JXB Advance Access originally published online on February 10, 2008
Journal of Experimental Botany 2008 59(2):203-212; doi:10.1093/jxb/erm296
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© 2008 The Author(s).
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
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RESEARCH PAPER |
Regulation of cell cycle activity in the embryo of barley seeds during germination as related to grain hydration
1Université Pierre et Marie Curie-Paris 6, EA2388 Physiologie des semences, Site d'Ivry, Boîte 152, 4 place Jussieu, F-75252 Paris Cedex 05, France
2IFEVA-Catedra de Cerealicultura, Facultad de Agronomia, Universidad de Buenos Aires/CONICET, Av. San Martin 4453, 1417 Buenos Aires, Argentina
* To whom correspondence should be addressed. E-mail: corbi{at}ccr.jussieu.fr
Various studies indicate that cell division is a post-germination phenomenon, with radicle protrusion occurring by cell elongation, while others demonstrate that induction of the cell cycle occurs in osmo-conditioned seeds prior to radicle growth. The aim of the present work was to investigate the occurrence of the cell cycle during germination as related to grain hydration, using: (i) a flow cytometry technique to estimate the percentage of cell nuclei in G1 and G2 phases of the cell cycle; and (ii) reverse transcription-PCR (RT-PCR) in order to characterize the expression of the genes encoding cyclin-dependent kinases (CDKA1, CDKB1, and CDKD1) and cyclins (CYCA3, CYCB1, and CYCD4), the main genes involved in the cell cycle and its regulation. Radicle tips of embryos were isolated from seeds placed for various times on water at 30 °C and from grains partially hydrated at moisture contents ranging from 11% to 51% fresh weight (FW), which prevent radicle elongation. Abscisic acid (ABA) contents of the embryos during seed germination at 30 °C and after 48 h of partial hydration were also measured. In dry embryos, cells are mostly arrested in the G1 phase of the cell cycle (82%), the remaining cells being in the G2 phase, and the ABA content of the embryo was 432.7 ng g–1 dry weight (DW). Seed imbibition was associated with a sharp decrease in ABA content as early as 5 h, while the cell cycle reactivation was a late process taking place 
4–6 h prior to radicle protrusion. Hydration of seeds resulted in a decrease in embryo ABA content, but it remained at a high level (207–273 ng g–1 DW) even after 48 h at 0.41–0.51 g H2O g–1 FW. The cell population of the radicle tips in the G2 phase of the cell cycle, i.e. 4C nuclei, increased from 9% up to 34% at a moisture content of 51% FW. In dry seeds, CDKA1 and CDKD1 mRNAs were present at low levels, but transcripts of CDKB1, CYCA3, CYCB1, and CYCD4 were not detected. Radicle protrusion was associated with a higher expression of CDKA1, CDKB1, CYCA3, and CYCB1. Blockage of germination of partially hydrated grains resulted in a reduction in the expression of CDKA1 and CDKB1, and of CYCA3 and CYCB1, and in a reinforcement of that of CDKD1 and CYCD4. Patterns of gene expression show differential sensitivity of the genes studied to hydration of the grain. They will be discussed with regard to embryo ABA content and embryo sensitivity to ABA.
Key words: Abscisic acid, barley, cell cycle, cyclin, cyclin-dependent kinase, germination, Hordeum vulgare L., water content
Received 31 July 2007; Revised 30 October 2007 Accepted 2 November 2007