Thambyrajah Roshana, Ucanok Deniz, Jalali Maryam, Hough Yasmin, Wilkinson Robert Neil, McMahon Kathryn, Moore Chris, Gering Martin
School of Life Sciences, University of Nottingham, Queen's Medical Centre, Nottingham NG7 2UH, UK.
Department of Infection, Immunity & Cardiovascular Disease, University of Sheffield, Medical School, Beech Hill Road, Sheffield S10 2RX, UK; Bateson Centre, University of Sheffield, Firth Court, Western Bank, Sheffield S10 2TN, UK.
Dev Biol. 2016 Sep 1;417(1):25-39. doi: 10.1016/j.ydbio.2016.07.010. Epub 2016 Jul 16.
A transposon-mediated gene trap screen identified the zebrafish line qmc551 that expresses a GFP reporter in primitive erythrocytes and also in haemogenic endothelial cells, which give rise to haematopoietic stem and progenitor cells (HSPCs) that seed sites of larval and adult haematopoiesis. The transposon that mediates this GFP expression is located in intron 1 of the gfi1aa gene, one of three zebrafish paralogs that encode transcriptional repressors homologous to mammalian Gfi1 and Gfi1b proteins. In qmc551 transgenics, GFP expression is under the control of the endogenous gfi1aa promoter, recapitulates early gfi1aa expression and allows live observation of gfi1aa promoter activity. While the transposon integration interferes with the expression of gfi1aa mRNA in haematopoietic cells, homozygous qmc551 fish are viable and fertile, and display normal primitive and definitive haematopoiesis. Retained expression of Gfi1b in primitive erythrocytes and up-regulation of Gfi1ab at the onset of definitive haematopoiesis in homozygous qmc551 carriers, are sufficient to allow normal haematopoiesis. This finding contradicts previously published morpholino data that suggested an essential role for zebrafish Gfi1aa in primitive erythropoiesis.
一项转座子介导的基因捕获筛选鉴定出斑马鱼品系qmc551,其在原始红细胞以及造血内皮细胞中表达绿色荧光蛋白(GFP)报告基因,造血内皮细胞可产生造血干细胞和祖细胞(HSPCs),这些细胞可在幼虫和成年造血位点定植。介导这种GFP表达的转座子位于gfi1aa基因的内含子1中,gfi1aa基因是斑马鱼三个旁系同源基因之一,编码与哺乳动物Gfi1和Gfi1b蛋白同源的转录抑制因子。在qmc551转基因鱼中,GFP表达受内源性gfi1aa启动子控制,概括了早期gfi1aa表达,并允许对gfi1aa启动子活性进行实时观察。虽然转座子整合干扰了造血细胞中gfi1aa mRNA的表达,但纯合qmc551鱼是可行且可育的,并表现出正常的原始造血和定型造血。在纯合qmc551携带者中,原始红细胞中Gfi1b的持续表达以及定型造血开始时Gfi1ab的上调,足以实现正常造血。这一发现与先前发表的吗啉代数据相矛盾,后者表明斑马鱼Gfi1aa在原始红细胞生成中起重要作用。