Pagano Andrea, Araújo Susana de Sousa, Macovei Anca, Leonetti Paola, Balestrazzi Alma
Department of Biology and Biotechnology 'Lazzaro Spallanzani', University of Pavia, Pavia, Italy.
Plant Cell Biotechnology, Instituto de Tecnologia Química e Biológica António Xavier (ITQB-NOVA), Oeiras, Portugal.
Front Plant Sci. 2017 Nov 14;8:1972. doi: 10.3389/fpls.2017.01972. eCollection 2017.
This work provides novel insights into the effects caused by the histone deacetylase inhibitor trichostatin A (TSA) during seed germination, with emphasis on the seed repair response. Seeds treated with HO and TSA (10 and 20 μM) were collected during imbibition (8 h) and at the radicle protrusion phase. Biometric data showed delayed germination and impaired seedling growth in TSA-treated samples. Comet assay, performed on radicles at the protrusion phase and 4-days old seedlings, revealed accumulation of DNA strand breaks upon exposure to TSA. Activation of DNA repair toward TSA-mediated genotoxic damage was evidenced by the up-regulation of () gene involved in the removal of oxidative DNA lesions, () gene, a key determinant of seed quality, required for the rejoining of DNA double strand breaks and genes encoding the multipurpose DNA repair enzymes tyrosyl-DNA phosphodiesterases. Since radical scavenging can prevent DNA damage, the specific antioxidant activity (SAA) was measured by DPPH (1,1-diphenyl-2-picrylhydrazyl) and Folin-Ciocalteu reagent assays. Fluctuations of SAA were observed in TSA-treated seeds/seedlings concomitant with the up-regulation of antioxidant genes () and (). Chromatin remodeling, required to facilitate the access of DNA repair enzymes at the damaged sites, is also part of the multifaceted seed repair response. To address this aspect, still poorly explored in plants, the () gene was analyzed. TRRAP is a transcriptional adaptor, so far characterized only in human cells where it is needed for the recruitment of histone acetyltransferase complexes to chromatin during DNA repair. The gene and the predicted interacting partners () and () showed tissue- and dose-dependent fluctuations in transcript levels. PCA (Principal Component Analysis) and correlation analyses suggest for a new putative link between DNA repair and chromatin remodeling that involves and genes, in the context of seed germination. Interesting correlations also connect DNA repair and chromatin remodeling with antioxidant players and proliferation markers.
这项工作为组蛋白脱乙酰酶抑制剂曲古抑菌素A（TSA）在种子萌发过程中所产生的影响提供了新的见解，重点是种子修复反应。在吸胀（8小时）期间和胚根突出阶段收集用HO和TSA（10和20μM）处理的种子。生物统计学数据显示，经TSA处理的样品发芽延迟且幼苗生长受损。对胚根突出阶段的胚根和4日龄幼苗进行彗星试验，结果显示暴露于TSA后DNA链断裂积累。参与去除氧化性DNA损伤的（）基因、DNA双链断裂重新连接所需的种子质量关键决定因素（）基因以及编码多功能DNA修复酶酪氨酰-DNA磷酸二酯酶的基因上调，证明了针对TSA介导的基因毒性损伤的DNA修复激活。由于自由基清除可以防止DNA损伤，因此通过DPPH（1,1-二苯基-2-苦基肼）和福林-酚试剂测定法测量了特定抗氧化活性（SAA）。在经TSA处理的种子/幼苗中观察到SAA的波动，同时抗氧化基因（）和（）上调。促进DNA修复酶进入受损位点所需的染色质重塑也是多方面种子修复反应的一部分。为了解决植物中尚未充分探索的这一方面，对（）基因进行了分析。TRRAP是一种转录衔接子，迄今为止仅在人类细胞中得到表征，在DNA修复过程中，它是组蛋白乙酰转移酶复合物募集到染色质所必需的。（）基因以及预测的相互作用伙伴（）和（）在转录水平上表现出组织和剂量依赖性波动。主成分分析（PCA）和相关性分析表明，在种子萌发的背景下，DNA修复与染色质重塑之间存在一种新的假定联系，涉及（）和（）基因。有趣的相关性还将DNA修复和染色质重塑与抗氧化因子和增殖标记联系起来。
