Characterization and validation of TaAGL66, a gene related to fertility conversion of wheat in the presence of Aegilops kotschyi cytoplasm.

MAIN CONCLUSION: TaAGL66, a MADS-box transcription factor highly expressed in fertile anthers of KTM3315A, regulates anther and/or pollen development, as well as male fertility in wheat with Aegilops kotschyi cytoplasm. Male sterility, as a string of sophisticated biological processes in higher plants, is commonly regulated by transcription factors (TFs). Among them, MADS-box TFs are mainly participated in the processes of floral organ formation and pollen development, which are tightly related to male sterility, but they have been little studied in the reproductive development in wheat. In our study, TaAGL66, a gene that was specifically expressed in spikes and highly expressed in fertile anthers, was identified by RNA sequencing and the expression profiles data of these genes, and qRT-PCR analyses, which was localized to the nucleus. Silencing of TaAGL66 under fertility condition in KTM3315A, a thermo-sensitive male sterile line with Ae. kotschyi cytoplasm, displayed severe fertility reduction, abnormal anther dehiscence, defective pollen development, decreased viability, and low seed-setting. It can be concluded that TaAGL66 plays an important role in wheat pollen development in the presence of Ae. kotschyi cytoplasm, providing new insights into the utilization of male sterility.

HDACs MADS-domain protein interaction: a case study of HDA15 and XAL1 in Arabidopsis thaliana.

Cellular behavior, cell differentiation and ontogenetic development in eukaryotes result from complex interactions between epigenetic and classic molecular genetic mechanisms, with many of these interactions still to be elucidated. Histone deacetylase enzymes (HDACs) promote the interaction of histones with DNA by compacting the nucleosome, thus causing transcriptional repression. MADS-domain transcription factors are highly conserved in eukaryotes and participate in controlling diverse developmental processes in animals and plants, as well as regulating stress responses in plants. In this work, we focused on finding out putative interactions of Arabidopsis thaliana HDACs and MADS-domain proteins using an evolutionary perspective combined with bioinformatics analyses and testing the more promising predicted interactions through classic molecular biology tools. Through bioinformatic analyses, we found similarities between HDACs proteins from different organisms, which allowed us to predict a putative protein-protein interaction between the Arabidopsis thaliana deacetylase HDA15 and the MADS-domain protein XAANTAL1 (XAL1). The results of two-hybrid and Bimolecular Fluorescence Complementation analysis demonstrated in vitro and in vivo HDA15-XAL1 interaction in the nucleus. Likely, this interaction might regulate developmental processes in plants as is the case for this type of interaction in animals.

AabHLH48, a novel basic helix-loop-helix transcription factor from Adonis amurensis, promotes early flowering in Arabidopsis by activating FRUITFULL expression.

Basic helix-loop-helix (bHLH) transcription factors play various important roles in plant growth and development. In this study, a AabHLH48 was identified in the floral organ of Adonis amurensis, a perennial herb that can naturally complete flowering at extreme low temperatures. AabHLH48 was widely expressed in various tissues or organs of A. amurensis and was localized in the nucleus. Overexpression of AabHLH48 promotes early flowering in Arabidopsis under both photoperiod (12 h light/12 h dark and 16 h light/8 h dark) and temperature (22 and 18 °C) conditions. Transcriptome sequencing combined with quantitative real-time PCR analysis showed that overexpression of AabHLH48 caused a general upregulation of genes involved in floral development in Arabidopsis, especially for AtAGAMOUS-LIKE 8/FRUITFULL (AtAGL8/FUL). The yeast one-hybrid assay revealed that AabHLH48 has transcriptional activating activity and can directly bind to the promoter region of AtAGL8/FUL. These results suggest that the overexpression of AabHLH48 promoting early flowering in Arabidopsis is associated with the upregulated expression of AtAGL8/FUL activated by AabHLH48. This indicates that AabHLH48 can serve as an important genetic resource for improving flowering-time control in other ornamental plants or crops.

Analysis of pea mutants reveals the conserved role of FRUITFULL controlling the end of flowering and its potential to boost yield.

