MYBL2 Is a Sub-haploinsufficient Tumor Suppressor Gene in Myeloid Malignancy (RNAi)

A dosage-dependent role for tumor suppressor genes in the initiation of myeloid malignancies remains controversial. Here we show that MYBL2 is expressed at sharply reduced levels in CD34+ cells from most patients with myelodysplastic syndrome (MDS; 65%; n=26). In a murine competitive reconstitution model, Mybl2 knockdown by RNAi to 20-30% of normal levels in multipotent hematopoietic progenitors led to clonal dominance by these “sub-haploinsufficient” cells, affecting all blood cell lineages. By 6 months post-transplantation, the reconstituted mice had developed a myeloproliferative/myelodysplastic disorder originating from the cells with aberrantly reduced Mybl2 expression. Thus, downregulation of MYBL2 activity to levels below those predicted by classical haploinsufficiency drives the clonal expansion of hematopoietic progenitors in a large fraction of human MDS cases. Cryopreserved CD34+ bone marrow were cultured for 24 hours, transduced with shRNA-encoding vectors and selected with Puromycin for 48 hours. Vectors shRNA 1-8 are specifically targeting MYBL2, vectors shRNA A-D encoded an irrelevant control shRNAs and one culture was left untreated (no shRNA). Cells were harvested after Puromycin selection and total RNA was prepared using Trizol (LifeTechnologies).