PDF(Pubmed)
Abstract：
Lethal toxin (LT) is the critical virulence factor of Bacillus anthracis, the causative agent of anthrax. One common symptom observed in patients with anthrax is thrombocytopenia, which has also been observed in mice injected with LT. Our previous study demonstrated that LT induces thrombocytopenia by suppressing megakaryopoiesis, but the precise molecular mechanisms behind this phenomenon remain unknown. In this study, we utilized 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced megakaryocytic differentiation in human erythroleukemia (HEL) cells to identify genes involved in LT-induced megakaryocytic suppression. Through cDNA microarray analysis, we identified Dachshund homolog 1 (DACH1) as a gene that was upregulated upon TPA treatment but downregulated in the presence of TPA and LT, purified from the culture supernatants of B. anthracis. To investigate the function of DACH1 in megakaryocytic differentiation, we employed short hairpin RNA technology to knock down DACH1 expression in HEL cells and assessed its effect on differentiation. Our data revealed that the knockdown of DACH1 expression suppressed megakaryocytic differentiation, particularly in polyploidization. We demonstrated that one mechanism by which B. anthracis LT induces suppression of polyploidization in HEL cells is through the cleavage of MEK1/2. This cleavage results in the downregulation of the ERK signaling pathway, thereby suppressing DACH1 gene expression and inhibiting polyploidization. Additionally, we found that known megakaryopoiesis-related genes, such as FOSB, ZFP36L1, RUNX1, FLI1, AHR, and GFI1B genes may be positively regulated by DACH1. Furthermore, we observed an upregulation of DACH1 during in vitro differentiation of CD34-megakaryocytes and downregulation of DACH1 in patients with thrombocytopenia. In summary, our findings shed light on one of the molecular mechanisms behind LT-induced thrombocytopenia and unveil a previously unknown role for DACH1 in megakaryopoiesis.
摘要：
致死毒素(LT)是炭疽芽孢杆菌的关键毒力因子,炭疽病的病原体.在炭疽患者中观察到的一个常见症状是血小板减少症，在注射LT的小鼠中也观察到了这一点。我们先前的研究表明，LT通过抑制巨核细胞生成诱导血小板减少症，但是这种现象背后的确切分子机制仍然未知。在这项研究中,我们利用12-O-十四烷酰基佛波醇-13-乙酸盐(TPA)诱导的人红白血病(HEL)细胞巨核细胞分化来鉴定参与LT诱导的巨核细胞抑制的基因.通过cDNA微阵列分析，我们确定了腊肠同源物1（DACH1）是在TPA处理后上调但在TPA和LT存在下下调的基因，从炭疽芽孢杆菌的培养上清液中纯化。探讨DACH1在巨核细胞分化中的作用,我们采用短发夹RNA技术在HEL细胞中敲低DACH1表达并评估其对分化的影响。我们的数据显示，DACH1表达的敲低抑制了巨核细胞的分化，特别是在多倍体化方面。我们证明了炭疽芽孢杆菌LT诱导HEL细胞多倍体化抑制的一种机制是通过MEK1/2的裂解。这种切割导致ERK信号通路的下调，从而抑制DACH1基因表达并抑制多倍体化。此外,我们发现已知的巨核细胞生成相关基因，比如FOSB,ZFP36L1,RUNX1,FLI1,AHR,和GFI1B基因可能受DACH1正调控。此外，我们观察到血小板减少症患者在CD34巨核细胞体外分化过程中DACH1的上调和DACH1的下调。总之,我们的研究结果揭示了LT诱导的血小板减少症背后的一个分子机制,并揭示了DACH1在巨核细胞生成中的一个以前未知的作用.
