that was generated at the Jackson Laboratory as part of a large-scale N-ethyl-N-nitrosourea mutagenesis screen.
  
  
  METHODS
  The HLB219 mutation was identified by interval mapping of F2 mice generated from intercross breeding of HLB219 to both BALB/cByJ (BALB) and 129/SvImJ (129/Sv). Mpl was identified as a candidate gene and sequenced. The mutation was characterized in vivo in mouse hematopoietic stem/progenitor cell assays and in cell culture by expression in Ba/F3 cells.
  
  
  RESULTS
  A novel mutation in the thrombopoietin (TPO) receptor Mpl in HLB219 mice caused a Cys-->Arg substitution at codon 40 in the extracellular region of the receptor. Mice homozygous for the Mpl(hlb219) mutation had an 80% decrease in the number of platelets in comparison to the wild-type C57BL/6J strain, low numbers of bone marrow megakaryocytes, high TPO levels, and decreased competitive repopulating ability, consistent with a loss-of-function mutation in the receptor. Mice heterozygous for Mpl(hlb219) however, showed an overdominance effect with a significant increase in platelet number. Functional analysis in vitro demonstrated that Ba/F3 cells expressing the mutant MPL(hlb219) protein failed to activate extracellular signal-regulated kinase and signal transducers and activators of transcription 5, but proliferated in the absence of TPO and required constitutive phosphorylation of RAC-alpha serine/threonine protein kinase (AKT) for cytokine-independent growth.
  
  
  CONCLUSIONS
  Thrombocytopenia in HLB219 mice is caused by a recessive mutation in Mpl that abrogates mitogen-activated protein kinase-extracellular signal regulated kinase and janus kinase-signal transducers and activators of transcription signaling.
authors.affiliation: Department of Genetics, Case Western Reserve University, Cleveland, Ohio 44106-4955, USA.
authors.forename: E Ricky
authors.initials: ER
authors.lastname: Chan
authors.forename: Heather
authors.initials: H
authors.lastname: Lavender
authors.forename: Geqiang
authors.initials: G
authors.lastname: Li
authors.forename: Peter
authors.initials: P
authors.lastname: Haviernik
authors.forename: Kevin D
authors.initials: KD
authors.lastname: Bunting
authors.forename: Mark D
authors.initials: MD
authors.lastname: Adams
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D001947:Breeding; D002460:Cell Line; D005038:Ethylnitrosourea / Q000633:toxicity*; D005260:Female; D005808:Genes, Recessive; D006720:Homozygote; D020935:MAP Kinase Signaling System / Q000187:drug effects / Q000235:genetics; D051379:Mice; D008807:Mice, Inbred BALB C; D020929:Mitogen-Activated Protein Kinase Kinases / Q000235:genetics / Q000378:metabolism; D009153:Mutagens / Q000633:toxicity*; D020125:Mutation, Missense; D053628:Receptors, Thrombopoietin / Q000235:genetics* / Q000378:metabolism; D013921:Thrombocytopenia / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects / Q000235:genetics
chemical_list: C481270:Mpl protein, mouse; D009153:Mutagens; D053628:Receptors, Thrombopoietin; D020929:Mitogen-Activated Protein Kinase Kinases; D005038:Ethylnitrosourea