AGAMOUS-LIKE 24 Research Articles

The AGL6–ELF3–FT circuit controls flowering time in Arabidopsis

ABSTRACT Adjusting the timing of floral transition is essential for reproductive success in plants. A number of flowering regulators integrate internal and external signals to precisely determine the time to flower. We here report that the AGAMOUS-LIKE 6 (AGL6) – EARLY FLOWERING 3 (ELF3) module regulates flowering in the FLOWERING LOCUS T (FT)-dependent pathway in Arabidopsis. The AGL6 transcriptional repressor promotes floral transition by directly suppressing ELF3, which in turn directly represses FT expression that acts as a floral integrator. Indeed, ELF3 is epistatic to AGL6 in the control of floral transition. Overall, our findings propose that the AGL6–ELF3 module contributes to fine-tuning flowering time in plants.

AINTEGUMENTA and redundant AINTEGUMENTA-LIKE6 are required for bract outgrowth in Arabidopsis.

Plants consist of fundamental units of growth called phytomers (leaf or bract, axillary bud, node, and internode), which are repeated and modified throughout shoot development to give plants plasticity for survival and adaptation. One phytomer modification is the suppression or outgrowth of bracts, the leaves subtending the flowers. The floral meristem identity regulator LEAFY (LFY) and the organ boundary genes BLADE-ON-PETIOLE1 (BOP1) and BOP2 have been shown to suppress bract development in Arabidopsis, as mutations in these genes result in bract outgrowth. However, much less is known about the mechanisms that promote bract outgrowth in Arabidopsis mutants such as these. Further understanding of this mechanism may provide a potential tool for modifying leaf development. Here, we showed that the MADS-box genes SUPPRESSOR OF OVEREXPRESSION OF CONSTANS1 (SOC1), FRUITFUL (FUL), and AGAMOUS-LIKE24 (AGL24) play more important roles than BOP1/2 and LFY in bract suppression, and that AINTEGUMENTA (ANT) and the partially redundant AINTEGUMENTA-LIKE6 (AIL6) are necessary for bract outgrowth in these mutant backgrounds. We also demonstrated that misexpression of AIL6 alone is sufficient for bract outgrowth. Our data reveal a mechanism for bract suppression and outgrowth and provide insight into phytomer plasticity.

Multifaceted roles of transcription factors during plant embryogenesis.

Transcription factors (TFs) are diverse groups of regulatory proteins. Through their specific binding domains, TFs bind to their target genes and regulate their expression, therefore TFs play important roles in various growth and developmental processes. Plant embryogenesis is a highly regulated and intricate process during which embryos arise from various sources and undergo development; it can be further divided into zygotic embryogenesis (ZE) and somatic embryogenesis (SE). TFs play a crucial role in the process of plant embryogenesis with a number of them acting as master regulators in both ZE and SE. In this review, we focus on the master TFs involved in embryogenesis such as BABY BOOM (BBM) from the APETALA2/Ethylene-Responsive Factor (AP2/ERF) family, WUSCHEL and WUSCHEL-related homeobox (WOX) from the homeobox family, LEAFY COTYLEDON 2 (LEC2) from the B3 family, AGAMOUS-Like 15 (AGL15) from the MADS family and LEAFY COTYLEDON 1 (LEC1) from the Nuclear Factor Y (NF-Y) family. We aim to present the recent progress pertaining to the diverse roles these master TFs play in both ZE and SE in Arabidopsis, as well as other plant species including crops. We also discuss future perspectives in this context.

Regulatory mechanisms of the LBD40 transcription factor in Arabidopsis thaliana somatic embryogenesis.

