A Molecular Signal Integration Network Underpinning Arabidopsis Seed Germination.

Seed dormancy is an adaptive trait defining where and when plants are established. Diverse signals from the environment are used to decide when to initiate seed germination, a process driven by the expansion of cells within the embryo. How these signals are integrated and transduced into the biomechanical changes that drive embryo growth remains poorly understood. Using Arabidopsis seeds, we demonstrate that cell-wall-loosening EXPANSIN (EXPA) genes promote gibberellic acid (GA)-mediated germination, identifying EXPAs as downstream molecular targets of this developmental phase transition. Molecular interaction screening identified transcription factors (TFs) that bind to both EXPA promoter fragments and DELLA GA-response regulators. A subset of these TFs is targeted each by nitric oxide (NO) and the phytochrome-interacting TF PIL5. This molecular interaction network therefore directly links the perception of an external environmental signal (light) and internal hormonal signals (GA and NO) with downstream germination-driving EXPA gene expression. Experimental validation of this network established that many of these TFs mediate GA-regulated germination, including TCP14/15, RAP2.2/2.3/2.12, and ZML1. The reduced germination phenotype of the tcp14 tcp15 mutant seed was partially rescued through ectopic expression of their direct target EXPA9. The GA-mediated control of germination by TCP14/15 is regulated through EXPA-mediated control of cell wall loosening, providing a mechanistic explanation for this phenotype and a previously undescribed role for TCPs in the control of cell expansion. This network reveals the paths of signal integration that culminate in seed germination and provides a resource to uncover links between the genetic and biomechanical bases of plant growth.

GROWTH-REGULATING FACTORS Interact with DELLAs and Regulate Growth in Cold Stress.

DELLA proteins are repressors of the gibberellin (GA) hormone signaling pathway that act mainly by regulating transcription factor activities in plants. GAs induce DELLA repressor protein degradation and thereby control a number of critical developmental processes as well as responses to stresses such as cold. The strong effect of cold temperatures on many physiological processes has rendered it difficult to assess, based on phenotypic criteria, the role of GA and DELLAs in plant growth during cold stress. Here, we uncover substantial differences in the GA transcriptomes between plants grown at ambient temperature (21°C) and plants exposed to cold stress (4°C) in Arabidopsis (Arabidopsis thaliana). We further identify over 250, to the largest extent previously unknown, DELLA-transcription factor interactions using the yeast two-hybrid system. By integrating both data sets, we reveal that most members of the nine-member GRF (GROWTH REGULATORY FACTOR) transcription factor family are DELLA interactors and, at the same time, that several GRF genes are targets of DELLA-modulated transcription after exposure to cold stress. We find that plants with altered GRF dosage are differentially sensitive to the manipulation of GA and hence DELLA levels, also after cold stress, and identify a subset of cold stress-responsive genes that qualify as targets of this DELLA-GRF regulatory module.

Largely additive effects of gibberellin and strigolactone on gene expression in Arabidopsis thaliana seedlings.

The phytohormones gibberellin (GA) and strigolactone (SL) are involved in essential processes in plant development. Both GA and SL signal transduction mechanisms employ α/ß-hydrolase-derived receptors that confer E3 ubiquitin ligase-mediated protein degradation processes. This suggests a common evolutionary origin of these pathways and possibly a molecular interaction between them. One such indication stems from rice, where the DELLA protein of the GA pathway was reported to interact with the SL receptor. Here, we examine the physiological interaction between both pathways through the analysis of GA (ga1) and SL biosynthesis (max1 and max3) mutants. In ga1 max double mutants, we find indications only for additive interactions when examining several phenotypic readouts. We further identify short-term transcriptional responses to GA and the synthetic SL rac-GR24 through next-generation sequencing of poly-adenylated RNAs (RNA-seq) in ga1 max1. Remarkably, both hormones lead to predominantly additive transcriptional changes of a largely overlapping set of genes. The expression of only a few genes was altered in a synergistic manner but, interestingly, these include the genes encoding the GA catabolic enzyme GA2 OXIDASE2 (GA2ox2) as well as the SL pathway regulators BRANCHED1 (BRC1) and SUPPRESSOR OF max2 1-LIKE8 (SMXL8). We conclude that GA and rac-GR24 signaling in Arabidopsis seedlings converge at the level of transcription of a common gene-set.