Artificial and Solar UV Radiation Induces Strand Breaks and Cyclobutane Pyrimidine Dimers in Bacillus subtilis Spore DNA

The loss of stratospheric ozone and the accompanying increase in solar UV flux have led to concerns regarding decreases in global microbial productivity. Central to understanding this process is determining the types and amounts of DNA damage in microbes caused by solar UV irradiation. While UV irradiation of dormant Bacillus subtilis endospores results mainly in formation of the “spore photoproduct” 5-thyminyl-5,6-dihydrothymine, genetic evidence indicates that an additional DNA photoproduct(s) may be formed in spores exposed to solar UV-B and UV-A radiation (Y. Xue and W. L. Nicholson, Appl. Environ. Microbiol. 62:2221–2227, 1996). We examined the occurrence of double-strand breaks, single-strand breaks, cyclobutane pyrimidine dimers, and apurinic-apyrimidinic sites in spore DNA under several UV irradiation conditions by using enzymatic probes and neutral or alkaline agarose gel electrophoresis. DNA from spores irradiated with artificial 254-nm UV-C radiation accumulated single-strand breaks, double-strand breaks, and cyclobutane pyrimidine dimers, while DNA from spores exposed to artificial UV-B radiation (wavelengths, 290 to 310 nm) accumulated only cyclobutane pyrimidine dimers. DNA from spores exposed to full-spectrum sunlight (UV-B and UV-A radiation) accumulated single-strand breaks, double-strand breaks, and cyclobutane pyrimidine dimers, whereas DNA from spores exposed to sunlight from which the UV-B component had been removed with a filter (“UV-A sunlight”) accumulated only single-strand breaks and double-strand breaks. Apurinic-apyrimidinic sites were not detected in spore DNA under any of the irradiation conditions used. Our data indicate that there is a complex spectrum of UV photoproducts in DNA of bacterial spores exposed to solar UV irradiation in the environment.

平流层臭氧（stratospheric ozone）损耗与伴随的太阳紫外辐射通量（solar UV flux）升高，引发了学界对全球微生物生产力（microbial productivity）下降的广泛担忧。阐明这一过程的核心环节，在于明确太阳紫外辐照导致的微生物DNA损伤类型与负荷。针对休眠枯草芽孢杆菌（Bacillus subtilis）芽孢的人工紫外辐照研究显示，其主要会形成“芽孢光产物（spore photoproduct）”5-胸腺嘧啶基-5,6-二氢胸腺嘧啶（5-thyminyl-5,6-dihydrothymine）；但遗传学研究证据表明，当芽孢暴露于太阳UV-B与UV-A辐射时，可能会形成额外的DNA光产物（Y. Xue与W. L. Nicholson, Appl. Environ. Microbiol. 62:2221–2227, 1996）。本研究借助酶促探针技术结合中性或碱性琼脂糖凝胶电泳（agarose gel electrophoresis），在多种紫外辐照条件下检测了芽孢DNA中的双链断裂（double-strand breaks）、单链断裂（single-strand breaks）、环丁烷嘧啶二聚体（cyclobutane pyrimidine dimers）以及无嘌呤-无嘧啶位点（apurinic-apyrimidinic sites）。经人工254 nm UV-C辐射辐照的芽孢DNA，会积累单链断裂、双链断裂与环丁烷嘧啶二聚体；而经人工UV-B辐射（波长290至310 nm）处理的芽孢DNA，仅积累环丁烷嘧啶二聚体。暴露于全光谱太阳光（包含UV-B与UV-A辐射）的芽孢DNA，会同时积累单链断裂、双链断裂与环丁烷嘧啶二聚体；而经滤除UV-B组分的太阳光（“UV-A太阳光”）辐照的芽孢DNA，仅存在单链断裂与双链断裂的积累。在所有测试的辐照条件下，芽孢DNA中均未检测到无嘌呤-无嘧啶位点。本研究数据表明，环境中暴露于太阳紫外辐照的细菌芽孢DNA，其紫外光产物谱具有复杂多样的特征。