Somatic embryogenesis (SE) is a process by which an embryo is derived from somatic tissue. Transcription factors (TFs) have been identified that control this process. One such TF that promotes SE is AGAMOUS-like 15 (AGL15). Prior work has shown that AGL15 can both induce and repress gene expression. One way this type of dual function TF works is via protein interactions, so a yeast 2-hybrid (Y2H) screen was undertaken. One intriguing protein with which AGL15 interacted in Y2H was LBD40. LBD40 encodes a LATERAL ORGAN BOUNDARIES (LOB)-domain TF that is unique to plants and is primarily expressed during seed development. Here, we confirm the AGL15-LBD40 interaction by quantitative assays and in planta co-immunoprecipation. We also document a role for LBD40, and the closely related protein LBD41, in supporting SE. To determine downstream genes potentially controlled by LBD40, chromatin immunoprecipitation followed by high throughput sequencing (ChIP-seq) was used. More than 400 binding regions for LBD40 were consistently found genome-wide. To determine genes responsive to LBD40/41 accumulation, RNA-seq analysis of transcriptomes of wild-type control and loss-of-function lbd40/lbd41 was performed. Combining these datasets provides insight into genes directly and indirectly controlled by these LOB domain TFs. The gene ontology (GO) enrichment analysis of these regulated genes showed an overrepresentation of biological processes that are associated with SE, further indicating the importance of LBD40 in SE. This work provides insight into SE, a poorly understood, but essential process to generate transgenic plants to meet agricultural demands or test gene function. This manuscript reports on experiments to understand the role that LDB40, a TF, plays in support of SE by investigating genes directly and indirectly controlled by LBD40 and examining physical and genetic interactions with other TFs active in SE. We uncover targets of LBD40 and an interacting TF of the MADS family and investigate targets involvement in SE.

MicroRNAs associated with AGL6 and IAA9 function in tomato fruit set

ObjectiveFruit set is triggered after ovule fertilization, as a consequence of the downregulation of ovary growth repressors, such as the tomato transcription factors Auxin/indole-3-acetic acid 9 (IAA9) and Agamous-like 6 (AGL6). In a recent work, we developed a method to silence IAA9 and AGL6 in tomato ovaries using exogenous dsRNAs. We also produced small RNA libraries from IAA9- and AGL6-silenced ovaries to confirm the presence of siRNAs, derived from exogenous dsRNA, targeting IAA9 and AGL6. The objective of this work is to exploit these sRNA libraries to identify miRNAs differentially expressed in IAA9- and AGL6-silenced ovaries as compared with unpollinated control ovaries.ResultsWe identified by RNA sequencing 125 and 104 known and 509 and 516 novel miRNAs from reads mapped to mature or hairpin sequences, respectively. Of the known miRNAs, 7 and 45 were differentially expressed in IAA9- and AGL6-silenced ovaries compared to control ones, respectively. Six miRNAs were common to both datasets, suggesting their importance in the fruit set process. The expression pattern of two of these (miR393 and miR482e-5p) was verified by stem-loop qRT-PCR. The identified miRNAs represent a pool of regulatory sRNAs potentially involved in tomato fruit initiation.

Light induction of somatic embryogenesis in Arabidopsis is regulated by PHYTOCHROME E

The requirement of light on somatic embryogenesis (SE) has been documented in many species; however, no mechanism of action has been elucidated. Using Arabidopsis SE as a model, the effect of red light (660 nm) during the induction phase corresponding to the formation of the embryogenic tissue was examined. Analyses of several phytochrome mutants revealed that red light signaling, conducive to SE, was mediated by PHYTOCHROME E (PHYE). Both phyE and darkness were sufficient to repress the formation of somatic embryos and reduced the expression of CONSTITUTIVE PHOTOMORPHIC DWARF 3 (CPD3), a rate limiting step in brassinosteroid (BR) biosynthesis, as well as AGAMOUS LIKE 15 (AGL15), a key inducer of many SE genes. We further integrated BR signaling and nitric oxide (NO) with PHYE by demonstrating that applications of both compounds to phyE explants and WT explants cultured in the dark partially restored AGL15 expression. These results demonstrate that SE induction by red light operates via PHYE through BR signaling and NO required to induce AGL15.

Functional Divergence Analysis of AGL6 Genes in Prunus mume.