Monocarpic plants have a single reproductive phase in their life. Therefore, flower and fruit production are restricted to the length of this period. This reproductive strategy involves the regulation of flowering cessation by a coordinated arrest of the growth of the inflorescence meristems, optimizing resource allocation to ensure seed filling. Flowering cessation appears to be a regulated phenomenon in all monocarpic plants. Early studies in several species identified seed production as a major factor triggering inflorescence proliferative arrest. Recently, genetic factors controlling inflorescence arrest, in parallel to the putative signals elicited by seed production, have started to be uncovered in Arabidopsis, with the MADS-box gene FRUITFULL (FUL) playing a central role in the process. However, whether the genetic network regulating arrest is also at play in other species is completely unknown. Here, we show that this role of FUL is not restricted to Arabidopsis but is conserved in another monocarpic species with a different inflorescence structure, field pea, strongly suggesting that the network controlling the end of flowering is common to other plants. Moreover, field trials with lines carrying mutations in pea FUL genes show that they could be used to boost crop yield.

Evolutionary analysis of MADS-box genes in buckwheat species and functional study of FdMADS28 in flavonoid metabolism.

The MADS-box gene family is a transcription factor family that is widely expressed in plants. It controls secondary metabolic processes in plants and encourages the development of tissues like roots and flowers. However, the phylogenetic analysis and evolutionary model of MADS-box genes in Fagopyrum species has not been reported yet. This study identified the MADS-box genes of three buckwheat species at the whole genome level, and conducted systematic evolution and physicochemical analysis. The results showed that these genes can be divided into four subfamilies, with fragment duplication being the main way for the gene family expansion. During the domestication process from golden buckwheat to tartary buckwheat and the common buckwheat, the Ka/Ks ratio indicated that most members of the family experienced strong purification selection pressure, and with individual gene pairs experiencing positive selection. In addition, we combined the expression profile data of the MADS genes, mGWAS data, and WGCNA data to mine genes FdMADS28/48/50 that may be related to flavonoid metabolism. The results also showed that overexpression of FdMADS28 could increase rutin content by decreasing Kaempferol pathway content in hairy roots, and increase the resistance and growth of hairy roots to PEG and NaCl. This study systematically analyzed the evolutionary relationship of MADS-box genes in the buckwheat species, and elaborated on the expression patterns of MADS genes in different tissues under biotic and abiotic stresses, laying an important theoretical foundation for further elucidating their role in flavonoid metabolism.

[Brassica juncea BjuWRKY71-1 accelerates flowering by regulating the expression of SOC1].

Brassica juncea (mustard) is a vegetable crop of Brassica, which is widely planted in China. The yield and quality of stem mustard are greatly influenced by the transition from vegetative growth to reproductive growth, i.e., flowering. The WRKY transcription factor family is ubiquitous in higher plants, and its members are involved in the regulation of many growth and development processes, including biological/abiotic stress responses and flowering regulation. WRKY71 is an important member of the WRKY family. However, its function and mechanism in mustard have not been reported. In this study, the BjuWRKY71-1 gene was cloned from B. juncea. Bioinformatics analysis and phylogenetic tree analysis showed that the protein encoded by BjuWRKY71-1 has a conserved WRKY domain, belonging to class Ⅱ WRKY protein, which is closely related to BraWRKY71-1 in Brassica rapa. The expression abundance of BjuWRKY71-1 in leaves and flowers was significantly higher than that in roots and stems, and the expression level increased gradually along with plant development. The result of subcellular localization showed that BjuWRKY71-1 protein was located in nucleus. The flowering time of overexpressing BjuWRKY71-1 Arabidopsis plants was significantly earlier than that of the wild type. Yeast two-hybrid assay and dual-luciferase reporter assay showed that BjuWRKY71-1 interacted with the promoter of the flowering integrator BjuSOC1 and promoted the expression of its downstream genes. In conclusion, BjuWRKY71-1 protein can directly target BjuSOC1 to promote plant flowering. This discovery may facilitate further clarifying the molecular mechanism of BjuWRKY71-1 in flowering time control, and creating new germplasm with bolting and flowering tolerance in mustard.

