Castrillo Marta, Bernhardt Adrian, Ávalos Javier, Batschauer Alfred, Pokorny Richard
Department of Genetics, Faculty of Biology, University of Seville, Seville, Spain.
Department of Plant Physiology and Photobiology, Faculty of Biology, Philipps-University, Marburg, Germany.
Photochem Photobiol. 2015 Nov;91(6):1356-67. doi: 10.1111/php.12501. Epub 2015 Sep 11.
Proteins from the cryptochrome/photolyase family utilize UV-A, blue or even red light to achieve such diverse functions as repair of DNA lesions by photolyases and signaling by cryptochromes. DASH-type cryptochromes retained the ability to repair cyclobutane pyrimidine dimers (CPDs) in single-stranded DNA regions in vitro. However, most organisms possess conventional CPD photolyases responsible for repair of these lesions in vivo. Recent work showed that the DASH-type cryptochrome CryD plays a regulatory role in diverse light-dependent processes in Fusarium fujikuroi. Here, we report our in vitro studies on heterologously expressed FfCryD. The purified protein contains N(5) ,N(10) -methenyltetrahydrofolate and flavin adenine dinucleotide as cofactors. Photoreduction and DNA photorepair experiments confirmed that FfCryD is active in light-driven electron transfer processes. However, the protein showed comparable affinities for CPD-comprising and undamaged DNA probes. Surprisingly, after purification, full-length FfCryD as well as a truncated version containing only the PHR domain bound RNA which influenced their behavior in vitro. Moreover, binding of FfCryD to RNA indicates a putative role in RNA metabolism or in posttranscriptional control of gene expression.
隐花色素/光解酶家族的蛋白质利用紫外-A光、蓝光甚至红光来实现多种功能，如光解酶修复DNA损伤以及隐花色素进行信号传导。DASH型隐花色素在体外保留了修复单链DNA区域中环丁烷嘧啶二聚体（CPD）的能力。然而，大多数生物体拥有负责在体内修复这些损伤的传统CPD光解酶。最近的研究表明，DASH型隐花色素CryD在藤仓镰孢菌的多种光依赖过程中发挥调节作用。在此，我们报告了对异源表达的FfCryD的体外研究。纯化后的蛋白质含有N(5),N(10)-亚甲基四氢叶酸和黄素腺嘌呤二核苷酸作为辅因子。光还原和DNA光修复实验证实FfCryD在光驱动的电子转移过程中具有活性。然而，该蛋白质对包含CPD的DNA探针和未受损的DNA探针表现出相当的亲和力。令人惊讶的是，纯化后，全长FfCryD以及仅包含PHR结构域的截短版本都能结合RNA，这影响了它们在体外的行为。此外，FfCryD与RNA的结合表明其在RNA代谢或基因表达的转录后控制中可能发挥作用。
