Transcription Factor ABI5 Research Articles

BBX30/miP1b and BBX31/miP1a form a positive feedback loop with ABI5 to regulate ABA-mediated post-germination seedling growth arrest.

In plants the switch to autotrophic growth involves germination followed by post-germination seedling establishment. When environmental conditions are not favorable, the stress hormone Abscisic Acid (ABA) signals plants to postpone seedling establishment by inducing the expression of the transcription factor ABI5. The levels of ABI5 determine the efficiency of the ABA mediated post-germination developmental growth arrest. The molecular mechanisms regulating the stability and activity of ABI5 during the transition to light are less known. Using genetic, molecular and biochemical approach, we found that two B-box domain containing proteins BBX31 and BBX30 alongwith ABI5 inhibit post-germination seedling establishment in a partially interdependent manner. BBX31 and BBX30 are also characterized as microProteins miP1a and miP1b respectively, based on their small size, single domain and ability to interact with multi-domain proteins. miP1a/BBX31 and miP1b/BBX30 physically interact with ABI5 to stabilize it and promote its binding to promoters of downstream genes. ABI5 reciprocally induces the expression of BBX30 and BBX31 by directly binding to their promoter. ABI5 and the two microProteins thereby form a positive feedback loop to promote ABA-mediated developmental arrest of seedlings.

Impacts of drought and elevated temperature on the seeds of malting barley.

High seed quality is key to agricultural production, which is increasingly affected by climate change. We studied the effects of drought and elevated temperature during seed production on key seed quality traits of two genotypes of malting barley (Hordeum sativum L.). Plants of a "Hana-type" landrace (B1) were taller, flowered earlier and produced heavier, larger and more vigorous seeds that resisted ageing longer compared to a semi-dwarf breeding line (B2). Accordingly, a NAC domain-containing transcription factor (TF) associated with rapid response to environmental stimuli, and the TF ABI5, a key regulator of seed dormancy and vigour, were more abundant in B1 seeds. Drought significantly reduced seed yield in both genotypes, and elevated temperature reduced seed size. Genotype B2 showed partial thermodormancy that was alleviated by drought and elevated temperature. Metabolite profiling revealed clear differences between the embryos of B1 and B2. Drought, but not elevated temperature, affected the metabolism of amino acids, organic acids, osmolytes and nitrogen assimilation, in the seeds of both genotypes. Our study may support future breeding efforts to produce new lodging and drought resistant malting barleys without trade-offs that can occur in semi-dwarf varieties such as lower stress resistance and higher dormancy.

Regulatory dynamics distinguishing desiccation tolerance strategies within resurrection grasses.

Desiccation tolerance has evolved recurrently in grasses using two unique strategies of either protecting or dismantling the photosynthetic apparatus to minimize photooxidative damage under life without water (anhydrobiosis). Here, we surveyed chromatin architecture and gene expression during desiccation in two closely related grasses with distinguishing desiccation tolerance strategies to identify regulatory dynamics underlying these unique adaptations. In both grasses, we observed a strong association between nearby chromatin accessibility and gene expression in desiccated tissues compared to well-watered, reflecting an unusual chromatin stability under anhydrobiosis. Integration of chromatin accessibility (ATACseq) and expression data (RNAseq) revealed a core desiccation response across these two grasses. This includes many genes with binding sites for the core seed development transcription factor ABI5, supporting the long-standing hypothesis that vegetative desiccation tolerance evolved from rewiring seed pathways. Oropetium thomaeum has a unique set of desiccation induced genes and regulatory elements associated with photoprotection, pigment biosynthesis, and response to high light, reflecting its adaptation of protecting the photosynthetic apparatus under desiccation (homoiochlorophyly). By contrast, Eragrostis nindensis has unique accessible and expressed genes related to chlorophyll catabolism, scavenging of amino acids, and hypoxia, highlighting its poikilochlorophyllous adaptations of dismantling the photosynthetic apparatus and degrading chlorophyll under desiccation. Together, our results highlight the complex regulatory and expression dynamics underlying desiccation tolerance in grasses.