Genome-wide analysis of MADS-box transcription factor gene family in wild emmer wheat (Triticum turgidum subsp. dicoccoides).

The members of MADS-box gene family have important roles in regulating the growth and development of plants. MADS-box genes are highly regarded for their potential to enhance grain yield and quality under shifting global conditions. Wild emmer wheat (Triticum turgidum subsp. dicoccoides) is a progenitor of common wheat and harbors valuable traits for wheat improvement. Here, a total of 117 MADS-box genes were identified in the wild emmer wheat genome and classified to 90 MIKCC, 3 MIKC*, and 24 M-type. Furthermore, a phylogenetic analysis and expression profiling of the emmer wheat MADS-box gene family was presented. Although some MADS-box genes belonging to SOC1, SEP1, AGL17, and FLC groups have been expanded in wild emmer wheat, the number of MIKC-type MADS-box genes per subgenome is similar to that of rice and Arabidopsis. On the other hand, M-type genes of wild emmer wheat is less frequent than that of Arabidopsis. Gene expression patterns over different tissues and developmental stages agreed with the subfamily classification of MADS-box genes and was similar to common wheat and rice, indicating their conserved functionality. Some TdMADS-box genes are also differentially expressed under drought stress. The promoter region of each of the TdMADS-box genes harbored 6 to 48 responsive elements, mainly related to light, however hormone, drought, and low-temperature related cis-acting elements were also present. In conclusion, the results provide detailed information about the MADS-box genes of wild emmer wheat. The present work could be useful in the functional genomics efforts toward breeding for agronomically important traits in T. dicoccoides.

Exploring the molecular regulation of vernalization-induced flowering synchrony in Arabidopsis.

Many plant populations exhibit synchronous flowering, which can be advantageous in plant reproduction. However, molecular mechanisms underlying flowering synchrony remain poorly understood. We studied the role of known vernalization-response and flower-promoting pathways in facilitating synchronized flowering in Arabidopsis thaliana. Using the vernalization-responsive Col-FRI genotype, we experimentally varied germination dates and daylength among individuals to test flowering synchrony in field and controlled environments. We assessed the activity of flowering regulation pathways by measuring gene expression across leaves produced at different time points during development and through a mutant analysis. We observed flowering synchrony across germination cohorts in both environments and discovered a previously unknown process where flower-promoting and repressing signals are differentially regulated between leaves that developed under different environmental conditions. We hypothesized this mechanism may underlie synchronization. However, our experiments demonstrated that signals originating from sources other than leaves must also play a pivotal role in synchronizing flowering time, especially in germination cohorts with prolonged growth before vernalization. Our results suggest flowering synchrony is promoted by a plant-wide integration of flowering signals across leaves and among organs. To summarize our findings, we propose a new conceptual model of vernalization-induced flowering synchrony and provide suggestions for future research in this field.

SISTER OF FCA physically associates with SKB1 to regulate flowering time in Arabidopsis thaliana.

BACKGROUND: Proper flowering time is important for the growth and development of plants, and both too early and too late flowering impose strong negative influences on plant adaptation and seed yield. Thus, it is vitally important to study the mechanism underlying flowering time control in plants. In a previous study by the authors, genome-wide association analysis was used to screen the candidate gene SISTER OF FCA (SSF) that regulates FLOWERING LOCUS C (FLC), a central gene encoding a flowering suppressor in Arabidopsis thaliana. RESULTS: SSF physically interacts with Protein arginine methyltransferase 5 (PRMT5, SKB1). Subcellular co-localization analysis showed that SSF and SKB1 interact in the nucleus. Genetically, SSF and SKB1 exist in the same regulatory pathway that controls FLC expression. Furthermore, RNA-sequencing analysis showed that both SSF and SKB1 regulate certain common pathways. CONCLUSIONS: This study shows that PRMT5 interacts with SSF, thus controlling FLC expression and facilitating flowering time control.

