JAS:北大要茂盛和罗春雄课题组发展了基于单活体酵母菌实时在线

近日，国际知名学术期刊《JouRNAl of Aerosol science》在线发表了北京大学环境科学与工程学院要茂盛研究员、物理学院罗春雄副教授研究组合作的一篇研究论文，论文报道了研究组利用空气采样、微流控、荧光蛋白标记的酵母菌(Saccharomyces cerevisiae)以及单酵母菌蛋白荧光自动检测平台创建了大气PM2.5毒性实时在线监测系统-Single Living yEast PM Toxicity Sensor (SLEPTor-沉睡者)，能够对PM2.5多方面的毒性进行实时监测。论文的第一作者为环境学院的博士生魏恺，要茂盛、罗春雄为共同通讯作者。

空气污染特别是PM2.5成为当今人类面临的重要的环境问题之一。然而，目前PM2.5毒性的研究大多采用离线的方式，时效性差;而细胞染毒或动物暴露实验常常用的是高剂量、 灵敏度低且偏离实际环境暴露剂量。

课题组前期利用创建的SLEPTor系统对酵母菌的60多种蛋白进行了高通量的筛查，发现了HSP60、SSA1(分别为氧化损伤蛋白、DNA修复蛋白)对空气PM2.5响应比较灵敏，并且在单个活体酵母菌上发现DNA的修复发生在氧化损伤约20分钟后。Saccharomyces cerevisiae俗称酿酒酵母，繁殖快，其基因序列于1996年测序完成，为第一个完成基因测序的真核生物，被广泛地应用在人类疾病研究中，许多生命现象及蛋白的功能来自该酵母菌的研究。课题组利用S. cerevisiae研发的SLEPTor系统可通过同时监测不同标记荧光蛋白的表达来展现酵母菌整个生命体系从不同层面对颗粒物的实时响应，这为颗粒物对人体的健康效应机制的研究提供了类似于实时监测不同地区车辆行驶状况的开创性的研究思路和方法，从而有助于避免“盲人摸象”的弊端。

目前课题组正在利用该体系对不同国家、地区颗粒物的毒性进行研究，同时也在筛查更多的有响应的酵母菌蛋白，并研究其灵敏度、响应的毒性标定。课题组文章得到了审稿人的高度认可(“I think the method developed by the authors is highly promising and represents a significant multidisciplinary contribution combining aerosol, biology and toxicity.”)。

原文链接：

Single Living yEast PM Toxicity Sensor (SLEPTor) System

原文摘要：

Air pollution has become one of major environmental challenges facing mankind in this century. However, current atmospheric toxicity studies are most offline with low sensitivity and limited understanding of collective biological responses to ambient stimuli on the single cell level. Here, we developed an online analysis system named as SLEPTor (Single Living yEast PM Toxicity Sensor) which allows us to analyze aerosol particle toxicity in vivo. The SLEPTor is composed of an automated air sampling, microfluidics, GFP(green fluorescent protein)-tagged Saccharomyces cerevisiae yeast sensor as well as multiplexing automated fluorescence imaging system. In testing the SLEPTor, 63 yeast genes were screened simultaneously here for their responses to both man-made reactive oxygen species (ROS) and also Beijing aerosol samples. For ROS samples, MRS3, MRS4, BSD2 and ZRC1 proteins were detected in increased abundances, and the BSD2 protein expression, related to oxidative stress, was more pronounced, followed by MRS4 (for DNA repair) with a peak expression after 20 min exposure. In contrast, HSP60 (oxidative stress), SSA1 (DNA repair) and MSB1 (MAPK pathway) proteins were observed to have elevated expressions visualized with time-lapsed movies when challenged with Beijing air samples (150 µg/m3) for 100 min. The same experiments were repeated twice involving 63 genes using the SLEPTor and further confirmed by a spectra method. Among the screened proteins, SSA1 was found to be sensitive to ambient air. Using the SLEPTor developed, we have shown that PM exposure resulted in oxidative stress and DNA repair on living yeast cells. The demonstrated proof-of-concept technology here pioneers a new arena for investigating health impacts of aerosol particles in an interactive manner. Yet, its sensitivity and repeatability need to be further explored using a large set of different functional proteins.

doi:10.1016/j.jaerosci.2017.02.006

作者：要茂盛和罗春雄