Comprehensive analyses of ZFP gene family and characterization of expression profiles during plant hormone response in cotton.

BACKGROUND: Zinc finger proteins (ZFPs) containing only a single zinc finger domain play important roles in the regulation of plant growth and development, as well as in biotic and abiotic stress responses. To date, the evolutionary history and functions of the ZFP gene family have not been identified in cotton. RESULTS: In this paper, we identified 29 ZFP genes in Gossypium hirsutum. This gene family was divided into seven subfamilies, 22 of which were distributed over 17 chromosomes. Bioinformatic analysis revealed that 20 GhZFP genes originated from whole genome duplications and two originated from dispersed duplication events, indicating that whole genome duplication is the main force in the expansion of the GhZFP gene family. Most GhZFP8 subfamily genes, except for GhZFP8-3, were highly expressed during fiber cell growth, and were induced by brassinosteroids in vitro. Furthermore, we found that a large number of GhZFP genes contained gibberellic acid responsive elements, auxin responsive elements, and E-box elements in their promoter regions. Exogenous application of these hormones significantly stimulated the expression of these genes. CONCLUSIONS: Our findings reveal that GhZFP8 genes are involved in cotton fiber development and widely induced by auxin, gibberellin and BR, which provides a foundation for the identification of more downstream genes with potential roles in phytohormone stimuli, and a basis for breeding better cotton varieties in the future.

Comprehensive expression analysis of Arabidopsis GA2-oxidase genes and their functional insights.

Bioactive gibberellins (GAs) play multiple roles in plant development and stress responses. GA2-oxidases (GA2oxs) are a class of 2-oxoglutarate-dependent dioxygenases that regulate the deactivation of bioactive GAs. In this study, we investigated the phylogeny and domain structures of the seven GA2ox genes present in the Arabidopsis thaliana genome. Comprehensive expression analysis using translational reporter lines showed that the seven GA2ox genes are differentially expressed during Arabidopsis growth and development: GA2ox1 is specifically expressed in the hypocotyl and lateral root primordium; GA2ox2 is highly expressed in aboveground tissues; GA2ox3 is expressed in the chalazal endosperm of the early embryo sac and inflorescences; GA2ox4 is expressed in the shoot apical meristem and during lateral root initiation; GA2ox6 is expressed in the maturation zone, but not in the meristem or elongating zone of the root; GA2ox7 is constitutively expressed during almost all developmental stages; and GA2ox8 is exclusively expressed in stomatal cells. Overexpression of each of these GA2ox genes inhibited high temperature-induced hypocotyl elongation in both wild-type and elongated hypocotyl 5 plants, which have an elongated hypocotyl phenotype, suggesting that these genes negatively regulate hypocotyl elongation by reducing bioactive GA levels. This study provides a valuable resource for further elucidating the roles of GA2ox genes during different stages of development.

Arabidopsis defense mutant ndr1-1 displays accelerated development and early flowering mediated by the hormone gibberellic acid.

NONRACE-SPECIFIC DISEASE RESISTANCE (NDR1) is a widely characterized gene that plays a key role in defense against multiple bacterial, fungal, oomycete and nematode plant pathogens. NDR1 is required for activation of resistance by multiple NB and LRR-containing (NLR) protein immune sensors and contributes to basal defense. The role of NDR1 in positively regulating salicylic acid (SA)-mediated plant defense responses is well documented. However, ndr1-1 plants flower earlier and show accelerated development in comparison to wild type (WT) Arabidopsis plants, indicating that NDR1 is a negative regulator of flowering and growth. Exogenous application of gibberellic acid (GA) further accelerates the early flowering phenotype in ndr1-1 plants, while the GA biosynthesis inhibitor paclobutrazol attenuated the early flowering phenotype of ndr1-1, but not to WT levels, suggesting partial resistance to paclobutrazol and enhanced GA response in ndr1-1 plants. Mass spectroscopy analyses confirmed that ndr1-1 plants have 30-40% higher levels of GA3 and GA4, while expression of various GA metabolic genes and major flowering regulatory genes is also altered in the ndr1-1 mutant. Taken together this study provides evidence of crosstalk between the ndr1-1-mediated defense and GA-regulated developmental programs in plants.

The hydrogen sulfide signal enhances seed germination tolerance to high temperatures by retaining nuclear COP1 for HY5 degradation.