The RNA N6 -methyladenosine demethylase ALKBH9B modulates ABA responses in Arabidopsis.

The mRNA modification N6 -methyladenosine (m6 A) plays vital roles in plant development and biotic and abiotic stress responses. The RNA m6 A demethylase ALKBH9B can remove m6 A in alfalfa mosaic virus RNA and plays roles in alfalfa mosaic virus infection in Arabidopsis. However, it is unknown whether ALKBH9B also exhibits demethylation activity and has a biological role in endogenous plant mRNA. We demonstrated here that mRNA m6 A modification is induced by the phytohormone abscisic acid (ABA) and that ALKBH9B has m6 A demethylation activity on endogenous mRNA. Knocking out ALKBH9B led to hypersensitivity to ABA treatment during seed germination and early seedling development. We further showed that ALKBH9B removes the m6 A modification in the ABA INSENSITIVE 1 (ABI1) and BRI1-EMS-SUPPRESSOR 1 (BES1) transcripts following ABA treatment, affecting the stability of these mRNAs. Furthermore, we determined that ALKBH9B acts genetically upstream of the transcription factors ABI3 and ABI5, and its regulatory function in ABA responses depended on ABI3 and ABI5. Our findings reveal the important roles of the m6 A modification in ABA responses and highlight the role of ALKBH9B-mediated m6 A demethylation in regulating ABA responses post-transcriptionally.

Seed-to-Seedling Transition in Pisum sativum L.: A Transcriptomic Approach.

The seed-to-seedling transition is a crucial step in the plant life cycle. The transition occurs at the end of seed germination and corresponds to the initiation of embryonic root growth. To improve our understanding of how a seed transforms into a seedling, we germinated the Pisum sativum L. seeds for 72 h and divided them into samples before and after radicle protrusion. Before radicle protrusion, seeds survived after drying and formed normally developed seedlings upon rehydration. Radicle protrusion increased the moisture content level in seed axes, and the accumulation of ROS first generated in the embryonic root and plumule. The water and oxidative status shift correlated with the desiccation tolerance loss. Then, we compared RNA sequencing-based transcriptomics in the embryonic axes isolated from pea seeds before and after radicle protrusion. We identified 24,184 differentially expressed genes during the transition to the post-germination stage. Among them, 2101 genes showed more prominent expression. They were related to primary and secondary metabolism, photosynthesis, biosynthesis of cell wall components, redox status, and responses to biotic stress. On the other hand, 415 genes showed significantly decreased expression, including the groups related to water deprivation (eight genes) and response to the ABA stimulus (fifteen genes). We assume that the water deprivation group, especially three genes also belonging to ABA stimulus (LTI65, LTP4, and HVA22E), may be crucial for the desiccation tolerance loss during a metabolic switch from seed to seedling. The latter is also accompanied by the suppression of ABA-related transcription factors ABI3, ABI4, and ABI5. Among them, HVA22E, ABI4, and ABI5 were highly conservative in functional domains and showed homologous sequences in different drought-tolerant species. These findings elaborate on the critical biochemical pathways and genes regulating seed-to-seedling transition.

Studies on the interactions of AFPs and bZIP transcription factor ABI5

AFP1 interacts with ABI5 and negatively regulates the abscisic acid (ABA) signaling by accelerating ABI5's degradation during the seed germination phase in Arabidopsis, but the underlying mechanism remains unclear. Moreover, the molecular basis of the interaction between AFP1 homologs and ABI5 has yet to be elucidated. In this study, the patterns of their interactions with ABI5 were investigated in detail. We found that AFP2/3/4 can bind two regions of ABI5, one is ABI51aa to 135aa and another is ABI5202aa to 213aa. However, AFP1 only interacts with the second region of ABI5, i.e. ABI5202aa to 213aa. Prior research has shown that ABI51aa to 135aa is related to the transcriptional activity of ABI5. Thus, our results suggest that AFPs may also modulate ABI5, by directly binding to its transcriptional activation domain, thereby influencing its transcriptional activity. Further, interactions between AFPs and ABI5 were not affected if the Ser42th in the ABI5-SnRK2 motif were mutated respectively to Glu or Ala. Nevertheless, interactions between AFPs and ABI5 were eliminated if the Thr47th and Thr206th of ABI5 were mutated respectively to Glu or Ala. Since the two residues of Thr47th and Thr206th were located in the phosphorylation motifs of CKII, AFPs might regulate the activities of ABI5 transcription factor through a CKII-dependent pathway.

