Cecchini Sylvain, Masson Christel, La Madeleine Carole, Huels Michael A, Sanche Léon, Wagner J Richard, Hunting Darel J
Center for Research in Radiology (CR2), Department of Nuclear Medicine and Radiobiology, Faculty of Medicine, Université de Sherbrooke, Québec, Canada J1H 5N4.
Biochemistry. 2005 Dec 27;44(51):16957-66. doi: 10.1021/bi050799x.
DNA interstrand cross-links (ICL) can be induced both by natural products (e.g., psoralens with UVA) and by chemical agents, some of which are used in chemotherapy (e.g., Carboplatin and mitomycin C). Here, we report the formation of ICL by UV radiation in brominated DNA, but only for very specific conformations. The quantum yields for strand break and cross-link formation depend on the wavelength with a maximum near 280 nm. It is known that the photosensitization of DNA by bromodeoxyuridine (BrdUrd) results mainly from the electron affinity of bromine, leading to the irreversible formation of 2'-deoxyuridin-5-yl radicals (dUrd*) upon the addition of an electron from an adjacent adenosine. It is well documented that the photolytic loss of the bromine atom is greatly suppressed in single-stranded DNA versus that in double-stranded DNA. To study this behavior, we have used two models of BrdUrd-mediated sensitization: one consists of a DNA duplex containing a bulge, formed by five mismatched bases, including the BrdUrd, and the other consists of completely duplex DNA. UV irradiation induces much higher levels of single-strand breaks (ssb) in the completely duplex DNA at the BrdUrd site compared to the DNA with a bulge. However, in completely duplex DNA, ssb appear only in the brominated strand, whereas in the bulged duplex DNA, ssb occur on both strands. Most importantly, we also observe formation of interstrand cross-links in bulged duplex DNA in the BrdUrd region. Thus, we propose that UV irradiation of cells containing BrdUrd incorporated randomly into duplex DNA will create many ssb, whereas BrdUrd present in DNA bulges or open regions in double-stranded DNA (transcription bubbles, replication forks) will lead to potentially lethal damage in both strands in the form of ICL. These findings may help explain the potent clinical antiviral activity of IdUrd and BrdUrd (e.g., IdUrd is used to treat eye infections caused by the herpes virus) and suggest that ICL formation may be a very specific probe for identifying single-stranded regions in the DNA of living cells. In addition, this model system provides an excellent means of introducing ICL for studies on their repair and biological consequences.
DNA链间交联（ICL）可由天然产物（如补骨脂素与紫外线A）和化学试剂诱导产生，其中一些化学试剂用于化疗（如卡铂和丝裂霉素C）。在此，我们报道了紫外线辐射在溴化DNA中诱导形成ICL，但仅针对非常特定的构象。链断裂和交联形成的量子产率取决于波长，在280nm附近有最大值。已知溴脱氧尿苷（BrdUrd）对DNA的光敏化主要源于溴的电子亲和力，在从相邻腺苷添加一个电子后导致2'-脱氧尿苷-5-基自由基（dUrd*）的不可逆形成。有充分文献记载，与双链DNA相比，单链DNA中溴原子的光解损失受到极大抑制。为研究这种行为，我们使用了两种BrdUrd介导的光敏化模型：一种由包含一个凸起的DNA双链体组成，该凸起由五个错配碱基形成，包括BrdUrd，另一种由完全双链的DNA组成。与有凸起的DNA相比，紫外线照射在完全双链DNA的BrdUrd位点诱导产生的单链断裂（ssb）水平要高得多。然而，在完全双链DNA中，ssb仅出现在溴化链中，而在有凸起的双链DNA中，两条链上都会出现ssb。最重要的是，我们还观察到在有凸起的双链DNA的BrdUrd区域形成了链间交联。因此，我们提出，紫外线照射随机掺入双链DNA中的含有BrdUrd的细胞会产生许多ssb，而存在于双链DNA的凸起或开放区域（转录泡、复制叉）中的BrdUrd将以ICL的形式导致两条链上潜在的致死性损伤。这些发现可能有助于解释碘苷和溴脱氧尿苷强大的临床抗病毒活性（如碘苷用于治疗由疱疹病毒引起的眼部感染），并表明ICL的形成可能是识别活细胞DNA中单链区域的一种非常特异的探针。此外，这个模型系统为引入ICL以研究其修复和生物学后果提供了一个极好的手段。
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