title: Protoplast: a more efficient system to study nucleo-cytoplasmic interactions.
abstract: Agrobacterium tumefaciens-mediated genetic transformation is a powerful tool for plant research, but it can be labor-intensive and time-consuming. Here, we report a protoplast-based approach to study nucleo-cytoplasmic interactions, such as cytoplasmic male sterility/fertility restoration (CMS/Rf) and organellar RNA editing. To test the system, we transfected the fertility restorer gene Rf5, which is involved in the rice HL-CMS/Rf system, into rice protoplasts prepared from the HL-CMS line. As the Rf5 protein accumulated in the transformed protoplasts, the CMS-associated transcripts were endonucleolytically cleaved. There were much lower levels of the CMS-associated protein ORFH79 in the transfected protoplasts than in the mock-transfected protoplasts. Next, we used a dsRNA-mediated gene silencing approach to down-regulate the pentatricopeptide protein gene MPR25, which participates in RNA editing of the organellar transcript nad5. The editing efficiency of mitochondrial transcripts of nad5 at nucleotide 1580 was much lower in the transfected protoplasts than in the mock-transfected protoplasts. Together, these results show that protoplast is a simple and efficient system to study interactions between the nucleus and organelles.
authors.affiliation: State Key Laboratory of Hybrid Rice, College of Life Sciences, Wuhan University, China.
authors.forename: Changchun
authors.initials: C
authors.lastname: Yu
authors.affiliation: State Key Laboratory of Hybrid Rice, College of Life Sciences, Wuhan University, China.
authors.forename: Lili
authors.initials: L
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Hybrid Rice, College of Life Sciences, Wuhan University, China.
authors.forename: Cong
authors.initials: C
authors.lastname: Chen
authors.affiliation: State Key Laboratory of Hybrid Rice, College of Life Sciences, Wuhan University, China.
authors.forename: Chunlan
authors.initials: C
authors.lastname: He
authors.affiliation: State Key Laboratory of Hybrid Rice, College of Life Sciences, Wuhan University, China.
authors.forename: Jun
authors.initials: J
authors.lastname: Hu
authors.affiliation: State Key Laboratory of Hybrid Rice, College of Life Sciences, Wuhan University, China. Electronic address: zhuyg@public.wh.hb.cn.
authors.forename: Yingguo
authors.initials: Y
authors.lastname: Zhu
authors.affiliation: State Key Laboratory of Hybrid Rice, College of Life Sciences, Wuhan University, China. Electronic address: gangday@163.com.
authors.forename: Wenchao
authors.initials: W
authors.lastname: Huang
mesh_terms: D002467:Cell Nucleus / Q000378:metabolism*; D003593:Cytoplasm / Q000378:metabolism*; D012275:Oryza / Q000196:embryology; D011523:Protoplasts / Q000378:metabolism*; D012639:Seeds / Q000378:metabolism
keywords: Cytoplasmic male sterility; Fertility restoration; Nucleo-cytoplasmic interaction; Protoplast; RNA editing; Rice