Aberrant splicing of U12-type introns is the hallmark of ZRSR2 mutant myelodysplastic syndrome

Somatic mutations in the spliceosome gene ZRSR2— located on the X chromosome — are associated with myelodysplastic syndrome (MDS). ZRSR2 is involved in the recognition of 3΄ splice site during the early stages of spliceosome assembly; however, its precise role in RNA splicing has remained unclear. Here, we characterize ZRSR2 as an essential component of the minor spliceosome (U12-dependent) assembly. shRNA mediated knockdown of ZRSR2 leads to impaired splicing of the U12-type introns, and RNA-Sequencing of MDS bone marrow reveals that loss of ZRSR2 activity causes increased mis-splicing. These splicing defects involve retention of the U12-type introns while splicing of the U2-type introns remain mostly unaffected. ZRSR2 deficient cells also exhibit reduced proliferation potential and distinct alterations in myeloid and erythroid differentiation in vitro. These data identify a specific role for ZRSR2 in RNA splicing and highlight dysregulated splicing of U12-type introns as a characteristic feature of ZRSR2 mutations in MDS. RNA sequencing was performed on 16 bone marrow samples (MDS and normal) and six samples of control or ZRSR2 shRNA transduced TF-1 cells and data was analysed for aberrant splicing caused by ZRSR2 mutations/deficiency.

位于X染色体的剪接体（spliceosome）基因ZRSR2的体细胞突变（somatic mutation）与骨髓增生异常综合征（myelodysplastic syndrome, MDS）相关。ZRSR2参与剪接体组装早期阶段的3'剪接位点（3' splice site）识别，但其在RNA剪接（RNA splicing）中的具体功能仍未明确。本研究将ZRSR2鉴定为次要剪接体（minor spliceosome，U12依赖型）组装的必需组分。shRNA介导的ZRSR2基因敲低会导致U12型内含子（U12-type introns）剪接受损；对MDS患者骨髓样本的RNA测序（RNA-Sequencing）分析显示，ZRSR2活性缺失会引发异常剪接事件增多。此类剪接缺陷表现为U12型内含子滞留，而U2型内含子（U2-type introns）的剪接基本不受影响。体外实验中，ZRSR2缺陷细胞还表现出增殖能力下降，以及髓系与红系分化的显著改变。本研究数据明确了ZRSR2在RNA剪接中的特异性功能，并指出U12型内含子剪接失调是MDS患者中ZRSR2突变的标志性特征。本研究对16份骨髓样本（包含MDS患者样本与正常对照样本）以及6份对照或ZRSR2 shRNA转染的TF-1细胞样本进行了RNA测序，并针对ZRSR2突变/缺陷引发的异常剪接展开数据分析。