Loss of XRN4 Function Can Trigger Cosuppression in a Sequence-Dependent Manner
Loss of XRN4 Function Can Trigger Cosuppression in a Sequence-Dependent Manner
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DOI：
10.1093/pcp/pcs078
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发表时间：
2012-07-01
影响因子：
4.9
通讯作者：
Nishimura, Mikio
中科院分区：
文献类型：
--
作者：
Hayashi, Makoto;Nanba, Chieko;Nishimura, Mikio
OLE 1编码一种油质蛋白同工蛋白，一种种子中被称为油体的脂质储备细胞器的主要膜蛋白。产生转基因拟南芥以含有由OLE 1和绿色荧光蛋白（GFP）组成的人工嵌合转基因。过表达的融合蛋白允许可视化的油体的大小和结构在活细胞中使用荧光显微镜。从转基因植物的诱变后代中分离出两种积累增大的油体且GFP荧光降低的突变体xrn 4 -8（OleG）和xrn 4 -9（OleG）。这两种突变体都含有外切核糖核酸酶4（XRN 4）的缺陷，XRN 4是一种已知编码核糖核酸酶的基因，可特异性降解未加帽的mRNA。在这些突变体中转基因表达沉默，如转基因mRNA及其产物OLE 1-GFP水平降低所证明的。XRN 4功能丧失也引发共抑制，即转基因和共享相同序列区域的内源性OLE 1基因的表达同时减少。在xrn 4 -8（OleG）和xrn 4 -9（OleG）突变体中分别由于内源性OLE 1和转基因产物OLE 1-GFP的减少而形成了表现出降低的GFP荧光的增大的油体。由xrn 4突变触发的共抑制也发生在其他基因中，例如PYK 10，其编码内质网（ER）体内驻留的β-葡萄糖苷酶。总体结果表明，XRN 4功能的丧失可能以序列依赖性方式触发共抑制。
OLE1 encodes an oleosin isoprotein, a major membrane protein of the lipid-reserve organelle in seeds known as the oil body. Transgenic Arabidopsis were generated to contain an artificial chimeric transgene composed of OLE1 and green fluorescent protein (GFP). Overexpression of the fusion protein allowed visualization of the oil body size and structure in living cells using fluorescence microscopy. Two mutants, xrn4-8(OleG) and xrn4-9(OleG), accumulating enlarged oil bodies with reduced GFP fluorescence were isolated from the mutagenized progeny of a transgenic plant. Both mutants contained a defect in EXORIBONUCLEASE4 (XRN4), a gene known to encode a ribonuclease that specifically degrades uncapped mRNAs. Transgene expression was silenced in these mutants, as demonstrated by the reduced levels of the transgene mRNA and its product, OLE1-GFP. XRN4 loss of function also triggered cosuppression, i.e. simultaneous reduction in expression of the transgene and an endogenous OLE1 gene that shared a region of identical sequence. The enlarged oil bodies exhibiting reduced GFP fluorescence were formed in the xrn4-8(OleG) and xrn4-9(OleG) mutants due to the reduction of the endogenous OLE1 and the transgene product, OLE1-GFP, respectively. Cosuppression triggered by the xrn4 mutation also occurs for other genes such as PYK10, which encodes an endoplasmic reticulum (ER) body-resident beta-glucosidase. The overall results indicate that a loss of XRN4 function can potentially trigger the cosuppression in a sequence-dependent manner.