Expression of Let-7 Micro RNAs in Transgenic Mice Overexpressing Truncated Hmga2

Abstract 3470

Expression of HMGA2 is negatively regulated by binding of let-7-family micro RNAs (miRNAs) to specific sites in 3' untranslated region (UTR) of HMGA2. Either downregulation of let-7 or truncation of HMGA2 is thought to cause overexpression of HMGA2. Overexpression of HMGA2 mRNA due to truncation of its 3'UTR by chromosomal translocation was reported in 2 patients with paroxysmal nocturnal hemoglobinuria (PNH; Inoue et al, Blood, 2006). Accordingly, we recently established transgenic ΔHmga2 mice with 3'UTR-truncated Hmga2 cDNA, which revealed a clonal growth advantage of hematopoietic cells overexpressing Hmga2 mRNA that may lead to expansion of a PNH cell (Ikeda et al, Blood, 2011). It has also been reported that HMGA2 mRNA is overexpressed whereas let-7b and −7c are downregulated in peripheral blood cells from the majority of patients with PNH (Murakami et al, BJH, 2012). Although downregulation of let-7-family miRNAs may in part explain overexpression of HMGA2 mRNA, not only full-length HMGA2 mRNA but also short transcript variants of HMGA2 mRNA without 3'UTR are overexpressed in most of these patients. Thus, we aimed to investigate the influence of overexpression of 3'UTR-truncated Hmga2 mRNA, which mimics its short transcript variant, on expression of let-7-family miRNAs. We evaluated expression of let-7a, -7b, and -7c miRNAs in bone marrow (BM) and spleen cells of 12 week-old ΔHmga2 mice (n= 3–4) and wild-type (WT) mice (n= 3–4) using quantitative real-time RT-PCR. Relative expression levels of let-7a (mean ± SD, 1.14 ± 0.57 and 1.62 ± 1.05, respectively), -7b (0.76 ± 0.38 and 1.14 ± 0.77), and -7c (0.62 ± 0.47 and 0.96 ± 0.51) of BM cells were not significantly different between WT mice and ΔHmga2 mice. However, relative expression level of let-7c was significantly higher in spleen cells of ΔHmga2 mice (11.75 ± 2.10) compared with WT mice (1.47 ± 1.19; P < .01). Expression of let-7a (2.46 ± 2.72 and 20.20 ± 25.22) and −7b (1.43 ± 1.62 and 5.27 ± 4.05) was not significantly different in spleen between WT mice and ΔHmga2 mice. Relative expression levels of let-7c were higher in spleen cells compared with those in BM cells in ΔHmga2 mice (P < .01). Therefore, differences in cell lineage or differentiation status might alter expression of let-7-family miRNAs and 3'UTR-truncated Hmga2 may further influence let-7 expression in a tissue specific manner. However, the cause of downregulation of let-7b and -7c, and overexpression of 3'UTR truncated variants of HMGA2 in patients with PNH remains to be elucidated.