ENAP1 retrains seed germination via H3K9 acetylation mediated positive feedback regulation of ABI5.

Histone acetylation is involved in the regulation of seed germination. The transcription factor ABI5 plays an essential role in ABA- inhibited seed germination. However, the molecular mechanism of how ABI5 and histone acetylation coordinate to regulate gene expression during seed germination is still ambiguous. Here, we show that ENAP1 interacts with ABI5 and they co-bind to ABA responsive genes including ABI5 itself. The hypersensitivity to ABA of ENAP1ox seeds germination is recovered by the abi5 null mutation. ABA enhances H3K9Ac enrichment in the promoter regions as well as the transcription of target genes co-bound by ENAP1 and ABI5, which requires both ENAP1 and ABI5. ABI5 gene is directly regulated by ENAP1 and ABI5. In the enap1 deficient mutant, H3K9Ac enrichment and the binding activity of ABI5 in its own promoter region, along with ABI5 transcription and protein levels are all reduced; while in the abi5-1 mutant, the H3K9Ac enrichment and ENAP1 binding activity in ABI5 promoter are decreased, suggesting that ENAP1 and ABI5 function together to regulate ABI5- mediated positive feedback regulation. Overall, our research reveals a new molecular mechanism by which ENAP1 regulates H3K9 acetylation and mediates the positive feedback regulation of ABI5 to inhibit seed germination.

High oil accumulation in tuber of yellow nutsedge compared to purple nutsedge is associated with more abundant expression of genes involved in fatty acid synthesis and triacylglycerol storage

BackgroundYellow nutsedge is a unique plant species that can accumulate up to 35% oil of tuber dry weight, perhaps the highest level observed in the tuber tissues of plant kingdom. To gain insight into the molecular mechanism that leads to high oil accumulation in yellow nutsedge, gene expression profiles of oil production pathways involved carbon metabolism, fatty acid synthesis, triacylglycerol synthesis, and triacylglycerol storage during tuber development were compared with purple nutsedge, the closest relative of yellow nutsedge that is poor in oil accumulation.ResultsCompared with purple nutsedge, high oil accumulation in yellow nutsedge was associated with significant up-regulation of specific key enzymes of plastidial RubisCO bypass as well as malate and pyruvate metabolism, almost all fatty acid synthesis enzymes, and seed-like oil-body proteins. However, overall transcripts for carbon metabolism toward carbon precursor for fatty acid synthesis were comparable and for triacylglycerol synthesis were similar in both species. Two seed-like master transcription factors ABI3 and WRI1 were found to display similar transcript patterns but were expressed at 6.5- and 14.3-fold higher levels in yellow nutsedge than in purple nutsedge, respectively. A weighted gene co-expression network analysis revealed that ABI3 was in strong transcriptional coordination with WRI1 and other key oil-related genes.ConclusionsThese results implied that pyruvate availability and fatty acid synthesis in plastid, along with triacylglycerol storage in oil bodies, rather than triacylglycerol synthesis in endoplasmic reticulum, are the major factors responsible for high oil production in tuber of yellow nutsedge, and ABI3 most likely plays a critical role in regulating oil accumulation. This study is of significance with regard to understanding the molecular mechanism controlling carbon partitioning toward oil production in oil-rich tuber and provides a valuable reference for enhancing oil accumulation in non-seed tissues of crops through genetic breeding or metabolic engineering.

Bacterial Quorum-Sensing Signaling-Related drr1 Mutant Influences Abscisic Acid Responsiveness in Arabidopsis thaliana L.