Two SEPALLATA MADS-Box Genes, SlMBP21 and SlMADS1, Have Cooperative Functions Required for Sepal Development in Tomato.

MADS-box transcription factors have crucial functions in numerous physiological and biochemical processes during plant growth and development. Previous studies have reported that two MADS-box genes, SlMBP21 and SlMADS1, play important regulatory roles in the sepal development of tomato, respectively. However, the functional relationships between these two genes are still unknown. In order to investigate this, we simultaneously studied these two genes in tomato. Phylogenetic analysis showed that they were classified into the same branch of the SEPALLATA (SEP) clade. qRT-PCR displayed that both SlMBP21 and SlMADS1 transcripts are preferentially accumulated in sepals, and are increased with flower development. During sepal development, SlMBP21 is increased but SlMADS1 is decreased. Using the RNAi, tomato plants with reduced SlMBP21 mRNA generated enlarged and fused sepals, while simultaneous inhibition of SlMBP21 and SlMADS1 led to larger (longer and wider) and fused sepals than that in SlMBP21-RNAi lines. qRT-PCR results exhibited that the transcripts of genes relating to sepal development, ethylene, auxin and cell expansion were dramatically changed in SlMBP21-RNAi sepals, especially in SlMBP21-SlMADS1-RNAi sepals. Yeast two-hybrid assay displayed that SlMBP21 can interact with SlMBP21, SlAP2a, TAGL1 and RIN, and SlMADS1 can interact with SlAP2a and RIN, respectively. In conclusion, SlMBP21 and SlMADS1 cooperatively regulate sepal development in tomato by impacting the expression or activities of other related regulators or via interactions with other regulatory proteins.

Transcriptome sequencing and screening of genes related to the MADS-box gene family in Clematis courtoisii.

The MADS-box gene family controls plant flowering and floral organ development; therefore, it is particularly important in ornamental plants. To investigate the genes associated with the MADS-box family in Clematis courtoisii, we performed full-length transcriptome sequencing on C. courtoisii using the PacBio Sequel third-generation sequencing platform, as no reference genome data was available. A total of 12.38 Gb of data, containing 9,476,585 subreads and 50,439 Unigenes were obtained. According to functional annotation, a total of 37,923 Unigenes (75.18% of the total) were assigned with functional annotations, and 50 Unigenes were identified as MADS-box related genes. Subsequently, we employed hmmerscan to perform protein sequence similarity search for the translated Unigene sequences and successfully identified 19 Unigenes associated with the MADS-box gene family, including MIKC*(1) and MIKCC (18) genes. Furthermore, within the MIKCC group, six subclasses can be further distinguished.

IiAGL6 participates in the regulation of stamen development and pollen formation in Isatis indigotica.

The AGL6 (AGMOUSE LIKE 6) gene is a member of the SEP subfamily and functions as an E-class floral homeotic gene in the development of floral organs. In this study, we cloned IiAGL6, the orthologous gene of AGL6 in Isatis indigotica. The constitutive expression of IiAGL6 in Arabidopsis thaliana resulted in a late-flowering phenotype and the development of curly leaves during the vegetative growth period. Abnormal changes in floral organ development were observed during the reproductive stage. In woad plants, suppression of IiAGL6 using TRV-VIGS (tobacco rattle virus-mediated virus-induced gene silencing) decreased the number of stamens and led to the formation of aberrant anthers. Similar changes in stamen development were also observed in miRNA-AGL6 transgenic Arabidopsis plants. Yeast two-hybrid and BiFC tests showed that IiAGL6 can interact with other MADS-box proteins in woad; thus, playing a key role in defining the identities of floral organs, particularly during stamen formation. These findings might provide novel insights and help investigate the biological roles of MADS transcription factors in I. indigotica.

COOLAIR and PRC2 function in parallel to silence FLC during vernalization.

