AFP2 inhibits ABA responses during germination without ABI5 degradation but DWAs reduce desiccation tolerance

Overexpression of ABI5/ABF interacting proteins (AFPs) results in extreme ABA resistance of seeds and failure to acquire desiccation tolerance, at least in part through effects on chromatin modification. This study tests the hypothesis that the AFPs promote germination by also functioning as adapters for E3 ligases that ubiquitinate ABI5, leading to its degradation. Interactions between AFPs and two well-characterized classes of E3 ligases targeting ABI5, DWD HYPERSENSITIVE TO ABA (DWA)s and KEEP ON GOING (KEG), were analyzed by yeast two-hybrid, bimolecular fluorescence complementation, and genetic assays. Although the AFPs and E3 ligases showed weak direct interactions, loss of function for the E3 ligases did not impair ABA-resistance conferred by overexpression of the YFP-AFP2 fusion. Comparison of ABI5 and AFP2 levels in these lines showed that AFP2 accumulation increased during germination, but that ABI5 degradation followed germination, demonstrating that AFP2 controls ABA sensitivity during germination independently of ABI5 degradation. Surprisingly, AFP2 overexpression in the dwa1 dwa2 mutant background produced the unusual combination of extreme ABA resistance and desiccation tolerance, creating an opportunity to separate the underlying biochemical characteristics of ABA sensitivity and desiccation tolerance that we investigated by quantitative proteomics. Our analysis identified at least three-fold more differentially accumulated seed proteins than previous studies. Comparison of dry seed proteomes of the different genotypes allowed us to separate and refine the changes in protein accumulation patterns correlating with desiccation tolerance independently of ABA sensitivity, or vice versa, to a subset of cold-induced and defense stress-responsive proteins and signaling regulators.

Regulation of drought tolerance in Arabidopsis involves PLATZ4-mediated transcriptional suppression of PIP2;8

PLATZ transcription factors play important roles in plant growth, development, biotic and abiotic stress responses. However, how PLATZ regulates plant drought tolerance and ABA sensitivity remains largely unknown. Here, we show that PLATZ4 increases drought tolerance and ABA sensitivity in Arabidopsis thaliana by suppressing the expression of PIP2;8, while upregulating expression of ABI3, ABI4 and ABI5. PLATZ4 directly binds A/T-rich sequences within the PIP2;8 promoter. Consistent with this, PIP2;8 acts epistatically to PLATZ4. Furthermore, the aquaporin activity of PIP2;8 was confirmed in Xenopus laevis oocytes in response to osmotic stress. Analysis of water loss of seedlings overexpressing PIP2;8 or lacking PIP2;8 function indicated that PIP2;8-mediated water flow is particularly active in response to drought stress in planta. In platz4 mutant and PLATZ4-overexpressing plants, water loss and stomatal closure changed oppositely to those in pip2;8 mutants and PIP2;8-overexpressing plants, respectively. In addition, the interaction between PLATZ4 and AITR6 was confirmed by several assays, and the binding of PIP2;8 promoter by PLATZ4 was strengthened by an interaction with AITR6. Collectively, our findings reveal that PLATZ4 interacts with AITR6 to increase ABA sensitivity and drought tolerance by upregulating expression of ABI3, ABI4 and ABI5 while inhibiting the expression of PIP2;8 and associated genes.

Genome-wide association study revealed genetic variations of ABA sensitivity controlled by multiple stress-related genes in rice

Abscisic acid (ABA) is a critical phytohormone that regulates multiple physiological processes including plant growth and stress tolerance. The core ABA signaling pathway has been well established, but genetic variations mediating ABA responses remain largely unknown. In this study, we performed genome-wide association study (GWAS) to identify loci and genes associated with ABA sensitivity (reflected by seed germination inhibition by ABA) in a panel of 425 rice accessions. The seed germination assay revealed that Aus and indica rice had stronger ABA sensitivity than japonica rice. A total of 48 non-redundant association loci were detected in the indica subpopulation and whole population, and 386 genes in these loci were responsive to ABA or abiotic stresses. Eight association loci were overlapped with previously reported loci for yield under drought stress or for drought-indicative image traits. Haplotype analyses of important candidate genes such as OsSAPK6, a key component in the ABA signaling core, were performed to identify key SNPs/InDels that may affect gene functions through promoter activity regulation, amino acid variation, or gene splicing. These results provide insights into the genetic basis of ABA sensitivity related to stress responses.

