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Plant Physiology Preview Published on January 25, 2008; 10.1104/pp.107.113274
OPEN ACCESS ARTICLE
Received November 15, 2007 A plasma membrane-anchored fluorescent protein fusion illuminates sieve element plasma membranes in Arabidopsis thaliana and Nicotiana tabacum
Department of Cell Biology and Molecular Genetics, University of Maryland, College Park, Maryland, 20742, USA * Corresponding author; email: mvthom{at}umd.edu.
Rapid acquisition of quantitative anatomical data from the sieve tubes of angiosperm phloem has been confounded by their small size, their distance from organ surfaces, and the time-consuming nature of traditional methods, such as TEM. To improve access to these cells – for which good anatomical data are critical – a monomeric, yellow fluorescent protein (mCitrine) was N-terminally fused to a small (
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6 kDa) membrane protein (AtRCI2A) and stably expressed in Arabidopsis thaliana (Col-0 ecotype) and Nicotiana tabacum (cv Samsun) under the control of a companion cell-specific promoter (AtSUC2p). The construct, called by its abbreviation SUmCR, yielded stable sieve element plasma membrane fluorescence-labeling, even after plastic (methacrylate) embedding. In conjunction with wide-field fluorescence measurements of sieve pore number and position using aniline blue-stained callose, mCitrine-labeled material was used to calculate rough estimates of sieve tube specific conductivity for both species. The SUmCR construct also revealed a hitherto unknown expression domain of the AtSUC2 sucrose-H+ symporter in the epidermis of the cell division zone of developing root tips. The success of this construct in targeting plasma membrane-anchored fluorescent proteins to sieve elements could be attributable to the small size of AtRCI2A or to the presence of other signals innate to AtRCI2A which permit the protein to be trafficked to sieve elements. The construct provides a hitherto unique entrée into companion cell-to-sieve element protein targeting, as well as a new tool for studying whole-plant phloem anatomy and architecture.
