Resistance to arginine deiminase treatment in melanoma cells is associated with induced argininosuccinate synthetase expression involving c-Myc/HIF-1alpha/Sp4.
Resistance to arginine deiminase treatment in melanoma cells is associated with induced argininosuccinate synthetase expression involving c-Myc/HIF-1alpha/Sp4.
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DOI：
10.1158/1535-7163.mct-09-0794
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发表时间：
2009-12
影响因子：
5.7
通讯作者：
Kuo MT
中科院分区：
文献类型：
--
作者：
Tsai WB;Aiba I;Lee SY;Feun L;Savaraj N;Kuo MT
基于精氨酸脱亚胺酶（ADI）的精氨酸去除是一种治疗恶性黑色素瘤的新策略，临床试验具有良好的效果。黑色素瘤对ADI治疗的敏感性是基于其精氨酸营养缺陷型，这是由于缺乏精氨酸琥珀酸合成酶（AS）表达所致，AS是精氨酸从头生物合成的限速酶。我们发现ADI可在黑色素瘤细胞系A2058和SK-MEL-2中转录诱导AS表达，但在A375细胞中不诱导AS表达;并且这种诱导与ADI治疗的抗性相关。AS启动子的近端区域包含一个由c-Myc和HIF-1α识别的E-box和一个由Sp 4识别的GC-box。通过染色质免疫沉淀分析，我们证明了在非诱导条件下，E-box在所有三种黑素瘤细胞系中均与HIF-1α结合。在精氨酸耗竭条件下，A2058和SK-MEL-2细胞中HIF-1α被c-Myc取代，而A375细胞中没有。无论所有三种细胞系中的精氨酸可用性如何，Sp 4均与GC盒组成性结合。通过转染过表达c-Myc上调A2058和SK-MEL-2细胞中AS的表达;而与HIF-1α共转染抑制c-Myc诱导的AS表达。这些结果表明，AS表达的调控涉及转录正调控因子c-Myc和Sp 4以及负调控因子HIF-1α之间的相互作用，HIF-1α赋予A2058和SK-MEL-2细胞对ADI处理的抗性。在精氨酸耗竭条件下，A375细胞中AS诱导的无能与c-Myc与AS启动子相互作用的失败有关。
Arginine deiminase (ADI)-based arginine depletion is a novel strategy under clinical trials for the treatment of malignant melanoma with promising results. The sensitivity of melanoma to ADI treatment is based on its auxotrophy for arginine due to a lack of argininosuccinate synthetase (AS) expression, the rate-limiting enzyme for the de novo biosynthesis of arginine. We show here that AS expression can be transcriptionally induced by ADI in melanoma cell lines A2058 and SK-MEL-2 but not in A375 cells; and this inducibility was correlated with resistance to ADI treatment. The proximal region of the AS promoter contains an E-box that is recognized by c-Myc and HIF-1α and a GC-box by Sp4. By chromatin immunoprecipitation asssays, we demonstrated that under noninduced conditions, the E-box was bound by HIF-1α in all the three melanoma cell lines. Under arginine depletion conditions, HIF-1α was replaced by c-Myc in A2058 and SK-MEL-2 cells but not in A375 cells. Sp4 was constitutively bound to the GC-box regardless of arginine availability in all three cell lines. Overexpressing c-Myc by transfection upregulated AS expression in A2058 and SK-MEL-2 cells; whereas co-transfection with HIF-1α suppressed c-Myc-induced AS expression. These results suggest that regulation of AS expression involves interplay among positive transcriptional regulators c-Myc and Sp4, and negative regulator HIF-1α that confers resistance to ADI treatment in A2058 and SK-MEL-2 cells. Inability of AS induction in A375 cells under arginine depletion conditions was correlated by the failure of c-Myc to interact with the AS promoter.