Hall Megan R, Gracey Andrew Y
Department of Biological Sciences, University of Southern California, Los Angeles, CA, United States.
Front Physiol. 2021 Dec 9;12:647482. doi: 10.3389/fphys.2021.647482. eCollection 2021.
One of the challenges facing efforts to generate molecular biomarkers for toxins is distinguishing between markers that are indicative of exposure and markers that provide evidence of the effects of toxicity. Phenotypic anchoring provides an approach to help segregate markers into these categories based on some phenotypic index of toxicity. Here we leveraged the mussel embryo-larval toxicity assay in which toxicity is estimated by the fraction of larvae that exhibit an abnormal morphology, to isolate subsets of larvae that were abnormal and thus showed evidence of copper-toxicity, versus others that while exposed to copper exhibited normal morphology. Mussel larvae reared under control conditions or in the presence of increasing levels of copper (3-15 μg/L Cu) were physically sorted according to whether their morphology was normal or abnormal, and then profiled using RNAseq. Supervised differential expression analysis identified sets of genes whose differential expression was specific to the pools of abnormal larvae versus normal larvae, providing putative markers of copper toxicity versus exposure. Markers of copper exposure and copper-induced abnormality were involved in many of the same pathways, including development, shell formation, cell adhesion, and oxidative stress, yet unique markers were detected in each gene set. Markers of effect appeared to be more resolving between phenotypes at the lower copper concentration, while markers of exposure were informative at both copper concentrations.
生成毒素分子生物标志物的努力面临的挑战之一是区分指示暴露的标志物和提供毒性作用证据的标志物。表型锚定提供了一种基于某种毒性表型指标将标志物分为这些类别的方法。在这里，我们利用贻贝胚胎-幼虫毒性试验，其中毒性通过表现出异常形态的幼虫比例来估计，以分离出形态异常从而显示铜毒性证据的幼虫亚群，与其他虽暴露于铜但形态正常的幼虫进行对比。将在对照条件下或在铜水平不断增加（3 - 15μg/L铜）的环境中饲养的贻贝幼虫，根据其形态正常与否进行物理分类，然后使用RNA测序进行分析。监督差异表达分析确定了那些差异表达在异常幼虫组与正常幼虫组中具有特异性的基因集，从而提供了铜毒性与暴露的推定标志物。铜暴露和铜诱导异常的标志物涉及许多相同的途径，包括发育、壳形成、细胞粘附和氧化应激，但在每个基因集中都检测到了独特的标志物。在较低铜浓度下，效应标志物在区分表型方面似乎更具分辨力，而暴露标志物在两种铜浓度下都具有信息价值。