The AGAMOUS-LIKE6 (AGL6) lineage is an important clade of MADS-box transcription factors that play essential roles in floral organ development. The genome of Prunus mume contains two homoeologous AGL6 genes that are replicated as gene fragments. In this study, two AGL6 homologs, PmAGL6-1 and PmAGL6-2, were cloned from P. mume and then functionally characterized. Sequence alignment and phylogenetic analyses grouped both genes into the AGL6 lineage. The expression patterns and protein-protein interaction patterns showed significant differences between the two genes. However, the ectopic expression of the two genes in Arabidopsis thaliana resulted in similar phenotypes, including the promotion of flowering, alteration of floral organ structure, participation in the formation of the floral meristem and promotion of pod bending. Therefore, gene duplication has led to some functional divergence of PmAGL6-1 and PmAGL6-2 but their functions are similar. We thus speculated that AGL6 genes play a crucial role in flower development in P. mume.

Delphinieae flowers originated from the rewiring of interactions between duplicated and diversified floral organ identity and symmetry genes.

Species of the tribe Delphinieae (Ranunculaceae) have long been the focus of morphological, ecological, and evolutionary studies due to their highly specialized, nearly zygomorphic (bilaterally symmetrical) spiral flowers with nested petal and sepal spurs and reduced petals. The mechanisms underlying the development and evolution of Delphinieae flowers, however, remain unclear. Here, by conducting extensive phylogenetic, comparative transcriptomic, expression, and functional studies, we clarified the evolutionary histories, expression patterns, and functions of floral organ identity and symmetry genes in Delphinieae. We found that duplication and/or diversification of APETALA3-3 (AP3-3), AGAMOUS-LIKE6 (AGL6), CYCLOIDEA (CYC), and DIVARICATA (DIV) lineage genes was tightly associated with the origination of Delphinieae flowers. Specifically, an AGL6-lineage member (such as the Delphinium ajacis AGL6-1a) represses sepal spur formation and petal development in the lateral and ventral parts of the flower while determining petal identity redundantly with AGL6-1b. By contrast, two CYC2-like genes, CYC2b and CYC2a, define the dorsal and lateral-ventral identities of the flower, respectively, and form complex regulatory links with AP3-3, AGL6-1a, and DIV1. Therefore, duplication and diversification of floral symmetry genes, as well as co-option of the duplicated copies into the preexisting floral regulatory network, have been key for the origin of Delphinieae flowers.

Revisiting AGAMOUS-LIKE15, a Key Somatic Embryogenesis Regulator, Using Next Generation Sequencing Analysis in Arabidopsis

AGAMOUS-like 15 (AGL15) is a member of the MADS-domain transcription factor (TF) family. MADS proteins are named for a conserved domain that was originally from an acronym derived from genes expressed in a variety of eukaryotes (MCM1-AGAMOUS-DEFICIENS-SERUM RESPONSE FACTOR). In plants, this family has expanded greatly, with more than one-hundred members generally found in dicots, and the proteins encoded by these genes have often been associated with developmental identity. AGL15 transcript and protein accumulate primarily in embryos and has been found to promote an important process called plant regeneration via somatic embryogenesis (SE). To understand how this TF performs this function, we have previously used microarray technologies to assess direct and indirect responsive targets of this TF. We have now revisited this question using next generation sequencing (NGS) to both characterize in vivo binding sites for AGL15 as well as response to the accumulation of AGL15. We compared these data to the prior microarray results to evaluate the different platforms. The new NGS data brought to light an interaction with brassinosteroid (BR) hormone signaling that was "missed" in prior Gene Ontology analysis from the microarray studies.