Seed germination is a critical stage during the initiation of the plant lifecycle and is strongly affected by endogenous phytohormones and environmental stress. High temperature (HT) upregulates endogenous abscisic acid (ABA) to suppress seed germination, and ABA-INSENSITIVE 5 (ABI5) is the key positive regulator in the ABA signal-mediated modulation of seed germination. In plants, hydrogen sulfide (H2S) is a small gas messenger that participates in multiple physiological processes, but its role in seed germination thermotolerance has not been thoroughly elucidated to date. In this study, we found that H2S enhanced the seed germination rate under HT. Moreover, HT accelerates the efflux of the E3 ligase CONSTITUTIVE PHOTOMORPHOGENESIS 1 (COP1) from the nucleus to the cytoplasm, which results in increased nuclear accumulation of ELONG HYPCOTYL 5 (HY5) to activate the expression of ABI5 and thereby suppress seed germination. However, the H2S signal reversed the HT effect, as characterized by increased COP1 in the nucleus, which resulted in increased degradation of HY5 and reduced expression of ABI5 and thereby enhanced the seed germination thermotolerance. Thus, our findings reveal a novel role for the H2S signal in the modulation of seed germination thermotolerance through the nucleocytoplasmic partitioning of COP1 and the downstream HY5 and ABI5 pathways.

Auxin and GA signaling play important roles in the maize response to phosphate deficiency.

Phytohormone signaling is involved in the low-phosphate (LP) response and causes root system changes. To understand the roles of auxin and gibberellic acid (GA) in the maize response to LP stress, inbred line Q319 was used to identify the changes in root morphology and the gene expression response to LP stress with or without exogenous auxin, GA or their inhibitors. The root morphology, IAA and GAs concentration and genes related to the LP response, cell elongation and division, auxin transport and signaling, and GA synthesis and signaling were analyzed. The LP-induced maize root morphological adaption was dependent on changes in the expression of related genes, like IPS1, pht1;1 LPR1b, KRPs, and EXPB1-4. The altered local auxin concentration and signaling were involved in promoting axial root elongation and reducing lateral root density and length under LP conditions, which were regulated by PID and PP2A activity and the auxin signaling pathway. The upregulation of the GA synthesis genes AN1, GA20ox1, and GA20ox2 and the downregulation of the GA inactive genes GA2ox1 and GA2ox2 were observed in maize roots subjected to LP stress, and the increased GA biosynthesis and signaling were involved in root growth. Both hormones participate in LP stress response and jointly regulated root modification and LP acclimation in maize.

Gibberellins Play a Role in Regulating Tomato Fruit Ripening.

Although exogenous applications of gibberellins (GAs) delay tomato ripening, the regulatory mechanisms of GAs in the process have never been well recognized. Here, we report that the concentration of endogenous GAs is declined before the increase of ethylene production in mature-green to breaker stage fruits. We further demonstrate that reductions in GA levels via overexpression of a GA catabolism gene SlGA2ox1 specifically in fruit tissues lead to early ripening. Consistently, we have also observed that application of a GA biosynthetic inhibitor, prohexadione-calcium, at the mature-green stage accelerates fruit ripening, while exogenous GA3 application delays the process. Furthermore, we demonstrate that ethylene biosynthetic gene expressions and ethylene production are activated prematurely in GA-deficient fruits but delayed/reduced in exogenous GA3-treated WT fruits. We also show that the GA deficiency-mediated activation of ethylene biosynthesis is due to the activation of the ripening regulator genes RIN, NOR and CNR. In conclusion, our results demonstrate that GAs play a negative role in tomato fruit ripening.

Hormonal control of parthenocarpic fruit set in 'Rojo Brillante' persimmon (Diospyros kaki Thunb.).

