MiR-128 is a novel PHF6 targeting regulatory non-coding RNA acting as a T-ALL oncogene

T-cell acute lymphoblastic leukemia (T-ALL) arises through accumulation and interaction between multiple genetic events that perturb normal T-cell differentiation and drive the cellular hallmarks of oncogenesis. Through our previous work, we identified the plant homeodomain finger (PHF6) gene as a novel bona fide tumor suppressor gene in T-ALL (Van Vlierberghe et al. Nature Genetics 2010) and uncovered a mechanism of convergent multi-miRNA targeting of several important T-ALL tumor suppressor genes including PHF6 (Mavrakis et al. Nature Genetics 2011). In order to unravel the complete network of miRNAs convergently targeting PHF6, a miRNAome wide screen for the PHF6 3'UTR was performed in triplicate which yielded a total of 23 positive hits. Following assessment of miRNA expression levels in primary T-ALLs and normal thymocyte subsets, we selected miR-128 as a strong candidate for T-ALL specific modulation of PHF6 activity. MiR-128 showed a higher expression in T-ALL samples as compared to normal thymocytes and showed a distinct pattern of expression in normal developing T-cells with high expression in CD34+ cells and reduced expression in maturing thymocytes. The PHF6 3'UTR was found to contain three distinct 7-mer seed regions for miR-128 and independent 3'UTR luciferase assays and rescue experiments confirmed specific binding of miR-128. Based on correlation analysis between miR-128 and mRNA expression data in T-ALL samples, and subsequent gene set enrichment analysis, we identified a gene set described by Chiaretti et al. (Blood 2004) linked to T-cell differentiation suggesting that this miRNA could directly interfere with thymocyte maturation through dampening normal PHF6 function. A similar approach and KEGG pathway analysis yielded links for MAPK and insulin signaling pathways. In vitro modulation and gene expression analysis showed that miR-128 can modulate PHF6 expression on mRNA and protein level in T-ALL cell lines. Presently, we are testing the oncogenic potential of ectopic expression of miR-128 in a NOTCH1-induced T-ALL mouse model. Finally, we also looked for cooperative effects through targeting of other T-ALL tumor suppressor genes and identified NF1 as a bona fide target (Paschou et al. Plos One 2012) and IKFZ1 and SUZ12 as predicted targets of miR-128. Taken together, we identify miR-128 as a novel T-ALL oncogene with PHF6 as confirmed target and NF1, IKFZ1 and SUZ12 as further possible targets contributing to a cooperative effect on T-ALL oncogenesis. Our study adds T-ALL to a growing number of cancer entities in which miR-128 acts as oncogene by controlling apoptosis, survival and proliferation. In the light of recent successes in miRNA therapeutics, we propose miR-128 as a strong candidate marker for pre-clinical testing in T-ALL.

Jaar van publicatie:2013