Characterization of the activity of a plastid-targeted green fluorescent protein in Arabidopsis

In Arabidopsis thaliana the PALE CRESS (PAC) gene product is required for b oth chloroplast and cell differentiation. Transgenic Arabidopsis plants exp ressing a translational fusion of the N-terminal part of the PAC protein ha rboring the complete plastid-targeting sequence and the green fluorescent p rotein (GFP) exhibit high GFP fluorescence. Detailed analyses based on conf ocal imaging of various tissues and cell types revealed that the PAC-GFP fu sion protein accumulates in chloroplasts of mature stomatal guard cells. Th e GFP fluorescence within the guard cell chloroplasts is not evenly distrib uted and appears to be concentrated in suborganellar regions. GFP localizat ion studies demonstrate that thin tubular projections emanating from chloro plasts and etioplasts often connect the organelles with each other. Further more, imaging of non-green and etiolated tissue further revealed that GFP f luorescence is present in proplastids, etioplasts, chromoplasts, and amylop lasts. Even photobleaching of carotenoid-free plastids does not affect PAC- GFP accumulation in the organelles of the guard cells indicating that the p rotein translocation machinery is functional in all types of plastids. The specific accumulation of GFP in guard cell chloroplasts, their tubular conn ections, the translocation of the precursor polypeptide into the different types of organelles, as well as the use of a plastid-targeted GFP protein a s a versatile marker is discussed in the context of previously described ob servations.