title: Whole-mount confocal imaging of nuclei in giant feeding cells induced by root-knot nematodes in Arabidopsis.
abstract: • Excellent visualization of nuclei was obtained here using a whole-mount procedure adapted to provide high-resolution images of large, irregularly shaped nuclei. The procedure is based on tissue clearing, and fluorescent staining of nuclear DNA with the dye propidium iodide. • The method developed for standard confocal imaging was applied to large multicellular root swellings, named galls, induced in plant hosts by the root-knot nematode Meloidogyne incognita. • Here, we performed a functional analysis, and examined the nuclear structure in giant feeding cells overexpressing the cell cycle inhibitor Kip-related protein 4 (KRP4). Ectopic KRP4 expression in galls led to aberrant nuclear structure, disturbing giant cell expansion and nematode reproduction. In vivo live-cell imaging of GFP-KRP4 demonstrated that this protein co-localizes to chromosomes from prophase to late anaphase during cell cycle progression. • The data presented here suggest the involvement of KRP4 during mitotic progression in plant cells. The detailed results obtained using confocal analysis also demonstrate the potential utility of a rapid, easy-to-use clearing method for the analysis of the nuclei of certain Arabidopsis mutants and other complex plant nuclei.
authors.affiliation: Institut National de la Recherche Agronomique, UMR 1355 ISA, 400 route des Chappes, Sophia-Antipolis, France.|Centre National de la Recherche Scientifique, UMR 7254 ISA, 400 route des Chappes, Sophia-Antipolis, France.|Université de Nice-Sophia Antipolis, UMR ISA, 400 route des Chappes, Sophia-Antipolis, France.
authors.forename: Paulo
authors.initials: P
authors.lastname: Vieira
authors.affiliation: Institut National de la Recherche Agronomique, UMR 1355 ISA, 400 route des Chappes, Sophia-Antipolis, France.|Centre National de la Recherche Scientifique, UMR 7254 ISA, 400 route des Chappes, Sophia-Antipolis, France.|Université de Nice-Sophia Antipolis, UMR ISA, 400 route des Chappes, Sophia-Antipolis, France.
authors.forename: Gilbert
authors.initials: G
authors.lastname: Engler
authors.affiliation: Institut National de la Recherche Agronomique, UMR 1355 ISA, 400 route des Chappes, Sophia-Antipolis, France.|Centre National de la Recherche Scientifique, UMR 7254 ISA, 400 route des Chappes, Sophia-Antipolis, France.|Université de Nice-Sophia Antipolis, UMR ISA, 400 route des Chappes, Sophia-Antipolis, France.
authors.forename: Janice
authors.initials: J
authors.lastname: de Almeida Engler
mesh_terms: D000818:Animals; D017360:Arabidopsis / Q000166:cytology / Q000469:parasitology*; D029681:Arabidopsis Proteins / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism*; D048430:Cell Shape; D050756:Cyclin-Dependent Kinase Inhibitor Proteins / Q000378:metabolism; D015726:Giant Cells / Q000166:cytology* / Q000378:metabolism / Q000469:parasitology; D049452:Green Fluorescent Proteins / Q000378:metabolism; D018613:Microscopy, Confocal / Q000379:methods*; D009348:Nematoda / Q000502:physiology*; D059828:Plant Cells / Q000469:parasitology; D010935:Plant Diseases / Q000469:parasitology*; D018517:Plant Roots / Q000166:cytology* / Q000469:parasitology; D010941:Plant Tumors / Q000469:parasitology; D011419:Propidium / Q000378:metabolism; D021381:Protein Transport; D013194:Staining and Labeling
chemical_list: D029681:Arabidopsis Proteins; D050756:Cyclin-Dependent Kinase Inhibitor Proteins; C550809:KRP4 protein, Arabidopsis; D049452:Green Fluorescent Proteins; D011419:Propidium