Reporting Form from miR-1202 acts as anti-oncomiR in myeloid leukaemia by down-modulating GATA-1_S expression

Transient abnormal myelopoiesis (TAM) is a Down syndrome-related pre-leukemic condition characterized by somatic mutations in the hematopoietic transcription factor GATA-1 that result in exclusive production of its shorter isoform (GATA-1S). Given the common hallmark of altered miRNA expression profiles in haematological malignancies and the pro-leukemic role of GATA-1S, we aimed to search for miRNAs potentially able to modulate the expression of GATA-1 isoforms. Starting from an in silico prediction of miRNA binding sites in the GATA-1 transcript, miR-1202 came into our sight as potential regulator of GATA-1 expression. Expression studies in K562 cells revealed that miR-1202 directly targets GATA-1, negatively regulates its expression, impairs GATA-1S production, reduces cell proliferation, and increases apoptosis sensitivity. Furthermore, data from TAM and myeloid leukaemia patients provided substantial support to our study by showing that miR-1202 down-modulation is accompanied by increased GATA-1 levels, with more marked effects on GATA-1S. These findings indicate that miR-1202 acts as an anti-oncomiR in myeloid cells and may impact leukemogenesis at least in part by down-modulating GATA-1S levels

Supplemental Experimental Procedures from miR-1202 acts as anti-oncomiR in myeloid leukaemia by down-modulating GATA-1_S expression

Northern blot analysis and MicroRNAs cloning and capture procedur

Supplementary Tables S1-S4 from miR-1202 acts as anti-oncomiR in myeloid leukaemia by down-modulating GATA-1_S expression

Table S1. Clinical and molecular characterization of TMD/AML-DS patients with GATA-1 mutations. Table S2. Mutant sequences within or outside the putative hsa-miR-1202 binding site generated in the psiCHECK™-2 Vector GATA1 exon2 construct. Table S3. Primer sequences used for quantitative Real-time PCR analysis.Table S4. Antibodies used for western blot analysis

Original blot images from miR-1202 acts as anti-oncomiR in myeloid leukaemia by down-modulating GATA-1_S expression

Transient abnormal myelopoiesis (TAM) is a Down syndrome-related pre-leukemic condition characterized by somatic mutations in the hematopoietic transcription factor GATA-1 that result in exclusive production of its shorter isoform (GATA-1S). Given the common hallmark of altered miRNA expression profiles in haematological malignancies and the pro-leukemic role of GATA-1S, we aimed to search for miRNAs potentially able to modulate the expression of GATA-1 isoforms. Starting from an in silico prediction of miRNA binding sites in the GATA-1 transcript, miR-1202 came into our sight as potential regulator of GATA-1 expression. Expression studies in K562 cells revealed that miR-1202 directly targets GATA-1, negatively regulates its expression, impairs GATA-1S production, reduces cell proliferation, and increases apoptosis sensitivity. Furthermore, data from TAM and myeloid leukaemia patients provided substantial support to our study by showing that miR-1202 down-modulation is accompanied by increased GATA-1 levels, with more marked effects on GATA-1S. These findings indicate that miR-1202 acts as an anti-oncomiR in myeloid cells and may impact leukemogenesis at least in part by down-modulating GATA-1S levels