Increased Abscisic Acid Sensitivity And Drought Stress By Overexpression of Abscisic Acid Receptors In Arabidopsis Thaliana

Abscisic acid (ABA) is a key plant hormone that regulates plant growth development and stress response. ABA is recognized and bound by ABA Receptor PYR/PYL/RCAR (referred to as PYLs). However, little is known about the PYLs gene family in Populus euphratica. Here, we identified 12 PYLs in P. euphratica and named PePYL1-12. Phylogenetic analysis divided the 12 PePYLs into three subfamilies. Subcellular localization showed that PePYL2, PePYL4, PePYL5, PePYL6, and PePYL9 were located in the cytoplasm and nucleus, PePYL10 localized in the nucleus. The promoter of 12 PePYLs contains hormones- and abiotic stress-related cis-acting elements. Moreover, ABA and drought significantly up-regulation the expression of PePYL6 and PePYL9. To study the performance of PePYLs under ABA and drought stress, we generated transgenic Arabidopsis plants overexpressing PePYL6 and PePYL9. Compared with wild type, transgenic Arabidopsis enhanced ABA sensitivity during seed germination and root growth, improved water use efficiency and drought resistance. Taken together, our results confirmed that PePYL6 and PePYL9 play a positive role in ABA-mediated stress responses in P. euphratica.

Comparative transcriptomic and physiological analyses of weedy rice and cultivated rice to identify vital differentially expressed genes and pathways regulating the ABA response

Weedy rice is a valuable germplasm resource characterized by its high tolerance to both abiotic and biotic stresses. Abscisic acid (ABA) serves as a regulatory signal in plant cells as part of their adaptive response to stress. However, a global understanding of the response of weedy rice to ABA remains to be elucidated. In the present study, the sensitivity to ABA of weedy rice (WR04-6) was compared with that of temperate japonica Shennong9816 (SN9816) in terms of seed germination and post-germination growth via the application of exogenous ABA and diniconazole, an inhibitor of ABA catabolism. Physiological analysis and a transcriptomic comparison allowed elucidation of the molecular and physiological mechanisms associated with continuous ABA and diniconazole treatment. WR04-6 was found to display higher ABA sensitivity than SN9816, resulting in the rapid promotion of antioxidant enzyme activity. Comparative transcriptomic analyses indicated that the number of differentially expressed genes (DEGs) in WR04-6 seedlings treated with 2 μM ABA or 10 μM diniconazole was greater than that in SN9816 seedlings. Genes involved in stress defense, hormone signal transduction, and glycolytic and citrate cycle pathways were highly expressed in WR04-6 in response to ABA and diniconazole. These findings provide new insight into key processes mediating the ABA response between weedy and cultivated rice.

An ABA-GA bistable switch can account for natural variation in the variability of Arabidopsis seed germination time

Genetically identical plants growing in the same conditions can display heterogeneous phenotypes. Here we use Arabidopsis seed germination time as a model system to examine phenotypic variability and its underlying mechanisms. We show extensive variation in seed germination time variability between Arabidopsis accessions and use a multiparent recombinant inbred population to identify two genetic loci involved in this trait. Both loci include genes implicated in modulating abscisic acid (ABA) sensitivity. Mutually antagonistic regulation between ABA, which represses germination, and gibberellic acid (GA), which promotes germination, underlies the decision to germinate and can act as a bistable switch. A simple stochastic model of the ABA-GA network shows that modulating ABA sensitivity can generate the range of germination time distributions we observe experimentally. We validate the model by testing its predictions on the effects of exogenous hormone addition. Our work provides a foundation for understanding the mechanism and functional role of phenotypic variability in germination time.

The Arabidopsis SMALL AUXIN UP RNA32 Protein Regulates ABA-Mediated Responses to Drought Stress

SMALL AUXIN UP-REGULATED RNAs (SAURs) are recognized as auxin-responsive genes involved in the regulation of abiotic stress adaptive growth. Among the growth-limiting factors, water-deficit condition significantly affects plant growth and development. The putative function of SAUR family member AtSAUR32 has the potential to diminish the negative impact of drought stress, but the exact function and mode of action remain unclear in Arabidopsis. In the current study, AtSAUR32 gene was cloned and functionally analyzed. AtSAUR32 localized to the plasma membrane and nucleus was dominantly expressed in roots and highly induced by abscisic acid and drought treatment at certain time points. The stomatal closure and seed germination of saur32 were less sensitive to ABA relative to AtSAUR32-overexpressed line (OE32-5) and wild type (WT). Moreover, the saur32 mutant under drought stress showed increased ion leakage while quantum yield of photosystem II (ΦPSII) and endogenous ABA accumulation were reduced, along with the expression pattern of ABA/stress-responsive genes compared with WT and the OE32-5 transgenic line. Additionally, yeast two-hybrid (Y2H) and bimolecular fluorescence complementation (BiFC) assays showed that AtSAUR32 interacted with clade-A PP2C proteins (AtHAI1 and AtAIP1) to regulate ABA sensitivity in Arabidopsis. Taken together, these results indicate that AtSAUR32 plays an important role in drought stress adaptation via mediating ABA signal transduction.