Cloning, expression and functional analysis of the SOC1 homologous gene in pak choi (Brassica rapa ssp. Chinensis makino)

Flowering transition is important in plant growth and development, and the SUPPRESSOR OF OVEREXPRESSION OF CO 1 (SOC1) gene belongs to the MADS-box gene family, which is an integrator involved in the regulation of flower formation in plants. Current studies have shown that the structural domain of SOC1 is relatively conserved in a variety of plants. To further investigate the function of the SOC1 homologous gene in pak choi, the full-length coding sequence of pak choi BrcSOC1 was cloned, and the structure, subcellular localization and spatiotemporal expression pattern of BrcSOC1 were analyzed in this study. The tissue-specific expression results showed that BrcSOC1 was expressed in roots, leaves, bolts, flowers and pods. The expression results at different developmental stages showed that BrcSOC1 was expressed at the highest level at floral bud differentiation stage 5. When BrcSOC1 was overexpressed in Arabidopsis thaliana, the flowering time in all transgenic plants was significantly earlier than in the wild type, and the expression analysis indicated that BrcSOC1 positively regulated the expression of the downstream genes AGAMOUS-LIKE 24 (AGL24) and LEAFY (LFY). These results provide an important reference for further research on the mechanism of BrcSOC1 in the flowering pathway of pak choi.

Identification of key regulatory genes involved in the sporophyte and gametophyte development in Ginkgo biloba ovules revealed by in situ expression analyses

PremiseIn Arabidopsis thaliana, the role of the most important key genes that regulate ovule development is widely known. In nonmodel species, and especially in gymnosperms, the ovule developmental processes are still quite obscure. In this study, we describe the putative roles of Ginkgo biloba orthologs of regulatory genes during ovule development. Specifically, we studied AGAMOUS (AG), AGAMOUS‐like 6 (AGL6), AINTEGUMENTA (ANT), BELL1 (BEL1), Class III HD‐Zip, and YABBY Ginkgo genes.MethodsWe analyzed their expression domains through in situ hybridizations on two stages of ovule development: the very early stage that corresponds to the ovule primordium, still within wintering buds, and the late stage at pollination time.Results GBM5 (Ginkgo ortholog of AG), GbMADS8 (ortholog of AGL6) and GbC3HDZ1‐2‐3 were expressed in both the stages of ovule development, while GbMADS1, GbAGL6‐like genes (orthologs of AGL6), GbBEL1‐2 and YABBY Ginkgo orthologs (GbiYAB1B and GbiYABC) seem mostly involved at pollination time. GbANTL1 was not expressed in the studied stages and was different from GbANTL2 and GbBEL1, which seem to be involved at both stages of ovule development. In Ginkgo, the investigated genes display patterns of expression only partially comparable to those of other studied seed plants.ConclusionsThe expression of most of these regulatory genes in the female gametophyte region at pollination time leads to suggest a communication between the sporophytic maternal tissue and the developing female gametophyte, as demonstrated for well‐studied model angiosperms.

Circlize package in R and Analytic Hierarchy Process (AHP): Contribution values of ABCDE and AGL6 genes in the context of floral organ development.

The morphological diversity of floral organs can largely be attributed to functional divergence in the MADS-box gene family. Nonetheless, research based on the ABCDE model has yet to conclusively determine whether the AGAMOUS-LIKE 6 (AGL6) subgroup has a direct influence on floral organ development. In the current study, the ABCDE model was used to quantify the contributions of ABCDE and AGL6 genes in the emergence of floral organs. We determined that the flower formation contribution values of the ABCDE and AGL6 genes were as follows: A gene, 0.192; B gene, 0.231; CD gene, 0.192; E gene, 0.385; and AGL6, 0.077. As AGL6 does not directly influence floral structure formation, the contribution value of AGL6 to flower formation was low. Furthermore, the gradient values of the floral organs were as follows: sepals, 0.572; petals, 1.606; stamens, 2.409; and carpels, 2.288. We also performed detailed analysis of the ABCDE and AGL6 genes using the Circlize package in R. Our results suggest that these genes likely emerged in one of two orders: 1) B genes→CD genes→AGL6→E genes→A genes; or 2) B genes→CD genes→AGL6/E genes→A genes. We use the analytic hierarchy process (AHP) to prove the contribution values and gradient values of floral organs. This is the first study to understand the contribution values of ABCDE and AGL6 genes using the AHP and the Circlize package in R.

The wheat AGL6-like MADS-box gene is a master regulator for floral organ identity and a target for spikelet meristem development manipulation.

