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The plastidial 2-C-methyl-D-erythritol 4-phosphate pathway provides the isoprenyl moiety for protein geranylgeranylation in tobacco BY-2 cells

AuthorsGerber, Esther; Hemmerlin, Andréa; Hartmann, Michael; Heintz, Dimitri; Hartmann, Marie-Andrée; Mutterer, Jerome; Rodríguez Concepción, Manuel; Boronat, Albert ; Van Dorsselaer, Alain; Rohmer, Michel; Crowell, Dring N.; Bach, Thomas J.
Issue Date9-Jan-2009
PublisherAmerican Society of Plant Physiologists
CitationPlant Cell 21(1): 285-300 (2009)
AbstractProtein farnesylation and geranylgeranylation are important posttranslational modifications in eukaryotic cells. We visualized in transformed Nicotiana tabacum Bright Yellow-2 (BY-2) cells the geranylgeranylation and plasma membrane localization of GFP-BD-CVIL, which consists of green fluorescent protein (GFP) fused to the C-terminal polybasic domain (BD) and CVIL isoprenylation motif from the Oryza sativa calmodulin, CaM61. Treatment with fosmidomycin (Fos) or oxoclomazone (OC), inhibitors of the plastidial 2-C-methyl-d-erythritol 4-phosphate (MEP) pathway, caused mislocalization of the protein to the nucleus, whereas treatment with mevinolin, an inhibitor of the cytosolic mevalonate pathway, did not. The nuclear localization of GFP-BD-CVIL in the presence of MEP pathway inhibitors was completely reversed by all-trans-geranylgeraniol (GGol). Furthermore, 1-deoxy-d-xylulose (DX) reversed the effects of OC, but not Fos, consistent with the hypothesis that OC blocks 1-deoxy-d-xylulose 5-phosphate synthesis, whereas Fos inhibits its conversion to 2-C-methyl-d-erythritol 4-phosphate. By contrast, GGol and DX did not rescue the nuclear mislocalization of GFP-BD-CVIL in the presence of a protein geranylgeranyltransferase type 1 inhibitor. Thus, the MEP pathway has an essential role in geranylgeranyl diphosphate (GGPP) biosynthesis and protein geranylgeranylation in BY-2 cells. GFP-BD-CVIL is a versatile tool for identifying pharmaceuticals and herbicides that interfere either with GGPP biosynthesis or with protein geranylgeranylation.
Description17 pages, 11 figures.-- PMID: 19136647 [PubMed].-- PMCID: PMC2648074.-- Supplementay information available at: http://www.plantcell.org/cgi/content/full/tpc.108.063248/DC1
Publisher version (URL)http://dx.doi.org/10.1105/tpc.108.063248
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