In persimmon (Diospyros kaki Thunb.), one to three waves of fruit abscission can occur. The parthenocarpic cv. Rojo Brillante may abscise close to 50% of flowers, which implies a major economic losses. In order to study this process, 700 flowers were labelled, 600 had the lobes of the calyx removed at three stages to promote abscission. Half of them were also treated with gibberellic acid (GA3; 10 mg l-1) to counteract the effect, and 100 were used as control. In the second year, GA3 (25 mg l-1) was applied to whole trees. Calyx lobe removal (CLR) reduced fruit growth rates and advanced and increased fruitlet abscission, whereas GA3 counteracted this effect. Furthermore, when GA3 was applied to the whole tree, fruit set was increased. The time-course of fruit abscission paralleled a decreased in hormonal and carbohydrate contents. Control fruit showed a peak of gibberellin (GA1 and GA4) and IAA concentration at anthesis. Hexose concentrations remained almost constant from flower bud to fruit set, whereas that of sucrose diminished with time. A peak in ethylene production occurred at anthesis, which increased when CLR was performed prior to or at anthesis, but not when performed at fruit set, when ethylene was markedly smaller. GA3 also counteracted it. Accordingly, we suggest that fruit set depends on the induction of gibberellin (GA) and IAA responses in the persimmon, and since there is no shortage of hormones or carbohydrates at anthesis, ethylene production at anthesis seems the most plausible cause of the physiological fruitlet abscission.

MOTHER-OF-FT-AND-TFL1 represses seed germination under far-red light by modulating phytohormone responses in Arabidopsis thaliana.

Seed germination in many plant species is triggered by sunlight, which is rich in the red (R) wavelength and repressed by under-the-canopy light rich in far red (FR). R:FR ratios are sensed by phytochromes to regulate levels of gibberellins (GAs) and abscisic acid (ABA), which induce and inhibit germination respectively. In this study we have discovered that, under FR light conditions, germination is repressed by MOTHER-OF-FT-AND-TFL1 (MFT) through the regulation of the ABA and GA signaling pathways. We also show that MFT gene expression is tightly regulated by light quality. Previous work has shown that under FR light conditions the transcription factor PHYOCHROME-INTERACTING-FACTOR1 (PIF1) accumulates and promotes expression of SOMNUS (SOM) that, in turn, leads to increased ABA and decreased GA levels. PIF1 also promotes expression of genes encoding ABA-INSENSITIVE5 (ABI5) and DELLA growth-repressor proteins, which act in the ABA and GA signaling pathways, respectively. Here we show that MFT gene expression is promoted by FR light through the PIF1/SOM/ABI5/DELLA pathway and is repressed by R light via the transcription factor SPATULA (SPT). Consistent with this, we also show that SPT gene expression is repressed under FR light in a PIF1-dependent manner. Furthermore, transcriptomic analyses presented in this study indicate that MFT exerts its function by promoting expression of known ABA-induced genes and repressing cell wall expansion-related genes.

Mediation of Flower Induction by Gibberellin and its Inhibitor Paclobutrazol: mRNA and miRNA Integration Comprises Complex Regulatory Cross-Talk in Apple.

Guaranteeing successful flowering is very important in economic plant species, especially apple (Malus domestica Borkh.), which is difficult to induce to flower. However, the gene expression and networks involved in flowering have not been totally characterized. Here, we employed mRNA and microRNA (miRNA) sequencing to understand the different responses to gibberellin- and its inhibitor paclobutrazol- (PAC) mediated flower induction. Significant opposite cytological and morphological changes were observed in treated terminal buds, which led to a reduced flowering rate under gibberellin and an increased flowering rate under PAC. We also found that the differentially expressed mRNAs, miRNAs and miRNA target genes participated in different biological networks including hormones, photosynthesis, redox state and other metabolic processes, which provided important clues to understand the complex networks involved in apple flower induction. Additionally, we subsequently focused on one important candidate, MdSPL3, which is one of 31 apple SPL gene family members and whose transcription was inhibited by gibberellin but promoted by PAC. Functional investigation showed that MdSPL3 was located in the nucleus, and ectopic MdSPL3 activated floral meristem identity genes, promoted the formation of floral primordia and led to an earlier flowering phenotype in Arabidopsis. Our research identified critical mRNA and miRNA responsive to gibberellin or PAC, and provided a candidate framework for flower induction. This carefully orchestrated regulatory cross-talk highlighted potential targets for developing regulatory techniques and genetic improvement of flower induction in apple.

Plant Stature Related receptor-like Kinanse2 (PSRK2) acts as a factor that determines stem elongation toward gibberellins response in rice.

