Fuglerud Bettina M, Lemma Roza B, Wanichawan Pimthanya, Sundaram Arvind Y M, Eskeland Ragnhild, Gabrielsen Odd S
Department of Biosciences, University of Oslo, P.O.Box 1066 Blindern, N-0316 Oslo, Norway.
Institute for Experimental Medical Research, Oslo University Hospital and University of Oslo, P.O.Box 4956 Nydalen, N-0424 Oslo, Norway.
Nucleic Acids Res. 2017 Jul 27;45(13):7681-7696. doi: 10.1093/nar/gkx364.
The transcription factor c-Myb is involved in early differentiation and proliferation of haematopoietic cells, where it operates as a regulator of self-renewal and multi-lineage differentiation. Deregulated c-Myb plays critical roles in leukaemias and other human cancers. Due to its role as a master regulator, we hypothesized it might function as a pioneer transcription factor. Our approach to test this was to analyse a mutant of c-Myb, D152V, previously reported to cause haematopoietic defects in mice by an unknown mechanism. Our transcriptome data from K562 cells indicates that this mutation specifically affects c-Myb's ability to regulate genes involved in differentiation, causing failure in c-Myb's ability to block differentiation. Furthermore, we see a major effect of this mutation in assays where chromatin opening is involved. We show that each repeat in the minimal DNA-binding domain of c-Myb binds to histones and that D152V disrupts histone binding of the third repeat. ATAC-seq data indicates this mutation impairs the ability of c-Myb to cause chromatin opening at specific sites. Taken together, our findings support that c-Myb acts as a pioneer factor and show that D152V impairs this function. The D152V mutant is the first mutant of a transcription factor specifically destroying pioneer factor function.
转录因子c-Myb参与造血细胞的早期分化和增殖，在其中作为自我更新和多谱系分化的调节因子发挥作用。c-Myb失调在白血病和其他人类癌症中起关键作用。由于其作为主要调节因子的作用，我们推测它可能作为先驱转录因子发挥功能。我们测试这一推测的方法是分析c-Myb的一个突变体D152V，此前报道该突变体通过未知机制在小鼠中导致造血缺陷。我们来自K562细胞的转录组数据表明，这种突变特异性地影响c-Myb调节参与分化的基因的能力，导致c-Myb阻断分化的能力失效。此外，在涉及染色质开放的实验中，我们看到了这种突变的主要影响。我们表明，c-Myb最小DNA结合域中的每个重复序列都与组蛋白结合，并且D152V破坏了第三个重复序列与组蛋白的结合。ATAC-seq数据表明，这种突变损害了c-Myb在特定位点引起染色质开放的能力。综上所述，我们的研究结果支持c-Myb作为先驱因子发挥作用，并表明D152V损害了这一功能。D152V突变体是第一个特异性破坏先驱因子功能的转录因子突变体。