SummaryThe AGAMOUS‐LIKE6 (AGL6)‐like genes are ancient MADS‐box genes and are functionally studied in a few model plants. The knowledge of these genes in wheat remains limited. Here, by studying a ‘double homoeolog mutant’ of the AGL6 gene in tetraploid wheat, we showed that AGL6 was required for the development of all four whorls of floral organs with dosage‐dependent effect on floret fertility. Yeast two‐hybrid analyses detected interactions of AGL6 with all classes of MADS‐box proteins in the ABCDE model for floral organ development. AGL6 was found to interact with several additional proteins, including the G protein β and γ (DEP1) subunits. Analysis of the DEP1‐B mutant showed a significant reduction in spikelet number per spike in tetraploid wheat, while overexpression of AGL6 in common wheat increased the spikelet number per spike and hence the grain number per spike. RNA‐seq analysis identified the regulation of several meristem activity genes by AGL6, such as FUL2 and TaMADS55. Our work therefore extensively updated the wheat ABCDE model and proposed an alternative approach to improve wheat grain yield by manipulating the AGL6 gene.

Genome-wide identification of jujube witches’ broom phytoplasma effectors revealed the role of SJP3 in inducing phyllody

Jujube witches’ broom (JWB), associated with ‘Candidatus Phytoplasma ziziphi’, is a devastating disease in Chinese jujube (Ziziphus jujuba Mill.). One of the symptoms in plants infected with phytoplasmas is the change of floral organs into leaf-like structures, known as phyllody. Effector proteins secreted by phytoplasmas are key to inducing this and other witches’ broom symptoms. However, the phytoplasma effector proteins that induce phyllody in jujube remain unknown. We identified 43 candidate secreted JWB phytoplasma proteins (SJPs) in the JWB phytoplasma genome. Eight of the predicted candidate effector genes exhibited high transcript levels in infected flower buds. Among the eight candidate effectors, SJP3 experimentally showed the potential to induce phyllody. SJP3 expression in infected jujube was at a high level in young leaves and flowers. Ectopic expression of SJP3 in Arabidopsis induced leaf-like flowers and extra secondary flowers and significantly changed the transcript levels of PISTILLATA (PI), SHORT VEGETATIVE PHASE (SVP), AGAMOUS-LIKE 24 (AGL24) and SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1 (SOC1). The expression levels of these MADS-box homologous genes were also changed in infected jujube flower buds and all four whorls of the floral organs. The data showed that the effector SJP3 disrupts expression of several MADS-box genes associated with floral organ identity and flowering time and is likely key to inducing phyllody in phytoplasma-infected jujube. This study provides new insight into understanding the pathogenesis of JWB disease induced by phytoplasma.

Comparative Transcriptomics of Non-Embryogenic and Embryogenic Callus in Semi-Recalcitrant and Non-Recalcitrant Upland Cotton Lines.

Somatic embryogenesis-mediated plant regeneration is essential for the genetic manipulation of agronomically important traits in upland cotton. Genotype specific recalcitrance to regeneration is a primary challenge in deploying genome editing and incorporating useful transgenes into elite cotton germplasm. In this study, transcriptomes of a semi-recalcitrant cotton (Gossypium hirsutum L.) genotype ‘Coker312’ were analyzed at two critical stages of somatic embryogenesis that include non-embryogenic callus (NEC) and embryogenic callus (EC) cells, and the results were compared to a non-recalcitrant genotype ‘Jin668’. We discovered 305 differentially expressed genes in Coker312, whereas, in Jin668, about 6-fold more genes (2155) were differentially expressed. A total of 154 differentially expressed genes were common between the two genotypes. Gene enrichment analysis of the upregulated genes identified functional categories, such as lipid transport, embryo development, regulation of transcription, sugar transport, and vitamin biosynthesis, among others. In Coker312 EC cells, five major transcription factors were highly upregulated: LEAFY COTYLEDON 1 (LEC1), WUS-related homeobox 5 (WOX5), ABSCISIC ACID INSENSITIVE3 (ABI3), FUSCA3 (FUS3), and WRKY2. In Jin668, LEC1, BABY BOOM (BBM), FUS3, and AGAMOUS-LIKE15 (AGL15) were highly expressed in EC cells. We also found that gene expression of these embryogenesis genes was typically higher in Jin668 when compared to Coker312. We conclude that significant differences in the expression of the above genes between Coker312 and Jin668 may be a critical factor affecting the regenerative ability of these genotypes.

