A plastidial pathway for protein isoprenylation in tobacco cells.

Isoprenylation is an important posttranslational modification of proteins in eukaryotic cells, as it facilitates protein–membrane and protein–protein interactions. In plants, isoprenylated proteins (which include a number of small GTPases and the G-protein g-subunit) are involved in phytohormone signaling, meristem maintenance, cell cycle regulation, and stress responses (Crowell, 2000; Johnson et al., 2005). Isoprenylation involves the covalent attachment of a geranyl-geranyl or farnesyl moiety to a C-terminal Cys of the target protein. The resulting isoprenyl moiety often undergoes further modification, which may be related to its role in signaling. For example, it was recently shown that methylation of isoprenylated proteins plays an important role in regulating abscisic acid signaling (Huizinga et al., 2008). A key step in protein isoprenylation is the generation of the geranyl-geranyl diphosphate or farnesyl diphosphate substrates. In plants, generation of these compounds occurs via two pathways: the cytosolic mevalonate (MVA) pathway and the plastidial 2-C-methyl-Derythritol 4-phosphate (MEP) pathway. Previous work has shown that there is crosstalk between the two pathways, and their relative importance in protein isoprenylation in plants remains an open question. Gerber et al. (pages 285–300) use a fluorescent labeling assay to demonstrate that the plastidial MEP pathway plays the major role in providing the isoprenoid moiety for protein geranyl-geranylation in tobacco BY-2 cells. The authors devised a marker for monitoring isoprenylation of proteins, consisting of a green fluorescent protein that can be geranylgeranylated. When so modified, the GFP marker localizes to the plasma membrane, whereas the unmodified version is targeted to the cell nucleus (see figure). A series of experiments with specific chemical inhibitors showed that inhibition of the MEP pathway caused partial translocation of the marker to the nucleus, whereas inhibition of the MVA pathway had no effect, and inhibiting both pathways together caused complete translocation to the nucleus. The specificity of the inhibitors for each pathway was confirmed by complementation with corresponding pathway intermediates. Taken together, the results suggested that the MEP pathway is the principal route of geranyl-geranylation in these cells. It will be important to follow up these studies in plants, as BY-2 cells have specific properties that differ from whole plants in ways that could affect flux through these pathways. For example, unlike BY-2 cells, the plastidial MEP pathway in green leaves is directed toward producing photosynthetic pigments. This work nevertheless represents a major contribution to understanding isoprenylation in plant cells. In addition, the authors note that the development of the fluorescent marker and rapid, visual assay for inhibition of the MEP pathway could have important pharmaceutical and agricultural significance, as MEP pathway inhibitors may be useful as antibiotics, antimalarial drugs, or herbicides (Rodrı́guez-Concepción, 2004).