The role of cytidine deaminase and GATA1 mutations in the increased cytosine arabinoside sensitivity of Down syndrome myeloblasts and leukemia cell lines
The role of cytidine deaminase and GATA1 mutations in the increased cytosine arabinoside sensitivity of Down syndrome myeloblasts and leukemia cell lines
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DOI：
10.1158/0008-5472.can-03-2456
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发表时间：
2004-01-15
期刊：
影响因子：
11.2
通讯作者：
Taub, JW
中科院分区：
文献类型：
--
作者：
Ge, YB;Jensen, TL;Taub, JW
患有急性髓性白血病（AML）的唐氏综合征（DS）儿童的成髓细胞在体外对1-β-D-阿拉伯呋喃糖基胞嘧啶（ara-C）的敏感性显著高于非DS AML成髓细胞，并产生更高的1-β-D-阿拉伯呋喃糖基胞嘧啶5 '-三磷酸（ara-CTP）。半定量逆转录-PCR分析表明，胞苷脱氨酶（CDA）的成绩单DS比非DS成髓细胞低2.7倍。与此相反，胱硫醚-β-合酶和脱氧胞苷激酶的转录是一个中位数12.5倍和2.6倍高DS相比，非DS成髓细胞。脱氧胞苷激酶/ CDA转录本的比率与ara-C敏感性和ara-CTP生成显著相关。在临床相关AML细胞系模型中，DS CMK细胞中的高胱硫醚-β-合酶转录物伴随着比非DS CMS细胞高10倍的ara-C敏感性和高2.4倍的ara-CTP水平。非DS THP-1细胞中CDA的过表达与ara-C敏感性降低100倍和ara-CTP产生降低40倍相关。THP-1细胞将CDA分泌到培养液中，并在30 min内将胞外阿糖胞苷完全转化为1-β-D-阿拉伯呋喃糖基尿嘧啶。5 '-cDNA末端快速扩增（5'-RACE）和逆转录PCR检测鉴定了THP-1细胞中具有不同5'非翻译区和翻译起始位点的短型（sf）和长型（lf）CDA转录本;然而，只有后者导致主动CDA。尽管两种CDA转录物的5'侧翼序列在报告基因测定中表现出启动子活性，但CDAlf的活性较低。在CDAsf启动子中存在几个GATA 1结合位点，并且在DS巨核细胞白血病中一致检测到GATA 1突变，这表明GATA 1在调节CDA转录中的潜在作用，并且CDAsf启动子充当增强子。将GATA 1转染到果蝇Mel-2细胞中以剂量依赖性方式刺激CDAlf启动子。进一步鉴定DS和非DS成髓细胞之间阿糖胞苷代谢相关酶编码基因的差异表达机制可能会改善AML治疗。
Myeloblasts from Down syndrome (DS) children with acute myeloid leukemia (AML) are significantly more sensitive in vitro to 1-beta-D-arabinofuranosylcytosine (ara-C) and generate higher 1-beta-D-arabinofuranosylcytosine 5'-triphosphate (ara-CTP) than non-DS AML myeloblasts. Semi-quantitative reverse transcription-PCR analyses demonstrated that transcripts for cytidine deaminase (CDA) were 2.7-fold lower in DS than for non-DS myeloblasts. In contrast, transcripts of cystathionine-beta-synthase and deoxycytidine kinase were a median 12.5- and 2.6-fold higher in DS compared with non-DS myeloblasts. The ratio of deoxycytidine kinase/ CDA transcripts significantly correlated with ara-C sensitivities and ara-CTP generation. In clinically relevant AML cell line models, high cystathionine-beta-synthase transcripts in DS CMK cells were accompanied by 10-fold greater ara-C sensitivity and 2.4-fold higher levels of ara-CTP compared with non-DS CMS cells. Overexpression of CDA in non-DS THP-1 cells was associated with a 100-fold decreased ara-C sensitivity and 40-fold decreased ara-CTP generation. THP-1 cells secreted CDA into the incubation media and converted extracellular ara-C completely to 1-beta-D-arabinofuranosyluracil within 30 min. Rapid amplification of 5'-cDNA ends (5'-RACE) and reverse transcription-PCR assays identified short(sf) and long-form (lf) CDA transcripts in THP-1 cells with different 5' untranslated regions and translational start sites; however, only the latter resulted in the active CDA. Although 5' flanking sequences for both CDA transcripts exhibited promoter activity in reporter gene assays, activity for the CDAlf was low. The presence of several GATA1 binding sites in the CDAsf promoter and the uniform detection of GATA1 mutations in DS megakaryocytic leukemia suggested the potential role of GATA1 in regulating CDA transcription and the CDAsf promoter acting as an enhancer. Transfection of GATA1 into Drosophila Mel-2 cells stimulated the CDAlf promoter in a dose-dependent fashion. Additional identification of the mechanisms of differential expression of genes encoding enzymes involved in ara-C metabolism between DS and non-DS myeloblasts may lead to improvements in AML therapy.