The EAR Motif in the Arabidopsis MADS Transcription Factor AGAMOUS-Like 15 Is Not Necessary to Promote Somatic Embryogenesis.

AGAMOUS-like 15 (AGL15) is a member of the MADS domain family of transcription factors (TFs) that can directly induce and repress target gene expression, and for which promotion of somatic embryogenesis (SE) is positively correlated with accumulation. An ethylene-responsive element binding factor-associated amphiphilic repression (EAR) motif of form LxLxL within the carboxyl-terminal domain of AGL15 was shown to be involved in repression of gene expression. Here, we examine whether AGL15′s ability to repress gene expression is needed to promote SE. While a form of AGL15 where the LxLxL is changed to AxAxA can still promote SE, another form with a strong transcriptional activator at the carboxy-terminal end, does not promote SE and, in fact, is detrimental to SE development. Select target genes were examined for response to the different forms of AGL15.

Brassinosteroids repress the seed maturation program during the seed-to-seedling transition.

In flowering plants, repression of the seed maturation program is essential for the transition from the seed to the vegetative phase, but the underlying mechanisms remain poorly understood. The B3-domain protein VIVIPAROUS1/ABSCISIC ACID-INSENSITIVE3-LIKE 1 (VAL1) is involved in repressing the seed maturation program. Here we uncovered a molecular network triggered by the plant hormone brassinosteroid (BR) that inhibits the seed maturation program during the seed-to-seedling transition in Arabidopsis (Arabidopsis thaliana). val1-2 mutant seedlings treated with a BR biosynthesis inhibitor form embryonic structures, whereas BR signaling gain-of-function mutations rescue the embryonic structure trait. Furthermore, the BR-activated transcription factors BRI1-EMS-SUPPRESSOR 1 and BRASSINAZOLE-RESISTANT 1 bind directly to the promoter of AGAMOUS-LIKE15 (AGL15), which encodes a transcription factor involved in activating the seed maturation program, and suppress its expression. Genetic analysis indicated that BR signaling is epistatic to AGL15 and represses the seed maturation program by downregulating AGL15. Finally, we showed that the BR-mediated pathway functions synergistically with the VAL1/2-mediated pathway to ensure the full repression of the seed maturation program. Together, our work uncovered a mechanism underlying the suppression of the seed maturation program, shedding light on how BR promotes seedling growth.

VvDAM-SVPs genes are regulated by FLOWERING LOCUS T (VvFT) and not by ABA/low temperature-induced VvCBFs transcription factors in grapevine buds.

In deciduous fruit trees in which dormancy is induced by low temperatures, the expression of DORMACY-ASSOCIATED MADS-BOX genes (DAM) is regulated by CBF/DREB1 transcription factors. In Vitis vinifera, in which dormancy is induced by the photoperiod, VvDAM-SVPs gene expression is regulated by FLOWERING LOCUS T (VvFT). Using the sequences of the six peach (Prunus persica) DORMACY-ASSOCIATED MADS-box genes (DAM) as query, eight putative DAM genes belonging to the family of MADS-box transcription factors and related to the Arabidopsis floral regulators SHORT VEGETATIVE PHASE (SVP) and AGAMOUS LIKE 24 (AGL24) were identified in the V. vinifera genome. Among these, five belong to the subfamily SVP-like genes which have been associated with the regulation of flowering and dormancy in annual and perennial plants, respectively. It has been proposed that they play a direct role in the induction and maintenance of endodormancy (ED) through the regulation of the FLOWERING LOCUS T (FT) gene. In the present study, it is demonstrated that in V. vinifera: (1) VvDAM-SVPs genes are not regulated by ABA/low temperature-induced VvCBFs transcription factors as described for other species of deciduous fruit trees. (2) A contrasting expression pattern between VvDAM3-SVP and VvFT was found under different experimental conditions related to the entry and exit of grapevine buds from ED. (3) Overexpression of VvFT in somatic grapevine embryos (SGE) repressed the expression of VvDAM3-SVP and VvDAM4-SVP. Taken together, the results suggest that VvDAM3-SVP could be associated with ED in grapevine buds, and that its expression could be regulated by VvFT.

