Cessation of coleoptile elongation and loss of auxin sensitivity in developing rye seedlings: a quantitative...

The use of the grass coleoptile for the elucidation of the mechanism of cell elongation is a legacy of the classic experiments of Charles Darwin, who described this organ in 1880 as a "reddish sheath". In this study we quantified the growth of intact, etiolated rye (Secale cereale L.) seedlings and selected 3-day-old (growing) vs. 4-day-old (pierced) coleoptiles for a comparative analysis. Upon emergence of the reddish primary leaf on day 4 after sowing, growth slowed down by 70% and the sensitivity of the coleoptile to auxin (Indole-3-acetic acid) was lost, but turgor pressure was maintained. A quantitative comparison of the proteome (microsomal- and cytoplasmic protein fractions, respectively), using the two-dimensional difference gel electrophoresis (2-D DIGE)-technique, revealed that at least 28 proteins (spots) were differentially up- or down-regulated more than 1.5-fold. Eight of these proteins were identified by reverse-phase liquid chromatography-electrospray tandem mass spectrometry. Cessation of coleoptile growth was associated with the down-regulation (- 81 %) of subunit E of the vacuolar H(+)-ATPase (V-ATPase) and the up-regulation of enzymes involved in lignification (phenylalanine ammonia lyase) and wounding responses (xylanase inhibitor; two lipoxygenases). We conclude that the degradation of the V-ATPases, electrogenic proton pumps on the tonoplast and the membranes of the Golgi- dependent secretory pathway, may be the cause for the cessation of growth in turgid coleoptiles and the associated loss of auxin sensitivity. However, the intracellular signals that cause these proteomic changes have not yet been identified.