Loss of H3K27me3 imprinting in the Sfmbt2 miRNA cluster causes enlargement of cloned mouse placentas. Loss of H3K27me3 imprinting in the Sfmbt2 miRNA cluster causes enlargement of cloned mouse placentas

Abnormal placentation in cloned animals remains an unsolved problem. We demonstrated the involvement of micro RNAs (miRNAs) in the abnormal enlargement (hyperplasia) of placentas in cloned mice. Using a comparative transcriptome analysis of cloned placentas, we noted the consistent upregulation of clustered miRNAs within Sfmbt2, a paternally expressed imprinted gene. This region was biallelically activated by loss of imprinting (LOI) in cloned placentas. Deletion of the maternal allele of the whole miRNA cluster resulted in the correction of their expression levels and upregulation of their putative target genes with antitumor or apoptotic functions. Consequently, the placental size was reduced to the normal level and histology was ameliorated. In contrast, correcting the expression of the LOI genes (Sfmbt2, Gab1, and Scl38a4) in cloned placentas had no impact on placental size. Thus, we identified that LOI of clustered miRNAs within Sfmbt2 in cloned placentas was the major cause of abnormal placental enlargement. Overall design: Mouse total RNAs were collected from in vitro fertilized and somatic cell nuclear transferred placentas at E11.5 and their miRNAs were hybridized with Agilent microarray.

克隆动物的胎盘异常发育仍是尚未解决的科学难题。我们证实了微小RNA（micro RNAs, miRNAs）参与克隆小鼠胎盘的异常增生（hyperplasia）过程。通过对克隆胎盘开展比较转录组分析，我们发现位于父本表达印记基因Sfmbt2内的成簇miRNAs呈现持续上调表达。该区域在克隆胎盘中因印记丢失（loss of imprinting, LOI）发生双等位基因激活。删除整个miRNA簇的母本等位基因后，这些miRNAs的表达水平得以矫正，其推定的具有抗肿瘤或凋亡功能的靶基因表达也同步上调。最终，胎盘尺寸恢复至正常水平，组织学异常亦得到改善。与之相反，矫正克隆胎盘中LOI相关基因（Sfmbt2、Gab1和Scl38a4）的表达，对胎盘尺寸并无影响。因此，我们确定克隆胎盘中Sfmbt2基因座内成簇miRNAs的印记丢失是导致胎盘异常增大的主要原因。整体实验设计：于胚胎发育第11.5天（E11.5）采集体外受精及体细胞核移植胎盘的总RNA，采用安捷伦（Agilent）微阵列芯片对其miRNAs进行杂交检测。