Plant and Cell Physiology Advance Access published online on April 1, 2008
Plant and Cell Physiology, doi:10.1093/pcp/pcn051
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Tomato Rab11a characterization evidenced a difference between SYP121 dependent and SYP122 dependent exocytosis
1: Di.S.Te.B.A., Università del Salento; via prov. Lecce-Monteroni; 73100 – Lecce, Italy
2: Plant Sciences Division, University of Nottingham; Sutton Bonington Campus, Near Loughborough, LE12 5RD, United Kingdom
3: Department of Biology; Plant Biology, University of Fribourg; ch. Du Musée 10; CH-1700 - Fribourg, Switzerland
Corresponding autor: Prof. Gian-Pietro Di Sansebastiano, Address: Di.S.Te.B.A., Università del Salento; via prov. Lecce-Monteroni; 73100 – Lecce, Italy, Phone: +390832298714; Fax: +390832298858; e-mail: gp.disansebastiano{at}unile.it
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Abstract |
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The regulatory functions of Rab proteins in membrane trafficking lie in their ability to perform as molecular switches that oscillate between a GTP- and a GDP- bound conformation. The role of tomato LeRab11a in secretion was analysed in tobacco protoplasts. GFP/RFP-tagged LeRab11a was localized at the TGN in vivo. Two serines in the GTP-binding site of the protein were mutagenised, giving rise to the three mutants Rab11S22N, Rab11S27N, and Rab11S22/27N. The double mutation reduced secretion of a marker protein secRGUS, by half, whereas each of the single mutations alone had a much smaller effect, showing that both serines have to be mutated to obtain a dominant negative effect on LeRab11a function. The dominant negative mutant was used to determine whether Rab11 is involved in the pathway(s) regulated by the plasma membrane syntaxins SYP121 and SYP122. Co-expression of either of these GFP-tagged syntaxins with the dominant negative Rab11S22/27N mutant led to the appearance of endosomes but co-expression of GFP-tagged SYP122 also labelled the endoplasmic reticulum and dotted structures. However, co-expression of Rab11S22/27N with SYP121 dominant negative mutants decreased secretion of secRGUS further compared to the expression of Rab11S22/27N alone, whereas co-expression of Rab11S22/27N with SYP122 had no synergistic effect. With the same essay the difference between SYP121 and SYP122 dependent secretion was then evidenced. The results suggest that Rab11 regulates anterograde transport from the TGN to the PM and strongly implicate SYP122, rather than SYP121. LeRab11a differential effect supports the possibility that SYP121 and SYP122 drive independent secretory events.
Keywords: Dominant negative mutants - exocytosis - plasma membrane - Rab11 - Syntaxin
^: These authors equally contributed to this work.
Present address of Egidio Stigliano: Institute of Botany, University of Neuchatel, Rue E. Argand 9. 2007-CH
(Received December 3, 2007; Accepted March 21, 2008)
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