S phase of the cell cycle: a key phase for the regulation of thermodormancy in barley grain

The aim of the present work was to investigate the occurrence of the cell cycle during germination as related to thermodormancy in barley (Hordeum vulgare L., cv. Pewter) grains in relation with abscisic acid (ABA) by: (i) flow cytometry to determine the progression of the cell cycle; and (ii) reverse transcription-PCR to characterize the expression of some important genes involved in cell-cycle regulation. In dry embryos, cells are mostly (82%) arrested in G1 phase of the cell cycle, the remaining cells being in the G2 (17%) or S phase (0.9%). Germination at 20 °C was associated with an increase in the nuclei population in G2 and S (up to 32.5-44.5 and 9.2-11.3%, respectively, after 18-24h). At 30 °C, partial reactivation of the cell cycle occurred in embryos of dormant grains that did not germinate. Incubation with 50mM hydroxyurea suggests that thermodormancy resulted in a blocking of the nuclei in the S phase. In dry dormant grains, transcripts of CDKA1, CYCA3, KRP4, and WEE1 were present, while those of CDKB1, CDKD1, CYCB1, and CYCD4 were not detected. Incubation at 30 °C resulted in a strong reduction of CDKB1, CYCB1, and CYCD4 expression and overexpression of CDK1 and KRP4. ABA had a similar effect as incubation at 30 °C on the expression of CDKB1, CYCB1, and CYCD4, but did not increase that of CDK1 and KRP4. Patterns of gene expression are discussed with regard to thermodormancy expression and ABA.