N-acyl-L-homoserine lactones (AHLs) are involved in cell-to-cell communication in Gram-negative bacteria through a process termed quorum-sensing (QS). In this report, we evaluated the response of Arabidopsis thaliana primary roots to abscisic acid (ABA) in wild-type (WT) and decanamide resistant root 1 (drr1) mutant, previously reported to be resistant to N-decanoyl-L-homoserine lactone (C10-HL). When compared to WT seedlings, drr1 mutants were hypersensitive to ABA and had primary roots shorter, which correlated with lower cell division in meristems, a higher concentration of endogenous ABA, and a greater expression of ABI5 gene; and this shortened primary root phenotype was reversed in drr1abi5 double mutants. An analysis of expression of ABSCISIC ACID INSENSITIVE 4 (ABI4) showed an ABA-inducible pattern in primary root tips, which was further increased in drr1 mutant seedlings. Comparison of seed germination in WT, drr1, abi5, and drr1abi5 lines showed higher germination percentages in the following order under control condition: abi5 > drr1abi5 > WT > drr1, while abi5 and drr1abi5 germinated faster and drr1 germinated slower with respect to WT under ABA condition. Taken together, our results suggest that DRR1 is a negative regulator of ABA signaling probably acting upstream of the transcription factors ABI4 and ABI5, which influence ABA responsiveness in primary roots and seed germination.

Abscisic acid-mediated induction of FLAVIN-CONTAINING MONOOXYGENASE 2 leads to reduced accumulation of methylthioalkyl glucosinolates in Arabidopsis thaliana

Side-chain modification contributes to the structural diversity of aliphatic glucosinolates (GSLs), a class of sulfur-containing secondary metabolites found in Brassicales. The first step in side-chain modification of aliphatic GSLs is the S-oxygenation of the methylthioalkyl (MT) moiety to the methylsulfinylalkyl (MS) moiety. This reaction is catalyzed by flavin-containing monooxygenase (FMOGS-OX), which is encoded by seven genes in Arabidopsis thaliana. Therefore, the regulation of FMOGS-OX gene expression is key to controlling side-chain structural diversity. In this study, we demonstrated that the expression of FMOGS-OX2 and FMOGS-OX4 was induced by glucose treatment, independent of MYB28/29 and MYC2/3/4, the transcription factors that positively regulate aliphatic GSL biosynthesis. Glucose treatment of the abscisic acid (ABA)-related mutants indicated that glucose-triggered upregulation of FMOGS-OX2 and FMOGS-OX4 was partially regulated by ABA through the key negative regulators ABI1 and ABI2, and the positive regulator SnRK2, but not via the transcription factor ABI5. In wild-type plants, glucose treatment drastically reduced the accumulation of 4-methylthiobutyl (4MT) GSL, whereas a decrease in 4MT GSL was not observed in the fmogs-ox2, abi1-1, abi2-1, aba2-1, or aba3-1 mutants. This result indicated that the decreased accumulation of 4MT GSL by glucose treatment was attributed to upregulation of FMOGS-OX2 via the ABA signaling pathway.

The effect of ABRE BINDING FACTOR 4-mediated FYVE1 on salt stress tolerance in Arabidopsis

FYVE1 encodes a protein that is localized to the peripheral membrane of late endosomal compartments, and is involved in the regulation of mulitivesicular/prevacuolar compartment protein sorting. It was found that FYVE1 attenuates ABA signaling through degrading ABA receptors PYR1 and PYL4 by ESCRT pathway, and also interacts with transcription factors ABF4 and ABI5 to transcriptionally inhibit ABA signaling pathway by reducing their binding to the cis-regulatory sequences of their downstream genes. However, the mechanisms underlying the transcriptional regulation of FYVE1 and its biological function in salt stress are largely unknown. Here, we show that fyve1 knockdown-mutants show enhanced tolerance to salt stress, while overexpression of FYVE1 results in increased sensitivity to salt stress. Further analysis shows that FYVE1 negatively regulates salt stress tolerance, which is associated with ABA signaling pathway. ABRE BINDING FACTOR 4 (ABF4) directly binds to promoter of FYVE1 to activate its transcription. Moreover, FYVE1 interacts with and promotes degradation of all ABA PYR/PYL receptors. Thus, our results suggest that FYVE1 negatively modulates salt stress tolerance in Arabidopsis via a negative feedback loop.

