Regulation of FLT3LG and MCL1 by miR-424 and miR-181c in GATA2 Deficiency

Authors

• SH Droll
• W Wang
• Z Zhao
• AP Hsu
• SM Holland
• KR Calvo

Abstract

GATA binding protein 2 (GATA2) is a zinc finger transcription factor required for the survival of hematopoietic stem cells. Germline GATA2 mutations result in haploinsufficiency, which is associated with severe immunodeficiency, myelodysplasia, acute myeloid leukemia, and increased serum levels of FLT3LG. miRNAs are short, non-coding RNAs that inhibit translation of messenger RNAs by targeting the 3’ untranslated region (3’UTR). Previous analysis of EBV-immortalized B cells revealed that miR-424 expression was significantly decreased (fc=-7.96, p=.02), and miR-181c expression was increased, (fc=2.69, p=.02) in cells derived from GATA2 patients. Functional studies were performed in cell lines demonstrating that overexpression of GATA2 resulted in increased miR-424 expression in K562 (fc=2, p=.007) and HL60 (fc=1.6, p=.03) cell lines; while miR-181c expression was decreased in K562 (fc= -6.8, p=.02) and HL60 (fc= -1.8, p=.02). Predicted mRNA targets of miR-424 included FLT3LG; while MCL-1 was a predicted target for miR-181c. To investigate the relationship between these miRs and putative targets, miR-424 was overexpressed resulting in significantly decreased FLT3LG in K562 (fc= -5.8, p=.001) and HL60 (fc= -3.3, p=.03). To demonstrate specificity, the 3’UTRs of FLT3LG and MCL1 were sub-cloned into a luciferase reporter vector. Expression of miR-424 with the FLT3LG 3’UTR construct resulted in significantly decreased luciferase activity (fc= -2.6, p =.002); while expression of miR-181c with the MCL1 3’UTR construct resulted in significantly decreased luciferase activity (fc = 1.3, p =.002). These studies provide evidence that GATA2 regulates expression of miR-424 and miR-181c, resulting in altered levels of FLT3LG and MCL1.

Scientific Focus Area: Cancer Biology

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