Gibberellins (GAs) are a family of plant hormones that are important to multiple aspects of plant growth and development, especially stem elongation. A PSRK2 was obtained through screening and identifying RLK dominant negative mutants. Phenotype of the loss-of-function mutants, psrk2-DN and psrk2-RNAi, showed that PSRK2 could influence the length of the uppermost and fourth internodes, indicating that PSRK2 might regulate cell division in the intercalary meristems and/or cell elongation in the internodes. Moreover, the expression pattern showed that PSRK2 was strongly expressed in the joined-nodes after the start-up of reproductive growth, but undetectable in leaves. PSRK2 expression was also found to be induced by GA3, and PSRK2 was involved in GA signaling in cereal aleurone cells, and PSRK2 influence the relative length of the second leaf sheaths in seedling stage. These results indicate PSRK2 is a component of GA signaling pathway that controls stem elongation by negatively regulating GA responses. Abbreviations: Os: Oryza sativa; At: Arabidopsis thaliana; RNAi: RNA interfere; DN: Dominate Negative; SMART: Simple Modular Architecture Research Tool; Uni : Uniconazol; PSRK2: Plant Stature Related receptor-like Kinase 2; RLK: Receptor-like Kinase; GA: Gibberellin; IAA: indole-3-acetic acid; BL: Brassinosteroid.

Ethylene-gibberellin signaling underlies adaptation of rice to periodic flooding.

Most plants do poorly when flooded. Certain rice varieties, known as deepwater rice, survive periodic flooding and consequent oxygen deficiency by activating internode growth of stems to keep above the water. Here, we identify the gibberellin biosynthesis gene, SD1 (SEMIDWARF1), whose loss-of-function allele catapulted the rice Green Revolution, as being responsible for submergence-induced internode elongation. When submerged, plants carrying the deepwater rice-specific SD1 haplotype amplify a signaling relay in which the SD1 gene is transcriptionally activated by an ethylene-responsive transcription factor, OsEIL1a. The SD1 protein directs increased synthesis of gibberellins, largely GA4, which promote internode elongation. Evolutionary analysis shows that the deepwater rice-specific haplotype was derived from standing variation in wild rice and selected for deepwater rice cultivation in Bangladesh.

Suppression of DELLA signaling induces procambial cell formation in culture.

The post-embryonic growth of plants requires the activities of apical meristems and lateral meristems. In the meristems, self-proliferation and differentiation of stem cells is tightly modulated by plant hormone signaling networks and specific transcription factors. Despite extensive studies on stem cell maintenance in plants, the mechanism by which stem cells are initially established is largely unknown. Vascular stem cells consisting of procambial/cambial cells give rise to xylem and phloem cells. In this study, we analyzed the establishment of procambial cells using the in vitro culture system VISUAL, in which mesophyll cells rapidly differentiate into xylem tracheary elements and phloem sieve elements via procambial cells. We found that procambial cell formation in VISUAL is initiated by light, which can be replaced by application of gibberellin (GA). Gibberellin was able to promote procambial cell formation through degradation of DELLA, whereas light did not elevate the endogenous GA content. Indeed, light in combination with bikinin reduced the accumulation of DELLA protein in VISUAL. Consistently, overexpression of a constitutively active DELLA protein repressed vascular cell differentiation even under light. These combined results suggest that DELLA signaling suppresses procambial cell formation during vascular development in VISUAL.

Transcriptomic analysis reveals the roles of gibberellin-regulated genes and transcription factors in regulating bolting in lettuce (Lactuca sativa L.).

A cool temperature is preferred for lettuce cultivation, as high temperatures cause premature bolting. Accordingly, exploring the mechanism of bolting and preventing premature bolting is important for agriculture. To explore this relationship in depth, morphological, physiological, and transcriptomic analyses of the bolting-sensitive line S39 at the five-leaf stage grown at 37°C were performed in the present study. Based on paraffin section results, we observed that S39 began bolting on the seventh day at 37°C. During bolting in the heat-treated plants, GA3 and GA4 levels in leaves and the indoleacetic acid (IAA) level in the stem reached a maximum on the sixth day, and these high contents were maintained. Additionally, bolting begins in the fifth day after GA3 treatment in S39 plants, GA3 and GA4 increased and then decreased, reaching a maximum on the fourth day in leaves. Similarly, IAA contents reached a maximum in the stem on the fifth day. No bolting was observed in the control group grown at 25°C, and significant changes were not observed in GA3 and GA4 levels in the controls during the observation period. RNA-sequencing data implicated transcription factors (TFs) in regulating bolting in lettuce, suggesting that the high GA contents in the leaves and IAA in the stem promote bolting. TFs possibly modulate the expression of related genes, such as those encoding hormones, potentially regulating bolting in lettuce. Compared to the control group, 258 TFs were identified in the stem of the treatment group, among which 98 and 156 were differentially up- and down-regulated, respectively; in leaves, 202 and 115 TFs were differentially up- and down-regulated, respectively. Significant changes in the treated group were observed for C2H2 zinc finger, AP2-EREBP, and WRKY families, indicating that these TFs may play important roles in regulating bolting.

