Disruption of a Functional Relationship Between PU.1 and Mir-155 during the Pathogenesis of Chronic Lymphocytic Leukemia (CLL)

Role of small non-coding microRNAs (miR) in hematopoiesis has been recently established by work demonstrating critical importance of miR-223 for granulocytic development (Johnnidis 2008) and miR-155 in lymphocytic differentiation (Thai 2007) in mice. The importance of miR-155 is further supported by several independent studies demonstrating its increased levels in human diffuse large B-cell lymphoma (Eis 2005, Kluvier 2005, Volinia 2006, Lawrie 2007, Rai 2008) and in chronic lymphocytic leukemia (CLL) (Fulci 2007, Marton 2008). PU.1 is ETS family transcription factor involved in myelo-lymphoid differentiation and is directly negatively regulated by miR-155 (Vigorito 2007). Studying primary CLL (N=27) and normal (N=13) peripheral blood mononuclear cells, we found significantly increased miR-155 levels and also demonstrate that PU.1 levels are significantly decreased. Next, we have tested genes that are targets of both miR-155 and PU.1 including CSF1R and FOS and again demonstrated their significantly decreased expression in CLL. Using gene expression profiling of CLL peripheral lymphocyte samples compared to normal controls, we have determined significant downregulation of the PU.1 mRNA and of numerous miR-155 targets previously reported to be required for myelo-lymphoid differentiation. These findings suggest that the relationship between PU.1 and miR-155 becomes disrupted in CLL. This in turn leads to overexpression of miR-155, reduced levels of PU.1 and of numerous miR-155 targets (N=2481 and N=400 respectively) and to inhibition of PU.1 regulated differentiation and proliferation programs. Next, we correlated negative cytogenetic prognostic features (p53 and ATM gene deletion detected by FISH) of the CLL patients with the miR-155 and PU.1 levels, In a relatively small-size patient cohort studied so far, the low ratios of PU.1/miR- 155 levels are similarly distributed among low, intermediate and high risk CLL patients, suggesting that inverse correlation of PU.1 and miR-155 represents a hallmark of CLL and likely represents a primary event during CLL pathogenesis.