Control of seed coat rupture by ABA-INSENSITIVE 5 inArabidopsis thaliana

AbstractIn Arabidopsis, seed germination is a biphasic process involving rupture of the seed coat followed by emergence of the radicle through the micropylar endosperm. Embryo expansion results in seed coat rupture and removal of seed coat imposed dormancy with DELLA proteins blocking embryo expansion in the absence of gibberellins. Exogenous abscisic acid (ABA) treatment does not block seed coat rupture but does block radicle emergence. We used this limited effect of exogenous ABA to further investigate the mechanism by which it blocks the onset of germination marked by seed coat rupture. We show that physical nicking of the seed coat results in exogenous ABA treatment blocking both seed coat and endosperm rupture and this block requires the transcription factors ABI3 and ABI5, but not ABI4. Furthermore, we show that the repression of expression of severalEXPANSINgenes (EXPA1,EXPA2,EXPA3,EXPA9andEXPA20) by exogenous ABA requires ABI5. We conclude that ABI5 plays an important role in the ABA-mediated repression of germination through prevention of seed coat rupture and propose that this involves EXPANSIN related control of cell wall loosening.

The β5 subunit is essential for intact 26S proteasome assembly to specifically promote plant autotrophic growth under salt stress.

The ubiquitin 26S proteasome (26SP) system efficiently degrades many key regulators of plant development. 26SP consists of two subcomplexes: the catalytic 20S core particle (CP) and the 19S regulatory particle (RP). Previous studies have focused on 19S RP; whether there is a specific subunit in 20S CP that has a stress-related biological function in plants is unclear. PBE1, one of the β5 subunits of Arabidopsis proteasome CP, is essential for the assembly and proteolytic activity of 26SP in salt-stressed seedlings. The expression of PBE1 is stress-induced. During the transition from seed germination to autotrophic growth in salt-stressed seedlings, loss of PBE1 function results specifically in arrest in developmental transition but not in germination and post-germination growth. PBE1 is also important for other types of proteasome stress and Endoplasmic Reticulum (ER) stress. PBE1 modulates the protein level of the transcription factor ABI5 and thereby down-regulates the expression of several genes downstream of this key regulator which are known to be essential for plant growth under stress. Collectively, our results showed PBE1-mediated intact proteasome assembly that is essential for successful autotrophic growth, and revealed how PBE1 mediated stress proteasome functions to control both proteasome activity and abscisic acid (ABA)-mediated stress signaling in plants.

ABI4 regulates the floral transition independently of ABI5 and ABI3

Emerging evidence suggests that the stress hormone abscisic acid (ABA) is also involved in the floral transition control. The transcription factors ABA INSENSITIVE4 (ABI4) and ABI5 negatively regulate flowering by directly promoting FLOWERING LOCUS C expression, and ABI3 also negatively regulates the floral transition. However, the genetic relationships between ABI4 and both ABI5 and ABI3 remain elusive. Here, we generated transgenic plants overexpressing ABI4 in the abi5 (OE-ABI4::abi5) and abi3 backgrounds (OE-ABI4::abi3). The flowering phenotypic analysis demonstrated that OE-ABI4::abi5 and OE-ABI4::abi3 plants exhibited delayed flowering. These findings suggest that ABI4 independently regulates floral transition but not through ABI5 and ABI3 cascades.

