Mpl Genotype and Phenotype Correlations in the Chronic Myeloproliferative Disorders: Relationship of Mpl Mutations, JAK2V617F Status and Mpl Protein Expression.
Authors: Williams Donna M. , Kim Ann H. , Rogers Ophelia , Spivak Jerry L. , Moliterno Alison R. 
Published: October 2019

Abstract
The chronic myeloproliferative disorders (MPD), polycythemia vera (PV), essential thrombocytosis (ET) and idiopathic myelofibrosis (IMF), are linked by a common genetic lesion, JAK2V617F, and abnormalities of the thrombopoietin receptor, Mpl. JAK2 is an obligate Mpl chaperone and is responsible for its cell surface expression. We observed a reciprocal relationship between neutrophil JAK2V617F allele percentage and platelet Mpl expression in JAK2V617F-positive PV, IMF and ET patients. However, severely impaired platelet Mpl expression was also present in the majority of JAK2V617F-negative PV and IMF patients, with the most severe reductions in IMF. We previously identified a constitutional variation in the Mpl gene in the distal N-terminal domain (K39N) associated with thrombocytosis, and an Mpl splice variant involving the same region that was increased in PV and IMF patients compared to controls. Given the association of Mpl genetic lesions and protein expression abnormalities in the MPD, we hypothesized that other lesions in Mpl may be present in the MPD. To test this, we sequenced all of the exons, the flanking intervening sequences and the 5′-proximal untranslated region of the Mpl gene in 1 ET, 3 PV and 4 IMF patients, all of whom were JAK2V617F-negative and all of whom had markedly reduced (<5%) platelet Mpl protein. In one IMF patient, we identified a two base substitution at nucleotide 1543 of the Mpl gene which results in a tryptophan to lysine substitution at amino acid 515 (W515K); the wild-type sequence was not identified from either peripheral blood CD34+ cells or neutrophils isolated from this patient. The patient developed IMF at the age of 26, and had the disease 23 years before succumbing to complications of extensive extramedullary hematopoiesis and anemia. In a second male patient, we identified a heterozygous nucleotide transition in the intervening sequence distal to exon 10 close to the splice donor site; the mutation was also detected in buccal cell DNA. This patient developed IMF at age 35 and has had the disease for 31 years characterized by anemia and extramedullary hematopoiesis. To test whether this or other lesions in exon 10 of the Mpl gene were present, we directly sequenced exon 10 in 9 patients with JAK2V617F-negative PV, 21 with JAK2V617F-negative IMF, and 13 with JAK2V617F-positive IMF. Neither these nor other lesions in exon 10 were identified in these 43 MPD patients; no other unreported SNPs were identified either. We did not identify the recently described MplW515L reported as present in 4/44 JAK2V617F-negative IMF patients (PLoS Med.2006;3:e270) in our 21 JAK2V617F-negative IMF patients. These data indicate that, in contrast to other cytokine receptors, genetic and epigenetic variations of Mpl have unique roles in the pathogenesis of the chronic MPD. We conclude that significant disruptions of Mpl, whether acquired or constitutional, at the genomic level as described above or at the proteomic level as we have previously described, associate with a chronic MPD characterized by significant extramedullary hematopoiesis.