Castillo A G, Kong L J, Hanley-Bowdoin L, Bejarano E R
Department of Cellular Biology, Genetics and Animal Physiology, Málaga University, Málaga 29071, Spain.
J Virol. 2004 Mar;78(6):2758-69. doi: 10.1128/jvi.78.6.2758-2769.2004.
Geminiviruses are small DNA viruses that replicate in nuclei of infected plant cells after accumulation of host replication machinery. Tomato golden mosaic virus (TGMV) and Tomato yellow leaf curl Sardinia virus (TYLCSV) encode a protein, RepAC1 (or Rep), that is essential for viral replication. Rep/RepAC1 is an oligomeric protein that binds to double-stranded DNA, catalyzes cleavage and ligation of single-stranded DNA, and is sufficient for host induction. It also interacts with several host proteins, including the cell cycle regulator, retinoblastoma, and essential components of the cell DNA replication machinery, like proliferating nuclear cell antigen (PCNA) and RFC-1. To identify other cellular proteins that interact with Rep/RepAC1 protein, a Nicotiana benthamiana cDNA library was screened with a yeast two-hybrid assay. The host cell sumoylation enzyme, NbSCE1 (N. benthamiana SUMO-conjugating enzyme, homolog to Saccharomyces cerevisiae UBC9), was found to interact specifically with RepAC1. Mapping studies localized the interaction to the N-terminal half of RepAC1. Effects on geminivirus replication were observed in transgenic plants with altered levels of SUMO, the substrate for UBC9.
双生病毒是一类小型DNA病毒，在宿主复制机制积累后于受感染植物细胞的细胞核中进行复制。番茄金色花叶病毒（TGMV）和番茄黄化曲叶撒丁岛病毒（TYLCSV）编码一种蛋白质RepAC1（或Rep），它对病毒复制至关重要。Rep/RepAC1是一种寡聚蛋白，可与双链DNA结合，催化单链DNA的切割和连接，并且足以诱导宿主。它还与几种宿主蛋白相互作用，包括细胞周期调节因子视网膜母细胞瘤蛋白，以及细胞DNA复制机制的重要组成部分，如增殖细胞核抗原（PCNA）和RFC-1。为了鉴定与Rep/RepAC1蛋白相互作用的其他细胞蛋白，利用酵母双杂交试验筛选了本氏烟草cDNA文库。发现宿主细胞类泛素化酶NbSCE1（本氏烟草类泛素缀合酶，与酿酒酵母UBC9同源）与RepAC1特异性相互作用。定位研究将这种相互作用定位到RepAC1的N端一半区域。在类泛素（UBC9的底物）水平改变的转基因植物中观察到了对双生病毒复制的影响。
