Chromosome Instability Underlies Stem Cell Dysfunction and Neoplasia In Widespread Hematologic Disease Induced By Deregulated Cyclin E

The Fbw7 ubiquitin ligase controls the expression of a number of oncoprotein substrates including cyclin E, Notch, c-Jun, and c-Myc. Using a knock-in mouse model (cyclin ET74AT393A), in which mutations were introduced into the cyclin E1 allele (Ccne1) to disrupt Fbw7-mediated ubiquitination specifically, we previously found that cyclin E dysregulation in vivo induces anemia with defects in erythroid differentiation and morphologic dysplasia of erythroid progenitors. We also found that cyclin ET74AT393A mice have fewer hematopoietic stem cells (HSCs) during steady-state hematopoiesis compared to wild-type counterparts. We performed serial transplantation experiments to assay comprehensively the self-renewal and multi-lineage reconstitution capacities of cyclin ET74AT393A HSCs. Contrary to our expectations, cyclin ET74AT393A HSC self-renewal appears normal after three rounds of serial transplantation; however, we identified defects in their multi-lineage reconstitution function. In cyclin ET74A T393A bone marrow erythroid cells, induction of a p53-dependent DNA damage response pathway appears to promote compensated erythropoiesis. In cyclin ET74A T393A HSCs, we similarly observed induced expression of canonical p53 target genes. We studied the effect of p53-loss on cyclin ET74A T393A HSCs and found that p53-null; cyclin ET74A T393A HSCs exhibit defects in both self-renewal and multi-lineage reconstitution. By enumerating chromosomes in metaphase spreads, we found p53-null; cyclin ET74A T393A hematopoietic stem and progenitor cells (HSPCs) demonstrate significant chromosomal instability (CIN). Importantly, we can recapitulate the self-renewal defects and CIN of cyclin ET74A T393A HSPCs with intact p53 by treating recipient animals with a single dose of 5-fluorouracil (5-FU). Thus, chromosomal stability is a key determinant for the maintenance of HSC self-renewal, and hematologic stress appears to unmask the potential for impaired Fbw7-dependent cyclin E ubiquitination to engender CIN in the presence of intact p53. Moreover, CIN is a characteristic feature of fatal T-cell malignancies that ultimately develop in recipients of cyclin ET74A T393A; p53-null HSCs. In pre-malignant thymocytes isolated from recipients of cyclin ET74A T393A; p53-null HSCs, aneuploidy is associated with the marked potentiation of cyclin E kinase activity in these cells by p53-loss. In malignant thymocytes, comparative genome hybridization analysis demonstrates clonal CIN associated with deregulated cyclin E expression combined with p53-loss. In toto, our data demonstrate the functional importance of cyclin E regulation by the Fbw7 ubiquitin ligase to the hematopoietic system and highlight CIN as a key mechanism underlying HSC dysfunction and malignancy induced by deregulated cyclin E in vivo. Disclosures: No relevant conflicts of interest to declare.