The Asparagine-Rich Protein NRP Facilitates the Degradation of the PP6-type Phosphatase FyPP3 to Promote ABA Response in Arabidopsis

The phytohormone abscisic acid (ABA) plays critical roles in abiotic stress responses and plant development. In germinating seeds, the phytochrome-associated protein phosphatase, FyPP3, negatively regulates ABA signaling by dephosphorylating the transcription factor ABI5. However, whether and how FyPP3 is regulated at the posttranscriptional level remains unclear. Here, we report that an asparagine-rich protein, NRP, interacts with FyPP3 and tethers FyPP3 to SYP41/61-positive endosomes for subsequent degradation in the vacuole. Upon ABA treatment, the expression of NRP was induced and NRP-mediated FyPP3 turnover was accelerated. Consistently, ABA-induced FyPP3 turnover was abolished in an nrp null mutant. On the other hand, FyPP3 can dephosphorylate NRP in vitro, and overexpression of FyPP3 reduced the half-life of NRP in vivo. Genetic analyses showed that NRP has a positive role in ABA-mediated seed germination and gene expression, and that NRP is epistatic to FyPP3. Taken together, our results identify a new regulatory circuit in the ABA signaling network, which links the intracellular trafficking with ABA signaling.

CsWRKY46, a WRKY transcription factor from cucumber, confers cold resistance in transgenic-plant by regulating a set of cold-stress responsive genes in an ABA-dependent manner

Plant WRKY transcription factors are trans-regulatory proteins that are involved in plant immune responses, development and senescence; however, their roles in abiotic stress are still not well understood, especially in the horticultural crop cucumber. In this study, a novel cucumber WRKY gene, CsWRKY46 was cloned and identified, which was up-regulated in response to cold stress and exogenous abscisic acid (ABA) treatment. CsWRKY46 is belonging to group II of the WRKY family, CsWRKY46 was found exclusively in the nucleus, as indicated by a transient expression assay. Yeast one-hybrid assay shown that CsWRKY46 interact with the W-box in the promoter of ABI5. Transgenic Arabidopsis lines over-expressing CsWRKY46, WRK46-OE1 and WRK46-OE5 had higher seedling survival rates upon freezing treatment compared with that of the wild-type. The above over-expression lines also showed much a higher proline accumulation, less electrolyte leakage and lower malondialdehyde (MDA) levels. Furthermore, the CsWRKY46 overexpression lines were hypersensitive to ABA during seed germination, but the seedlings were not. Quantitative RT-PCR analyses revealed that the expression levels of the ABA-responsive transcription factor ABI5 were higher in the WRKY46-OE lines than in wild-type and that the overexpression of CsWRKY46 increased the expression of stress-inducible genes, including RD29A and COR47. Taken together, our results demonstrated that CsWRKY46 from cucumber conferred cold tolerance to transgenic plants and positively regulated the cold signaling pathway in an ABA-dependent manner.

Modulation of the Fragrance of Kam Sweet Rice by Jasmonic Acid and Abscisic Acid Might Involve the MYC2 and ABI5 Transcription Factors

Volatile compounds were analyzed using gas chromatography-mass spectrometry (GC-MS) and gas chromatography-olfactometry (GC-O). Results showed that nonanal had the highest odor activity value in the filling grains. Nonanal was enhanced by application of JA and inhibited by ABA in the seedlings of Kam sweet rice. C9-aldehydes are significant odorants in many rice types and known products of the 9-lipoxygenase and 9-hydroperoxide lyases encoded by the OsLOX3 and OsHPL1 genes, respectively. OsLOX3 mRNA expression was higher in aromatic rice compared to a nonaromatic cultivar. mRNA expression of ABI5 and MYC2 negatively correlated with OsLOX3 expression (r = −0.841, P < 0.01; and r = −0.512, P < 0.05). In conclusion, the transcription factors ABI5 and MYC2 might be involved in ABA and JA modulation of nonanal biosynthesis, which could be used to improve rice quality. Practical Applications Nonanal and 2-acetyl-1-pyrroline are important and potent flavor compounds in aromatic Kam sweet rice. The OsLOX3 gene was potentially identified as a critical gene in aromatic Kam sweet rice and the enhancing effect of JA on nonanal biosynthesis was confirmed. The transcription factors ABI5 and MYC2 seem to regulate the expression of the fragrance gene OsLOX3 while MYC2 act as a positive regulator involved in JA-mediated nonanal biosynthesis. ABI5 participate in the ABA signaling pathway to down-regulate the expression of OsLOX3. These results will help understand the pathways involved in the biosynthesis of rice fragrance, which could be used to improve the quality of rice.

