Molecular and reverse genetic characterization of NUCLEOSOME ASSEMBLY PROTEIN1 (NAP1) genes unravels their function in transcription and nucleotide excision repair in Arabidopsis thaliana
Molecular and reverse genetic characterization of NUCLEOSOME ASSEMBLY PROTEIN1 (NAP1) genes unravels their function in transcription and nucleotide excision repair in Arabidopsis thaliana
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核小体组装蛋白 1 (NAP1) 基因的分子和反向遗传表征揭示了它们在拟南芥转录和核苷酸切除修复中的功能
DOI：
10.1111/j.1365-313x.2009.03844.x
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发表时间：
2009-07-01
期刊：
影响因子：
7.2
通讯作者：
Shen, Wen-Hui
中科院分区：
文献类型：
--
作者：
Liu, Ziqiang;Zhu, Yan;Shen, Wen-Hui
与核小体组装蛋白1 （NAP1）作为组蛋白伴侣在核小体组装/拆卸中的生化功能相比，NAP1的生理作用在很大程度上仍未被明确。在这里，我们定义了拟南芥中NAP1基因家族成员，研究了它们的分子特性，并使用反向遗传学来表征它们的生物学作用。我们发现4种atnap1组蛋白可以形成同型二聚体和异源二聚体，可以结合组蛋白H2A，并且在稳定状态下大量定位于细胞质中，弱定位于细胞核中。AtNAP1;与其他菌株不同的是，4号菌株表现出对抑制剂敏感的核细胞质穿梭和组织特异性表达，仅限于根段和花粉粒。另外三个AtNAP1基因在植物和AtNAP1中普遍表达；在幼苗中检测到3蛋白为主要亚型。我们表明，在我们的实验室条件下，atnap1组基因的破坏不会影响正常的植物生长。有趣的是，两个等位基因三突变体，Atnap1；1 - 1 Atnap1;2 - 1 Atnap1;3-1和Atnap1；1 - 1 Atnap1;2 - 1 Atnap1;3-2，表现出基因组转录紊乱，并对UV-C照射引起的DNA损伤表现出超敏反应。我们证明了AtNAP1；3结合染色质，富集一些参与核苷酸切除修复（NER）途径的基因，这些基因的表达在三突变体中下调。综上所述，我们的研究结果突出了AtNAP1蛋白的保守性和异构体特异性，并揭示了它们在DNA损伤修复的NER途径中的功能。
P>Compared with the well-studied biochemical function of NUCLEOSOME ASSEMBLY PROTEIN1 (NAP1) as a histone chaperone in nucleosome assembly/disassembly, the physiological roles of NAP1 remain largely uncharacterized. Here, we define the NAP1 gene family members in Arabidopsis, examine their molecular properties, and use reverse genetics to characterize their biological roles. We show that the four AtNAP1-group proteins can form homodimers and heterodimers, can bind histone H2A, and are localized abundantly in the cytoplasm and weakly in the nucleus at steady state. AtNAP1;4 differs from the others by showing inhibitor-sensitive nucleocytoplasmic shuttling and tissue-specific expression, restricted to root segments and pollen grains. The other three AtNAP1 genes are ubiquitously expressed in plants and the AtNAP1;3 protein is detected as the major isoform in seedlings. We show that disruption of the AtNAP1-group genes does not affect normal plant growth under our laboratory conditions. Interestingly, two allelic triple mutants, Atnap1;1-1 Atnap1;2-1 Atnap1;3-1 and Atnap1;1-1 Atnap1;2-1 Atnap1;3-2, exhibit perturbed genome transcription, and show hypersensitivity to DNA damage caused by UV-C irradiation. We show that AtNAP1;3 binds chromatin, with enrichment at some genes involved in the nucleotide excision repair (NER) pathway, and that the expression of these genes is downregulated in the triple mutants. Taken together, our results highlight conserved and isoform-specific properties of AtNAP1 proteins, and unravel their function in the NER pathway of DNA damage repair.