Transcriptional mechanisms associated with seed dormancy and dormancy loss in the gibberellin-insensitive sly1-2 mutant of Arabidopsis thaliana

Abstract

While widespread transcriptome changes were previously observed with seed dormancy loss, this study specifically characterized transcriptional changes associated with the increased seed dormancy and dormancy loss of the gibberellin (GA) hormone-insensitive sleepy1-2 (sly1-2) mutant. The SLY1 gene encodes the F-box subunit of an SCF E3 ubiquitin ligase needed for GA-triggered proteolysis of DELLA repressors of seed germination. DELLA overaccumulation in sly1-2 seeds leads to increased dormancy that can be rescued without DELLA protein destruction either by overexpression of the GA receptor, GA-INSENSITIVE DWARF1b (GID1b-OE) (74% germination) or by extended dry after-ripening (11 months, 51% germination). After-ripening of sly1 resulted in different transcriptional changes in early versus late Phase II of germination that were consistent with the processes known to occur. Approximately half of the transcriptome changes with after-ripening appear to depend on SLY1-triggered DELLA proteolysis. Given that many of these SLY1/GA-dependent changes are genes involved in protein translation, it appears that GA signaling increases germination capacity in part by activating translation. While sly1-2 after-ripening was associated with transcript-level changes in 4594 genes over two imbibition timepoints, rescue of sly1-2 germination by GID1b-OE was associated with changes in only 23 genes. Thus, a big change in sly1-2 germination phenotype can occur with relatively little change in the global pattern of gene expression during the process of germination. Most GID1b-OE-responsive transcripts showed similar changes with after-ripening in early Phase II of imbibition, but opposite changes with after-ripening by late Phase II. This suggests that GID1b-OE stimulates germination early in imbibition, but may later trigger negative feedback regulation.

Lost in Translation: Physiological Roles of Stored mRNAs in Seed Germination

“…It is worth noting that transcriptomics highlighted a link between dormancy release and the accumulation of mRNAs associated with the protein synthesis machinery, which led to the concept that translational capacity plays a major role in the switch from dormant to non-dormant state [66]. Such translational control for breaking dormancy may be, at least partially, controlled by GA signaling as after-ripening and subsequent cold stratification resulted in massive mRNA increases for translation-associated genes in WT, but not in the GA-insensitive sly1-2 mutant [67]. Similarly, during dormancy release in response to cold stratification or exogenous nitrate, the accumulation of several proteins involved in translational machinery has been observed, suggesting that either the turnover of these specific proteins is affected when seeds are maintained in a dormant state or that the synthesis/stability of the corresponding mRNAs is enhanced upon dormancy release [68].…”

Exome sequencing of bulked segregants identified a novel TaMKK3-A allele linked to the wheat ERA8 ABA-hypersensitive germination phenotype

“…RNAseq was used to identify additional ERA8 candidate genes in the ERA8-linked region based on differential expression in imbibing ZakERA8 versus wild-type seeds. Differences in seed dormancy should result in many transcriptome differences downstream of the ERA8 mutation (Barrero et al 2009;Liu et al 2013a;Nelson and Steber 2017). To focus on transcriptome differences possibly resulting from an ERA8 promoter or intron mutation, transcriptome analysis was performed comparing ERA8 and Zak WT seed afterripened long enough to reduce differences in seed dormancy.…”

Exome sequencing of bulked segregants identified a novel TaMKK3-A allele linked to the wheat ERA8 ABA-hypersensitive germination phenotype

“…Embryos were dissected and frozen in liquid nitrogen under a green safelight to avoid induction of photosynthesis genes. RNA was extracted from 20 dissected embryos per sample using a modified version of an RNA extraction technique which yielded an average of 2,300 ng/μL of RNA with a RIN > 8.0 (Onate-Sánchez and Vicente-Carbajosa 2008; Nelson and Steber 2017). This was replicated three times for each parent sample.…”

“…RNA-seq was used to identify additional ERA8 candidate genes in the ERA8linked region based on differential expression in imbibing ZakERA8 versus wild-type seeds. Differences in seed dormancy should result in many transcriptome differences downstream of the ERA8 mutation (Barrero et al 2009;Liu et al 2013a;Nelson and Steber 2017). To focus on transcriptome differences possibly resulting from an ERA8 promoter or intron mutation, transcriptome analysis was performed comparing ERA8 and Zak WT seed after-ripened long enough to reduce differences in seed dormancy.…”