Noncoding transcription induces chromatin changes that can mediate environmental responsiveness, but the causes and consequences of these mechanisms are still unclear. Here, we investigate how antisense transcription (termed COOLAIR) interfaces with Polycomb Repressive Complex 2 (PRC2) silencing during winter-induced epigenetic regulation of Arabidopsis FLOWERING LOCUS C (FLC). We use genetic and chromatin analyses on lines ineffective or hyperactive for the antisense pathway in combination with computational modeling to define the mechanisms underlying FLC repression. Our results show that FLC is silenced through pathways that function with different dynamics: a COOLAIR transcription-mediated pathway capable of fast response and in parallel a slow PRC2 switching mechanism that maintains each allele in an epigenetically silenced state. Components of both the COOLAIR and PRC2 pathways are regulated by a common transcriptional regulator (NTL8), which accumulates by reduced dilution due to slow growth at low temperature. The parallel activities of the regulatory steps, and their control by temperature-dependent growth dynamics, create a flexible system for registering widely fluctuating natural temperature conditions that change year on year, and yet ensure robust epigenetic silencing of FLC.

Genome-wide association study identifies a novel BMI1A QTL allele that confers FLC expression diversity in Arabidopsis thaliana.

Identification and understanding of the genetic basis of natural variations in plants are essential for comprehending their phenotypic adaptation. Here, we report a genome-wide association study (GWAS) of FLOWERING LOCUS C (FLC) expression in 727 Arabidopsis accessions. We identified B LYMPHOMA MOLONEY MURINE LEUKEMIA VIRUS INSERTION REGION 1 HOMOLOG 1A (BMI1A) as a causal gene for one of the FLC expression quantitative trait loci (QTLs). Loss of function in BMI1A increases FLC expression and delays flowering time at 16 °C significantly compared with the wild type (Col-0). BMI1A activity is required for histone H3 lysine 27 trimethylation (H3K27me3) accumulation at the FLC, MADS AFFECTING FLOWERING 4 (MAF4), and MAF5 loci at low ambient temperature. We further uncovered two BMI1A haplotypes associated with the natural variation in FLC expression and flowering time at 16 °C, and demonstrated that polymorphisms in the BMI1A promoter region are the main contributor. Different BMI1A haplotypes are strongly associated with geographical distribution, and the low ambient temperature-sensitive BMI1A variants are associated with a lower mean temperature of the driest quarter of their collection sites compared with the temperature-non-responsive variants, indicating that the natural variations in BMI1A have adaptive functions in FLC expression and flowering time regulation. Therefore, our results provide new insights into the natural variations in FLC expression and flowering time diversity in plants.

HISTONE DEACETYLASE19 Controls Ovule Number Determination and Transmitting Tract Differentiation.

The gynoecium is critical for the reproduction of flowering plants as it contains the ovules and the tissues that foster pollen germination, growth, and guidance. These tissues, known as the reproductive tract (ReT), comprise the stigma, style, and transmitting tract (TT). The ReT and ovules originate from the carpel margin meristem (CMM) within the pistil. SHOOT MERISTEMLESS (STM) is a key transcription factor for meristem formation and maintenance. In all above-ground meristems, including the CMM, local STM downregulation is required for organ formation. However, how this downregulation is achieved in the CMM is unknown. Here, we have studied the role of HISTONE DEACETYLASE 19 (HDA19) in Arabidopsis (Arabidopsis thaliana) during ovule and ReT differentiation based on the observation that the hda19-3 mutant displays a reduced ovule number and fails to differentiate the TT properly. Fluorescence-activated cell sorting coupled with RNA-sequencing revealed that in the CMM of hda19-3 mutants, genes promoting organ development are downregulated while meristematic markers, including STM, are upregulated. HDA19 was essential to downregulate STM in the CMM, thereby allowing ovule formation and TT differentiation. STM is ectopically expressed in hda19-3 at intermediate stages of pistil development, and its downregulation by RNA interference alleviated the hda19-3 phenotype. Chromatin immunoprecipitation assays indicated that STM is a direct target of HDA19 during pistil development and that the transcription factor SEEDSTICK is also required to regulate STM via histone acetylation. Thus, we identified factors required for the downregulation of STM in the CMM, which is necessary for organogenesis and tissue differentiation.