AGL19 directly interacts with floral signal integrator AGL24 in flowering time control of Brassica juncea

SUPPRESSOR OF OVEREXPRESSION OF CONSTANS1 (SOC1) and AGAMOUS-LIKE 24 (AGL24) are pivotal flowering signal integrators, which respectively, belong to TM3/SOC1 and STMADS11 clades of MIKC-type MADS-box family. AGAMOUS-LIKE 19 (AGL19), one member of TM3/SOC1 clade family, acts as an activator of flowering time. Previous studies have suggested that AGL19 together with AGL24 and SOC1 coordinately regulate flowering time. However, it remains unknown how AGL19 interacts with SOC1 and AGL24 in Brassica juncea. Here, an AGL19-like (BjuAGL19) was cloned from B. juncea, which was expressed in roots, stems, leaves, buds and flowers. Furthermore, both yeast one-hybrid and Dual-Glo Luciferase assays confirmed that BjuAGL19 protein could not target the promoters of BjuSOC1 and BjuAGL24. Interestingly, based on yeast two-hybrid and pull-down assays, BjuAGL19 protein directly interacted with BjuAGL24 protein, instead of BjuSOC1 protein. It is speculated that BjuAGL19 can bind to BjuAGL24 to form protein complex of BjuAGL19-BjuAGL24 and thus regulates flowering time in B. juncea. The results are very helpful for further elucidating the molecular mechanism of BjuAGL19 in flowering time control.

AGL15 Controls the Embryogenic Reprogramming of Somatic Cells in Arabidopsis through the Histone Acetylation-Mediated Repression of the miRNA Biogenesis Genes

The embryogenic transition of somatic cells requires an extensive reprogramming of the cell transcriptome. Relevantly, the extensive modulation of the genes that have a regulatory function, in particular the genes encoding the transcription factors (TFs) and miRNAs, have been indicated as controlling somatic embryogenesis (SE) that is induced in vitro in the somatic cells of plants. Identifying the regulatory relationships between the TFs and miRNAs during SE induction is of central importance for understanding the complex regulatory interplay that fine-tunes a cell transcriptome during the embryogenic transition. Hence, here, we analysed the regulatory relationships between AGL15 (AGAMOUS-LIKE 15) TF and miR156 in an embryogenic culture of Arabidopsis. Both AGL15 and miR156 control SE induction and AGL15 has been reported to target the MIR156 genes in planta. The results showed that AGL15 contributes to the regulation of miR156 in an embryogenic culture at two levels that involve the activation of the MIR156 transcription and the containment of the abundance of mature miR156 by repressing the miRNA biogenesis genes DCL1 (DICER-LIKE1), SERRATE and HEN1 (HUA-ENHANCER1). To repress the miRNA biogenesis genes AGL15 seems to co-operate with the TOPLESS co-repressors (TPL and TPR1-4), which are components of the SIN3/HDAC silencing complex. The impact of TSA (trichostatin A), an inhibitor of the HDAC histone deacetylases, on the expression of the miRNA biogenesis genes together with the ChIP results implies that histone deacetylation is involved in the AGL15-mediated repression of miRNA processing. The results indicate that HDAC6 and HDAC19 histone deacetylases might co-operate with AGL15 in silencing the complex that controls the abundance of miR156 during embryogenic induction. This study provides new evidence about the histone acetylation-mediated control of the miRNA pathways during the embryogenic reprogramming of plant somatic cells and the essential role of AGL15 in this regulatory mechanism.