Hu Yuzhao, Rosado Daniele, Lindbäck Louise N, Micko Julie, Pedmale Ullas V
Cold Spring Harbor Laboratory, 1 Bungtown Road, Cold Spring Harbor, New York 11724.
bioRxiv. 2023 Jan 16:2023.01.15.524001. doi: 10.1101/2023.01.15.524001.
Cryptochromes (CRYs) are evolutionarily conserved blue-light receptors that evolved from bacterial photolyases that repair damaged DNA. Today, CRYs have lost their ability to repair damaged DNA; however, prior reports suggest that human CRYs can respond to DNA damage. Currently, the role of CRYs in the DNA damage response (DDR) is lacking, especially in plants. Therefore, we evaluated the role of plant CRYs in DDR along with UBP12/13 deubiquitinases, which interact with and regulate the CRY2 protein. We found that was hypersensitive, while was hyposensitive to UVC-induced DNA damage. Elevated UV-induced cyclobutane pyrimidine dimers (CPDs) and the lack of DNA repair protein RAD51 accumulation in plants indicate that CRYs are required for DNA repair. On the contrary, CPD levels diminished and RAD51 protein levels elevated in plants lacking UBP12 and UBP13, indicating their role in DDR repression. Temporal transcriptomic analysis revealed that DDR-induced transcriptional responses were subdued in , but elevated in compared to WT. Through transcriptional modeling of the time-course transcriptome, we found that genes quickly induced by UVC (15 min) are targets of CAMTA 1-3 transcription factors, which we found are required for DDR. This transcriptional regulation seems, however, diminished in the mutant, indicating that CAMTAs are required for CRY2-mediated DDR. Furthermore, we observed enhanced CRY2-UBP13 interaction and formation of CRY2 nuclear speckles under UVC, suggesting that UVC activates CRY2 similarly to blue light. Together, our data reveal the temporal dynamics of the transcriptional events underlying UVC-induced genotoxicity and expand our knowledge of the role of CRY and UBP12/13 in DDR.
隐花色素（CRYs）是进化上保守的蓝光受体，由修复受损DNA的细菌光解酶进化而来。如今，CRYs已失去修复受损DNA的能力；然而，先前的报道表明人类CRYs可对DNA损伤作出反应。目前，CRYs在DNA损伤反应（DDR）中的作用尚不清楚，尤其是在植物中。因此，我们评估了植物CRYs在DDR中的作用，以及与CRY2蛋白相互作用并对其进行调控的去泛素化酶UBP12/13的作用。我们发现，[具体植物名称1]对UVC诱导的DNA损伤高度敏感，而[具体植物名称2]则低敏感。[具体植物名称1]中紫外线诱导的环丁烷嘧啶二聚体（CPDs）升高以及DNA修复蛋白RAD51积累的缺乏表明，DNA修复需要CRYs。相反，在缺乏UBP12和UBP13的植物中，CPD水平降低，RAD51蛋白水平升高，表明它们在DDR抑制中发挥作用。时间转录组分析显示，DDR诱导的转录反应在[具体植物名称1]中受到抑制，但与野生型相比，在[具体植物名称2]中升高。通过对时间进程转录组的转录建模，我们发现UVC快速诱导（15分钟）的基因是CAMTA 1-3转录因子的靶标，我们发现这些转录因子是DDR所必需的。然而，这种转录调控在[具体植物名称1]突变体中似乎减弱，表明CAMTAs是CRY2介导的DDR所必需的。此外，我们观察到在UVC照射下，CRY2-UBP13相互作用增强，CRY2核斑点形成，这表明UVC与蓝光类似地激活CRY2。总之，我们的数据揭示了UVC诱导的遗传毒性背后转录事件的时间动态，并扩展了我们对CRY和UBP12/13在DDR中作用的认识。
