保守的核糖核酸内切酶YbeY是拟南芥叶绿体核糖体RNA加工所必需的。
The conserved endoribonuclease YbeY is required for chloroplast ribosomal RNA processing in Arabidopsis.
作者信息
Liu Jinwen, Zhou Wenbin, Liu Guifeng, Yang Chuanping, Sun Yi, Wu Wenjuan, Cao Shenquan, Wang Chong, Hai Guanghui, Wang Zhifeng, Bock Ralph, Huang Jirong, Cheng Yuxiang
机构信息
State Key Laboratory of Tree Genetics and Breeding, Northeast Forestry University, Harbin 150040, China (J.L., G.L., C.Y., S.C., C.W., G.H., Z.W., Y.C.);College of Life Science and Technology, Heilongjiang Bayi Agricultural University, Daqing 163319, China (J.L.);National Key Laboratory of Plant Molecular Genetics, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Science, Chinese Academy of Sciences, Shanghai 200032, China (W.Z., Y.S., W.W., J.H.); andMax-Planck-Institut für Molekulare Pflanzenphysiologie, D-14476 Potsdam-Golm, Germany (R.B.).
State Key Laboratory of Tree Genetics and Breeding, Northeast Forestry University, Harbin 150040, China (J.L., G.L., C.Y., S.C., C.W., G.H., Z.W., Y.C.);College of Life Science and Technology, Heilongjiang Bayi Agricultural University, Daqing 163319, China (J.L.);National Key Laboratory of Plant Molecular Genetics, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Science, Chinese Academy of Sciences, Shanghai 200032, China (W.Z., Y.S., W.W., J.H.); andMax-Planck-Institut für Molekulare Pflanzenphysiologie, D-14476 Potsdam-Golm, Germany (R.B.)
出版信息
Plant Physiol. 2015 May;168(1):205-21. doi: 10.1104/pp.114.255000. Epub 2015 Mar 25.
Maturation of chloroplast ribosomal RNAs (rRNAs) comprises several endoribonucleolytic and exoribonucleolytic processing steps. However, little is known about the specific enzymes involved and the cleavage steps they catalyze. Here, we report the functional characterization of the single Arabidopsis (Arabidopsis thaliana) gene encoding a putative YbeY endoribonuclease. AtYbeY null mutants are seedling lethal, indicating that AtYbeY function is essential for plant growth. Knockdown plants display slow growth and show pale-green leaves. Physiological and ultrastructural analyses of atybeY mutants revealed impaired photosynthesis and defective chloroplast development. Fluorescent microcopy analysis showed that, when fused with the green fluorescence protein, AtYbeY is localized in chloroplasts. Immunoblot and RNA gel-blot assays revealed that the levels of chloroplast-encoded subunits of photosynthetic complexes are reduced in atybeY mutants, but the corresponding transcripts accumulate normally. In addition, atybeY mutants display defective maturation of both the 5' and 3' ends of 16S, 23S, and 4.5S rRNAs as well as decreased accumulation of mature transcripts from the transfer RNA genes contained in the chloroplast rRNA operon. Consequently, mutant plants show a severe deficiency in ribosome biogenesis, which, in turn, results in impaired plastid translational activity. Furthermore, biochemical assays show that recombinant AtYbeY is able to cleave chloroplast rRNAs as well as messenger RNAs and transfer RNAs in vitro. Taken together, our findings indicate that AtYbeY is a chloroplast-localized endoribonuclease that is required for chloroplast rRNA processing and thus for normal growth and development.
叶绿体核糖体RNA（rRNA）的成熟包括几个内切核糖核酸酶和外切核糖核酸酶加工步骤。然而，对于所涉及的具体酶及其催化的切割步骤知之甚少。在这里，我们报道了拟南芥中一个编码假定的YbeY内切核糖核酸酶的单基因的功能特性。AtYbeY基因敲除突变体在幼苗期致死，这表明AtYbeY的功能对植物生长至关重要。基因敲低植株生长缓慢，叶片呈淡绿色。对atybeY突变体的生理和超微结构分析表明，光合作用受损，叶绿体发育存在缺陷。荧光显微镜分析显示，当与绿色荧光蛋白融合时，AtYbeY定位于叶绿体中。免疫印迹和RNA凝胶印迹分析表明，atybeY突变体中光合复合体的叶绿体编码亚基水平降低，但相应的转录本正常积累。此外，atybeY突变体在16S、23S和4.5S rRNA的5'和3'末端均存在成熟缺陷，并且叶绿体rRNA操纵子中包含的转运RNA基因的成熟转录本积累减少。因此，突变植株在核糖体生物合成方面存在严重缺陷，进而导致质体翻译活性受损。此外，生化分析表明，重组AtYbeY能够在体外切割叶绿体rRNA以及信使RNA和转运RNA。综上所述，我们的研究结果表明，AtYbeY是一种定位于叶绿体的内切核糖核酸酶，是叶绿体rRNA加工所必需的，因此对正常生长发育至关重要。
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