Elucidating the Germination Transcriptional Program Using Small Molecules    

Abstract

The transition from seed to seedling is mediated by germination, a complex process that starts with imbibition and completes with radicle emergence. To gain insight into the transcriptional program mediating germination, previous studies have compared the transcript profiles of dry, dormant, and germinating after-ripened Arabidopsis (Arabidopsis thaliana) seeds. While informative, these approaches did not distinguish the transcriptional responses due to imbibition, shifts in metabolism, or breaking of dormancy from those triggered by the initiation of germination. In this study, three mechanistically distinct small molecules that inhibit Arabidopsis seed germination (methotrexate, 2, 4-dinitrophenol, and cycloheximide) were identified using a small-molecule screen and used to probe the germination transcriptome. Germination-responsive transcripts were defined as those with significantly altered transcript abundance across all inhibitory treatments with respect to control germinating seeds, using data from ATH1 microarrays. This analysis identified numerous germination regulators as germination responsive, including the DELLA proteins GAI, RGA, and RGL3, the abscisic acid-insensitive proteins ABI4, ABI5, ABI8, and FRY1, and the gibberellin receptor GID1A. To help visualize these and other publicly available seed microarray data, we designed a seed mRNA expression browser using the electronic Fluorescent Pictograph platform. An overall decrease in gene expression and a 5-fold greater number of transcripts identified as statistically down-regulated in drug-inhibited seeds point to a role for mRNA degradation or turnover during seed germination. The genes identified in our study as responsive to germination define potential uncharacterized regulators of this process and provide a refined transcriptional signature for germinating Arabidopsis seeds.

Temperature variability is integrated by a spatially embedded decision-making center to break dormancy in Arabidopsis seeds

“…each of these proteins are not expressed at high levels in nongerminating Arabidopsis seeds (28,29) (Fig. 2G and SI Appendix, Supplementary Figs.…”

Induction of Dormancy in Arabidopsis Summer Annuals Requires Parallel Regulation of DOG1 and Hormone Metabolism by Low Temperature and CBF Transcription Factors

“…Given that low seed maturation temperature leads to high dormancy at warm imbibition temperatures ( Figure 1A), and at warmer temperatures FLC was shown to have little or no role (Chiang et al, 2009), we investigated whether CBF-dependent pathways have a role in dormancy induction by low maturation temperatures. Investigation of available microarray data using the EFP browser (Bassel et al, 2008) also suggested that CBFs might be expressed during seed development. To analyze the temperature regulation of CBF transcription in seeds, we performed time-course experiments in which seeds or seedlings were transferred from 22 to 48C and the expression of CBF1 recorded by RT-PCR.…”

A Guideline to Family-Wide Comparative State-of-the-Art Quantitative RT-PCR Analysis Exemplified with a Brassicaceae Cross-Species Seed Germination Case Study

“…For this in silico analysis, we used the seed-specific eFP browser and the eNorthern tool (available at www.bar.utoronto.ca) to access diverse transcriptome data sets not only of Arabidopsis seed germination but also of seed development as a distinct developmental process (Toufighi et al, 2005;Winter et al, 2007;Bassel et al, 2008). Altogether, 101 different microarray experiments focusing on diverse treatments, mutants, tissues, and developmental processes of Arabidopsis seeds were analyzed (see Supplemental Data Set 2 online).…”