A hub for ABA signaling to the nucleus: Significance of a cytosolic and nuclear dual-localized PPR protein SOAR1 acting downstream of Mg-chelatase H subunit

SOAR1 is a cytosol-nucleus dual-localized pentatricopeptide repeat (PPR) protein, which we indentified recently as a crucial regulator in the CHLH/ABAR (Mg-chelatase H subunit /putative ABA receptor)-mediated signaling pathway, acting downstream of CHLH/ABAR and upstream of a nuclear ABA-responsive bZIP transcription factor ABI5. Downregulation and upregulation of SOAR1 expression alter dramatically both ABA sensitivity and expression of a subset of key, nuclear ABA-responsive genes, suggesting that SOAR1 is a hub for ABA signaling to the nucleus, and CHLH/ABAR mediates a central signaling pathway to regulate downstream gene expression through SOAR1.

Arabidopsis pentatricopeptide repeat protein SOAR1 plays a critical role in abscisic acid signalling.

A dominant suppressor of the ABAR overexpressor, soar1-1D, from CHLH/ABAR [coding for Mg-chelatase H subunit/putative abscisic acid (ABA) receptor (ABAR)] overexpression lines was screened to explore the mechanism of the ABAR-mediated ABA signalling. The SOAR1 gene encodes a pentatricopeptide repeat (PPR) protein which localizes to both the cytosol and nucleus. Down-regulation of SOAR1 strongly enhances, but up-regulation of SOAR1 almost completely impairs, ABA responses, revealing that SOAR1 is a critical, negative, regulator of ABA signalling. Further genetic evidence supports that SOAR1 functions downstream of ABAR and probably upstream of an ABA-responsive transcription factor ABI5. Changes in the SOAR1 expression alter expression of a subset of ABA-responsive genes including ABI5. These findings provide important information to elucidate further the functional mechanism of PPR proteins and the complicated ABA signalling network.

In vitro binding of Sorghum bicolor transcription factors ABI4 and ABI5 to a conserved region of a GA 2-OXIDASE promoter: possible role of this interaction in the expression of seed dormancy.

The precise adjustment of the timing of dormancy release according to final grain usage is still a challenge for many cereal crops. Grain sorghum [Sorghum bicolor (L.) Moench] shows wide intraspecific variability in dormancy level and susceptibility to pre-harvest sprouting (PHS). Both embryo sensitivity to abscisic acid (ABA) and gibberellin (GA) metabolism play an important role in the expression of dormancy of the developing sorghum grain. In previous works, it was shown that, simultaneously with a greater embryo sensitivity to ABA and higher expression of SbABA-INSENSITIVE 4 (SbABI4) and SbABA-INSENSITIVE 5 (SbABI5), dormant grains accumulate less active GA4 due to a more active GA catabolism. In this work, it is demonstrated that the ABA signalling components SbABI4 and SbABI5 interact in vitro with a fragment of the SbGA 2-OXIDASE 3 (SbGA2ox3) promoter containing an ABA-responsive complex (ABRC). Both transcription factors were able to bind the promoter, although not simultaneously, suggesting that they might compete for the same cis-acting regulatory sequences. A biological role for these interactions in the expression of dormancy of sorghum grains is proposed: either SbABI4 and/or SbABI5 activate transcription of the SbGA2ox3 gene in vivo and promote SbGA2ox3 protein accumulation; this would result in active degradation of GA4, thus preventing germination of dormant grains. A comparative analysis of the 5′-regulatory region of GA2oxs from both monocots and dicots is also presented; conservation of the ABRC in closely related GA2oxs from Brachypodium distachyon and rice suggest that these species might share the same regulatory mechanism as proposed for grain sorghum.