Key Lab of Pediatrics Hematology/Oncology, Ministry of Health, Department of Hematology/Oncology, Shanghai Children's Medical Center, Shanghai Jiao Tong University, Shanghai, China.
Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou, China.
Haematologica. 2020 Apr;105(4):925-936. doi: 10.3324/haematol.2018.210286. Epub 2019 Aug 29.
Hematopoiesis is a finely regulated process in vertebrates under both homeostatic and stress conditions. By whole exome sequencing, we studied the genomics of acute lymphoblastic leukemia (ALL) patients who needed multiple red blood cell (RBC) transfusions after intensive chemotherapy treatment. , encoding a RhoA guanine nucleotide exchange factor, was found to be associated with chemotherapy-induced anemia by genome-wide association study analyses. A single nucleotide polymorphism (SNP) of located in an intron predicted to be a GATA1 binding site, rs10892563, is significantly associated with patients who need RBC transfusion (=3.469E-03, odds ratio 5.864). A luciferase reporter assay revealed that this SNP impairs GATA1-mediated trans-regulation of , and quantitative polymerase chain reaction studies confirmed that the homozygotes status is associated with an approximately 61% reduction in expression (=0.0088). Consequently, erythropoiesis was affected at the pro-erythroblast phases. The role of and its homologs in erythroid differentiation was confirmed in human K562 cells, mouse 32D cells and primary murine bone marrow cells. We further demonstrated in zebrafish by morpholino-mediated knockdown and CRISPR/Cas9-mediated knockout of that its reduction resulted in erythropoiesis defects. The p38 kinase pathway was affected by the ARHGEF12-RhoA signaling in K562 cells, and consistently, the Arhgef12-RhoA-p38 pathway was also shown to be important for erythroid differentiation in zebrafish as active RhoA or p38 readily rescued the impaired erythropoiesis caused by knockdown. Finally, ARHGEF12-mediated p38 activity also appeared to be involved in phenotypes of patients of the rs10892563 homozygous genotype. Our findings present a novel SNP of that may involve ARHGEF12-RhoA-p38 signaling in erythroid regeneration in ALL patients after chemotherapy.
造血是脊椎动物在稳态和应激条件下精细调节的过程。通过全外显子组测序，我们研究了需要在强化化疗后多次输注红细胞（RBC）的急性淋巴细胞白血病（ALL）患者的基因组学。通过全基因组关联研究分析，发现编码 RhoA 鸟嘌呤核苷酸交换因子的 与化疗诱导的贫血相关。位于预测为 GATA1 结合位点的内含子中的 单核苷酸多态性（SNP）rs10892563 与需要输血的患者显着相关（=3.469E-03，优势比 5.864）。荧光素酶报告基因测定显示该 SNP 可损害 GATA1 对 的中介转录调控，并且定量聚合酶链反应研究证实纯合子状态与 表达降低约 61%（=0.0088）相关。因此，红系前体细胞阶段的红细胞生成受到影响。在人类 K562 细胞、小鼠 32D 细胞和原代小鼠骨髓细胞中证实了 和其同源物在红细胞分化中的作用。我们通过 ARHGEF12 的形态发生素介导的敲低和 CRISPR/Cas9 介导的 KO 进一步在斑马鱼中证明了其减少导致红细胞生成缺陷。在 K562 细胞中，p38 激酶途径受 ARHGEF12-RhoA 信号的影响，一致地，Arhgef12-RhoA-p38 途径也被证明对斑马鱼中的红细胞分化很重要，因为活性 RhoA 或 p38 容易挽救由 敲低引起的受损的红细胞生成。最后，ARHGEF12 介导的 p38 活性似乎也与 rs10892563 纯合基因型患者的表型有关。我们的发现提出了一种新的 SNP，即 ，该 SNP 可能涉及 ALL 患者化疗后红细胞再生中的 ARHGEF12-RhoA-p38 信号。
