A study of 36 unrelated cases with pure erythrocytosis revealed three new mutations in the erythropoietin receptor gene.

Thirty-six unrelated cases with erythrocytosis of unknown origin were investigated. Exons 5-8 of the erythropoietin receptor gene (EPOR), the von Hippel-Lindau gene, and the prolyl hydroxylase domain protein 2 gene (PHD2) were screened by direct DNA sequencing. The Janus kinase 2 mutation, JAK2 (Val617Phe), was screened by allele specific PCR. In this study, three new mutations of EPOR causing deletions in exon 8 were found: the first led directly to a stop codon [g.5957_5958delTT (p.Phe424X)], the second to a stop codon after one residue [g.5828_5829delCC (p.Pro381GlnfsX1)] and the third to a stop codon following a frameshift sequence of 23 residues [g.5971delC (p.Leu429TrpfsX23)]. One patient had a previously reported EPOR mutation [g.6146A>G (p.Asn487Ser)] and another, a silent one (g.5799G>A). All were heterozygotes. In addition, 2 patients were positive for JAK2 (Val617Phe), and 2 reported elsewhere, were mutated in the PHD2 gene [c.606delG (p.Met202IlefsX71).

Disturbance in the HIF-1alpha pathway associated with erythrocytosis: further evidences brought by frameshift and nonsense mutations in the prolyl hydroxylase domain protein 2 (PHD2) gene.

The hypoxia-inducible factor HIF-1 is the key regulator in cellular adaptation to hypoxia. Acting through a complex pathway, interconnected with VHL and kinases, it regulates a large number of genes, such as those involved in erythropoiesis, glycolysis, pH regulation, and angiogenesis. Recently, a missense mutation [c.950C>G (p.Pro317Arg)] in the prolyl hydroxylase domain protein 2 (PHD2) gene, whose encoded protein has HIF-1alpha as a substrate, provided evidence of the PHD2 role in a case of familial erythrocytosis. In this study, we looked for mutations in the PHD2 gene, in 74 patients with unidentified erythrocytosis. We found two heterozygous carriers of frameshift mutations [c.606delG (p.Met202IlefsX71) and c.840_841insA (p.Arg281ThrfsX3)]; both located in exon 1 and a heterozygous carrier of a nonsense mutation [c.1129C>T (p. Gln377X)] in exon 3. As a result of these mutations the encoded PHD2, if synthesized, would lose its catalytic activity. The genetic defects herein described are the first frameshift and nonsense mutations reported in the PHD2 gene and, as the previous missense mutation described, suggest that a decreased prolyl hydroxylase activity disturbing the oxygen-sensing pathway might be the cause of erythrocytosis. In addition to erythrocytosis, other complications, such as inflammatory arthromyalgia, have been observed in one case.

Two French Caucasian families with dominant thalassemia-like phenotypes due to hyper unstable hemoglobin variants: Hb Sainte Seve [codon 118 (-T)] and codon 127 [CAG-->TAG (Gln-->stop]).

We report two French Caucasian families suffering from dominant thalassemia-like phenotypes due to hyper unstable hemoglobin (Hb) variants. In both cases, molecular analysis revealed a defect localized in the third exon of the beta-globin gene, resulting in dramatic changes of the Hb structure. The first one is a new variant, Hb Sainte Seve, that is associated with a frameshift mutation at codon 118 (-T). In the second family, the disease resulted from a truncated protein due to a stop mutation at codon 127 [CAG-->TAG (Gln-->Stop)]. These two observations are additional evidences of the important role played by helix H in Hb stability: its partial absence, or a large structural modification, seems to be the major reason for the hyper instability of such molecules.