Analysis of the JAK2 gene in patients Na1061 and Na1253 and proposition of a new pathogenic model for polycythemia vera. (A) Representation of the 46/1 haplotype. The 46/1 haplotype is an approximately 280 Kb-long region of chromosome 9p that includes the entire JAK2, INSL6 and INSL4 genes. (B) Schematic representation of the JAK2 gene. JAK2 exons are represented by black boxes. JAK2 SNP rs10429491 (in exon 6), rs7034539 (in intron 18) and rs2230724 (in exon 19) positions are indicated with black bars. (C) Analysis by direct sequencing of JAK2 SNPs and JAK2V617F allelic ratios in gDNA of granulocytes and CD3⁺ lymphocytes (used as a control, healthy cells) of PV patients Na1061 and Na1253 (see primers in supplemental Table 3 and supplemental Figure 1). Black arrows indicate the different SNPs and JAK2V617F. Both patients were heterozygous for SNPs in CD3⁺ lymphocytes yet had SNP rs12343867 C-allele ratios in granulocyte gDNA (80% and 100%) compatible with homologous recombination of JAK2. (D) Detailed view of the JAK2 region. Results of the distortion of SNP allelic differences showed HR of part of JAK2 (exons 6-25) for Na1061 and of the whole 46/1 haplotype for Na1253. Regions of pre-JAK2V617F homologous recombination, not readily visible unless one looks for them, are indicated by double black arrows. (E) Karyoview of chromosomal aberrations. Bars depict the physical position and size of the aberration (purple, homologous recombination; blue, uniparental disomy events). Black arrows indicate the chronology of events, as deduced from the rs12343867 and V617F allelic ratios. For both patients the distortion of SNP allelic differences because of homologous recombination was higher at the telomeric end than in the centromeric region of chromosome 9p indicating 2 distinct partial 9pUPDs for Na1061 and 1 partial 9pUPD for Na1253. For both patients, SNP allelic distortion revealed pre-JAK2 homologous recombination (in purple). (F) Main and new pathogenic models for polycythemia vera and other MPN. The current model states that MPN patients carry or acquire a predisposition to MPN and mutation in the JAK2 gene; the JAK2 GGCC haplotype is one such genetic predisposition. In other patients, another genetic abnormality, congenital or acquired, presumably in a myeloid progenitor, is responsible for clonality, growth advantage and eventually, acquisition of JAK2 mutation -V617F being the most frequent - and MPN phenotype. Because high JAK2V617F loads are usually acquired via 9pUPD and most frequent in PV, acquisition of the PV phenotype is assumed to result from 9pUPD facilitated by JAK2V617F. Both JAK2 mutation and 9pUPD may occur more than once, leading to the development of one or several JAK2V617F-homozygous subclone(s). Disease phenotype and evolution, and occurrence of 9pUPD, may vary depending on parallel genetic events (eg, TET2 mutations) and the type of JAK2 mutation (eg, high mutant loads and 9pUPD are rare in patients with JAK2 exon 12 mutations). The new model adds an early step to the conventional model, stating that subsets of patients carrying the JAK2 GGCC haplotype may be predisposed to homologous recombination (HR) of JAK2 associated with growth advantage, followed or not by mutation in the JAK2 gene on the recombined allele and high JAK2 mRNA expression. Early JAK2 HR is compatible with all of the later steps leading to MPN according to the conventional pathogenic model: JAK2 mutation, 9pUPD, acquisition of parallel events in genes other than JAK2. The new model allows that a non-identified genetic event may facilitate JAK2 recombination and subsequent genetic alterations eventually leading to PV phenotype.