Interplay between cytochrome c and gibberellins during Arabidopsis vegetative development.

We studied the effect of reducing the levels of the mitochondrial electron carrier cytochrome c (CYTc) in Arabidopsis thaliana. Plants with CYTc deficiency have delayed growth and development, and reach flowering several days later than the wild-type but with the same number of leaves. CYTc-deficient plants accumulate starch and glucose during the day, and contain lower levels of active gibberellins (GA) and higher levels of DELLA proteins, involved in GA signaling. GA treatment abolishes the developmental delay and reduces glucose accumulation in CYTc-deficient plants, which also show a lower raise in ATP levels in response to glucose. Treatment of wild-type plants with inhibitors of mitochondrial energy production limits plant growth and increases the levels of DELLA proteins, thus mimicking the effects of CYTc deficiency. In addition, an increase in the amount of CYTc decreases DELLA protein levels and expedites growth, and this depends on active GA synthesis. We conclude that CYTc levels impinge on the activity of the GA pathway, most likely through changes in mitochondrial energy production. In this way, hormone-dependent growth would be coupled to the activity of components of the mitochondrial respiratory chain.

Transcriptome analysis of Phelipanche aegyptiaca seed germination mechanisms stimulated by fluridone, TIS108, and GR24.

P. aegyptiaca is one of the most destructive root parasitic plants worldwide, causing serious damage to many crop species. Under natural conditions P. aegyptiaca seeds must be conditioned and then stimulated by host root exudates before germinating. However, preliminary experiments indicated that TIS108 (a triazole-type inhibitor of strigolactone) and fluridone (FL, an inhibitor of carotenoid-biosynthesis) both stimulated the germination of P. aegyptiaca seeds without a water preconditioning step (i.e. unconditioned seeds). The objective of this study was to use deep RNA sequencing to learn more about the mechanisms by which TIS108 and FL stimulate the germination of unconditioned P. aegyptiaca seeds. Deep RNA sequencing was performed to compare the mechanisms of germination in the following treatments: (i) unconditioned P. aegyptiaca seeds with no other treatment, (ii) unconditioned seeds treated with 100 mg/L TIS108, (iii) unconditioned seeds treated with 100 mg/L FL + 100 mg/L GA3, (iv) conditioned seeds treated with sterile water, and (v) conditioned seeds treated with 0.03 mg/L GR24. The de novo assembled transcriptome was used to analyze transcriptional dynamics during seed germination. The key gene categories involved in germination were also identified. The results showed that only 119 differentially expressed genes were identified in the conditioned treatment vs TIS108 treatment. This indicated that the vast majority of conditions for germination were met during the conditioning stage. Abscisic acid (ABA) and gibberellic acid (GA) played important roles during P. aegyptiaca germination. The common pathway of TIS108, FL+GA3, and GR24 in stimulating P. aegyptiaca germination was the simultaneous reduction in ABA concentrations and increase GA concentrations. These results could potentially aid the identification of more compounds that are capable of stimulating P. aegyptiaca germination. Some potential target sites of TIS108 were also identified in our transcriptome data. The results of this experiment suggest that TIS108 and FL+GA3 could be used to control P. aegyptiaca through suicidal germination.

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.

A modern Green Revolution gene for reduced height in wheat.

Increases in the yield of wheat during the Green Revolution of the late 20th century were achieved through the introduction of Reduced height (Rht) dwarfing genes. The Rht-B1 and Rht-D1 loci ensured short stature by limiting the response to the growth-promoting hormone gibberellin, and are now widespread through international breeding programs. Despite this advantage, interference with the plant's response to gibberellin also triggers adverse effects for a range of important agronomic traits, and consequently modern Green Revolution genes are urgently required. In this study, we revisited the genetic control of wheat height using an association mapping approach and a large panel of 1110 worldwide winter wheat cultivars. This led to the identification of a major Rht locus on chromosome 6A, Rht24, which substantially reduces plant height alone as well as in combination with Rht-1b alleles. Remarkably, behind Rht-D1, Rht24 was the second most important locus for reduced height, explaining 15.0% of the genotypic variance and exerting an allele substitution effect of -8.8 cm. Unlike the two Rht-1b alleles, plants carrying Rht24 remain sensitive to gibberellic acid treatment. Rht24 appears in breeding programs from all countries of origin investigated, with increased frequency over the last decades, indicating that wheat breeders have actively selected for this locus. Taken together, this study reveals Rht24 as an important Rht gene of commercial relevance in worldwide wheat breeding.

