Expression of Cell-Surface Marker ABCB5 Causes Characteristic Modifications of Glucose, Amino Acid and Phospholipid Metabolism in the G3361 Melanoma-Initiating Cell Line.
Expression of Cell-Surface Marker ABCB5 Causes Characteristic Modifications of Glucose, Amino Acid and Phospholipid Metabolism in the G3361 Melanoma-Initiating Cell Line.
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DOI：
10.1371/journal.pone.0161803
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发表时间：
2016
期刊：
影响因子：
3.7
通讯作者：
Frank MH
中科院分区：
文献类型：
--
作者：
Lutz NW;Banerjee P;Wilson BJ;Ma J;Cozzone PJ;Frank MH
我们提出了一个试点研究，旨在确定ATP结合盒成员B5（ABCB 5），以前描述的黑色素瘤起始细胞的标志物，细胞代谢的表达的影响。比较了两组人G3361黑素瘤细胞的代谢谱，即具有完整ABCB 5表达的野生型黑素瘤细胞（ABCB 5-WT）和通过shRNA介导的基因敲低具有抑制的ABCB 5表达的相应黑素瘤细胞变体（ABCB 5-KD）。通过使用细胞提取物的质子和磷NMR光谱进行全面的代谢组学分析，以检查水溶性代谢物和脂质。绝对和相对代谢物水平的参数和非参数统计分析产生了参与葡萄糖、氨基酸和磷脂（PL）代谢的化合物的显著差异。相比之下，能量代谢几乎不受ABCB 5表达的影响。ABCB 5-WT与ABCB 5-KD G3361变体相比，每个总蛋白的水溶性代谢物总和高17%，但未发现PL总和的差异。乳酸盐（+23%）和丙氨酸（+26%）的丰度增加尤其明显，表明糖酵解和潜在的β-氨基丁酸解增加。PL降解产物甘油磷酸胆碱和甘油磷酸乙醇胺（分别为+85%和123%）的增加以及PL池内的再分布表明ABCB 5表达导致膜PL周转率增加。在野生型G3361黑色素瘤细胞中采用单克隆抗体（mAb）依赖性ABCB 5蛋白抑制的功能研究支持了ABCB 5依赖性IL 1 β介导机制调节糖酵解的可能性。我们的代谢组学结果表明，潜在的生化途径可能为黑色素瘤治疗提供靶点，可能与其他治疗形式相结合。
We present a pilot study aimed at determining the effects of expression of ATP-binding cassette member B5 (ABCB5), a previously described marker for melanoma-initiating cells, on cellular metabolism. Metabolic profiles for two groups of human G3361 melanoma cells were compared, i.e. wildtype melanoma cells with intact ABCB5 expression (ABCB5-WT) and corresponding melanoma cell variants with inhibited ABCB5 expression, through shRNA-mediated gene knockdown (ABCB5-KD). A comprehensive metabolomic analysis was performed by using proton and phosphorus NMR spectroscopy of cell extracts to examine water-soluble metabolites and lipids. Parametric and non-parametric statistical analysis of absolute and relative metabolite levels yielded significant differences for compounds involved in glucose, amino acid and phospholipid (PL) metabolism. By contrast, energy metabolism was virtually unaffected by ABCB5 expression. The sum of water-soluble metabolites per total protein was 17% higher in ABCB5-WT vs. ABCB5-KD G3361 variants, but no difference was found for the sum of PLs. Enhanced abundance was particularly pronounced for lactate (+ 23%) and alanine (+ 26%), suggesting an increase in glycolysis and potentially glutaminolysis. Increases in PL degradation products, glycerophosphocholine and glycerophosphoethanolamine (+ 85 and 123%, respectively), and redistributions within the PL pool suggested enhanced membrane PL turnover as a consequence of ABCB5 expression. The possibility of glycolysis modulation by an ABCB5-dependent IL1β-mediated mechanism was supported by functional studies employing monoclonal antibody (mAb)-dependent ABCB5 protein inhibition in wildtype G3361 melanoma cells. Our metabolomic results suggest that the underlying biochemical pathways may offer targets for melanoma therapy, potentially in combination with other treatment forms.