Active protein transport through plastid tubules: velocity quantified by fluorescence correlation spectroscopy

Dynamic tubular projections emanate from plastids in certain cells of vascu lar plants and are especially prevalent in non-photosynthetic cells. Tubule s sometimes connect two or more different plastids and can extend over long distances within a cell, observations that suggest that the tubules may fu nction in distribution of molecules within, to and from plastids, In a new application of two-photon excitation (2PE) fluorescence correlation spectro scopy (FCS), we separated diffusion of fluorescent molecules from active tr ansport in vivo. We quantified the velocities of diffusion versus active tr ansport of green fluorescent protein (GFP) within plastid tubules and in th e cytosol in vivo. GFP moves by 3-dimensional (3-D) diffusion both in the c ytosol and plastid tubules, but diffusion in tubules is about 50 times and 100 times slower than in the cytosol and an aqueous solution, respectively. Unexpectedly larger GFP units within plastid tubules exhibited active tran sport with a velocity of about 0.12 mum/second. Active transport might play an important role in the long-distance distribution of large numbers of mo lecules within the highly viscous stroma of plastid tubules.