GA as a regulatory link between the showy floral traits color and scent.

Emission of volatiles at advanced stages of flower development is a strategy used by plants to lure pollinators to the flower. We reveal that GA negatively regulates floral scent production in petunia. We used Agrobacterium-mediated transient expression of GA-20ox in petunia flowers and a virus-induced gene silencing approach to knock down DELLA expression, measured volatile emission, internal pool sizes and GA levels by GC-MS or LC-MS/MS, and analyzed transcript levels of scent-related phenylpropanoid-pathway genes. We show that GA has a negative effect on the concentrations of accumulated and emitted phenylpropanoid volatiles in petunia flowers; this effect is exerted through transcriptional/post-transcriptional downregulation of regulatory and biosynthetic scent-related genes. Both overexpression of GA20-ox, a GA-biosynthesis gene, and suppression of DELLA, a repressor of GA-signal transduction, corroborated GA's negative regulation of floral scent. We present a model in which GA-dependent timing of the sequential activation of different branches of the phenylpropanoid pathway during flower development may represent a link between the showy traits controlling pollinator attraction, namely color and scent.

Hormonal control of the floral transition: Can one catch them all?

The transition to flowering marks a key adaptive developmental switch in plants which impacts on their survival and fitness. Different signaling pathways control the floral transition, conveying both endogenous and environmental cues. These cues are often relayed and/or modulated by different hormones, which might confer additional developmental flexibility to the floral process in the face of varying conditions. Among the different hormonal pathways, the phytohormone gibberellic acid (GA) plays a dominant role. GA is connected with the other floral pathways through the GA-regulated DELLA proteins, acting as versatile interacting modules for different signaling proteins. In this review, I will highlight the role of DELLAs as spatial and temporal modulators of different consolidated floral pathways. Next, building on recent data, I will provide an update on some emerging themes connecting other hormone signaling cascades to flowering time control. I will finally provide examples for some established as well as potential cross-regulatory mechanisms between hormonal pathways mediated by the DELLA proteins.

Axillary buds are dwarfed shoots that tightly regulate GA pathway and GA-inducible 1,3-ß-glucanase genes during branching in hybrid aspen.

Axillary buds (AXBs) of hybrid aspen (Populus tremula×P. tremuloides) contain a developing dwarfed shoot that becomes para-dormant at the bud maturation point. Para-dormant AXBs can grow out after stem decapitation, while dormant AXBs pre-require long-term chilling to release them from dormancy. The latter is mediated by gibberellin (GA)-regulated 1,3-ß-glucanases, but it is unknown if GA is also important in the development, activation, and outgrowth of para-dormant AXBs. The present data show that para-dormant AXBs up-regulate GA receptor genes during their maturation, but curtail GA biosynthesis by down-regulating the rate-limiting GIBBERELLIN 3-OXIDASE2 (GA3ox2), which is characteristically expressed in the growing apex. However, decapitation significantly up-regulated GA3ox2 and GA4-responsive 1,3-ß-glucanases (GH17-family; α-clade). In contrast, decapitation down-regulated γ-clade 1,3-ß-glucanases, which were strongly up-regulated in maturing AXBs concomitant with lipid body accumulation. Overexpression of selected GH17 members in hybrid aspen resulted in characteristic branching patterns. The α-clade member induced an acropetal branching pattern, whereas the γ-clade member activated AXBs in recurrent flushes during transient cessation of apex proliferation. The results support a model in which curtailing the final step in GA biosynthesis dwarfs the embryonic shoot, while high levels of GA precursors and GA receptors keep AXBs poised for growth. GA signaling, induced by decapitation, reinvigorates symplasmic supply routes through GA-inducible 1,3-ß-glucanases that hydrolyze callose at sieve plates and plasmodesmata.