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Published on September 28, 2007; 10.1104/pp.107.103945

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Received June 28, 2007
Accepted September 21, 2007

Fluorescent Reporter Proteins for the Tonoplast and the Vacuolar Lumen Identify a Single Vacuolar Compartment in Arabidopsis Cells

Paul Richard Hunter , Christian Paul Craddock , Sara Di Benedetto , Lynne Margaret Roberts , and Lorenzo Frigerio *

Department of Biological Sciences, University of Warwick, Coventry CV4 7AL, United Kingdom

* Corresponding author; email: l.frigerio{at}warwick.ac.uk.

We generated fusions between three Arabidopsis thaliana tonoplast intrinsic proteins ({alpha}, {gamma} and {delta}-TIP) and the yellow fluorescent protein (YFP). We also produced soluble reporters consisting of the monomeric red fluorescent protein (RFP) and either the C-terminal vacuolar sorting signal of phaseolin or the sequence-specific sorting signal of proricin. In transgenic Arabidopsis leaves, mature roots and root tips, all TIP fusions localized to the tonoplast of the central vacuole and both of the luminal RFP reporters were found within TIP-delimited vacuoles. In embryos from developing, mature and germinating seeds, all three TIPs localized to the tonoplast of the protein storage vacuoles. To determine the temporal TIP expression patterns and to rule out mistargeting due to overexpression, we generated plants expressing YFP fused to the complete genomic sequences of the three TIP isoforms. In transgenic Arabidopsis, {gamma}-TIP expression was limited to vegetative tissues but specifically excluded from root tips, whereas {alpha}-TIP was exclusively expressed during seed maturation. {delta}-TIP was expressed in vegetative tissues, but not root tips, at a later stage than {gamma}-TIP. Our findings indicate that – in the Arabidopsis tissues analyzed – two different vacuolar sorting signals target soluble proteins to a single vacuolar location. Moreover, TIP isoform distribution is tissue- and development-specific, rather than organelle-specific.




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