Kong L J, Orozco B M, Roe J L, Nagar S, Ou S, Feiler H S, Durfee T, Miller A B, Gruissem W, Robertson D, Hanley-Bowdoin L
Departments of Biochemistry and Botany, North Carolina State University, Raleigh, NC 27695, USA.
EMBO J. 2000 Jul 3;19(13):3485-95. doi: 10.1093/emboj/19.13.3485.
Geminiviruses replicate in nuclei of mature plant cells after inducing the accumulation of host DNA replication machinery. Earlier studies showed that the viral replication factor, AL1, is sufficient for host induction and interacts with the cell cycle regulator, retinoblastoma (pRb). Unlike other DNA virus proteins, AL1 does not contain the pRb binding consensus, LXCXE, and interacts with plant pRb homo logues (pRBR) through a novel amino acid sequence. We mapped the pRBR binding domain of AL1 between amino acids 101 and 180 and identified two mutants that are differentially impacted for AL1-pRBR interactions. Plants infected with the E-N140 mutant, which is wild-type for pRBR binding, developed wild-type symptoms and accumulated viral DNA and AL1 protein in epidermal, mesophyll and vascular cells of mature leaves. Plants inoculated with the KEE146 mutant, which retains 16% pRBR binding activity, only developed chlorosis along the veins, and viral DNA, AL1 protein and the host DNA synthesis factor, proliferating cell nuclear antigen, were localized to vascular tissue. These results established the importance of AL1-pRBR interactions during geminivirus infection of plants.
双生病毒在诱导宿主DNA复制机制积累后，在成熟植物细胞的细胞核中进行复制。早期研究表明，病毒复制因子AL1足以诱导宿主，并且与细胞周期调节因子视网膜母细胞瘤（pRb）相互作用。与其他DNA病毒蛋白不同，AL1不包含pRb结合共有序列LXCXE，而是通过一个新的氨基酸序列与植物pRb同源物（pRBR）相互作用。我们将AL1的pRBR结合域定位在第101至180位氨基酸之间，并鉴定出两个对AL1与pRBR相互作用有不同影响的突变体。用对pRBR结合呈野生型的E-N140突变体感染的植物，表现出野生型症状，并且在成熟叶片的表皮、叶肉和维管细胞中积累病毒DNA和AL1蛋白。接种保留16% pRBR结合活性的KEE146突变体的植物，仅沿叶脉出现黄化，并且病毒DNA、AL1蛋白和宿主DNA合成因子增殖细胞核抗原定位于维管组织。这些结果确立了AL1与pRBR相互作用在双生病毒感染植物过程中的重要性。