FLZ13 interacts with FLC and ABI5 to negatively regulate flowering time in Arabidopsis.

The transition from vegetative to reproductive growth, known as flowering, is a critical developmental process in flowering plants to ensure reproductive success. This process is strictly controlled by various internal and external cues; however, the underlying molecular regulatory mechanisms need to be further characterized. Here, we report a plant-specific protein, FCS-LIKE ZINC FINGER PROTEIN 13 (FLZ13), which functions as a hitherto unknown negative modulator of flowering time in Arabidopsis thaliana. Biochemical analysis showed that FLZ13 directly interacts with FLOWERING LOCUS C (FLC), a major flowering repressor, and that FLZ13 largely depends on FLC to repress the transcription of two core flowering integrators: FLOWERING LOCUS T and SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1. In addition, FLZ13 works together with ABSCISIC ACID INSENSITIVE 5 to activate FLC expression to delay flowering. Taken together, our findings suggest that FLZ13 is an important component of the gene regulatory network for flowering time control in plants.

The nuclear pore Y-complex functions as a platform for transcriptional regulation of FLOWERING LOCUS C in Arabidopsis.

The nuclear pore complex (NPC) has multiple functions beyond the nucleo-cytoplasmic transport of large molecules. Subnuclear compartmentalization of chromatin is critical for gene expression in animals and yeast. However, the mechanism by which the NPC regulates gene expression is poorly understood in plants. Here we report that the Y-complex (Nup107-160 complex, a subcomplex of the NPC) self-maintains its nucleoporin homeostasis and modulates FLOWERING LOCUS C (FLC) transcription via changing histone modifications at this locus. We show that Y-complex nucleoporins are intimately associated with FLC chromatin through their interactions with histone H2A at the nuclear membrane. Fluorescence in situ hybridization assays revealed that Nup96, a Y-complex nucleoporin, enhances FLC positioning at the nuclear periphery. Nup96 interacted with HISTONE DEACETYLASE 6 (HDA6), a key repressor of FLC expression via histone modification, at the nuclear membrane to attenuate HDA6-catalyzed deposition at the FLC locus and change histone modifications. Moreover, we demonstrate that Y-complex nucleoporins interact with RNA polymerase II to increase its occupancy at the FLC locus, facilitating transcription. Collectively, our findings identify an attractive mechanism for the Y-complex in regulating FLC expression via tethering the locus at the nuclear periphery and altering its histone modification.

Antagonistic MADS-box transcription factors SEEDSTICK and SEPALLATA3 form a transcriptional regulatory network that regulates seed oil accumulation.

Transcriptional regulation is essential for balancing multiple metabolic pathways that influence oil accumulation in seeds. Thus far, the transcriptional regulatory mechanisms that govern seed oil accumulation remain largely unknown. Here, we identified the transcriptional regulatory network composed of MADS-box transcription factors SEEDSTICK (STK) and SEPALLATA3 (SEP3), which bridges several key genes to regulate oil accumulation in seeds. We found that STK, highly expressed in the developing embryo, positively regulates seed oil accumulation in Arabidopsis (Arabidopsis thaliana). Furthermore, we discovered that SEP3 physically interacts with STK in vivo and in vitro. Seed oil content is increased by the SEP3 mutation, while it is decreased by SEP3 overexpression. The chromatin immunoprecipitation, electrophoretic mobility shift assay, and transient dual-luciferase reporter assays showed that STK positively regulates seed oil accumulation by directly repressing the expression of MYB5, SEP3, and SEED FATTY ACID REDUCER 4 (SFAR4). Moreover, genetic and molecular analyses demonstrated that STK and SEP3 antagonistically regulate seed oil production and that SEP3 weakens the binding ability of STK to MYB5, SEP3, and SFAR4. Additionally, we demonstrated that TRANSPARENT TESTA 8 (TT8) and ACYL-ACYL CARRIER PROTEIN DESATURASE 3 (AAD3) are direct targets of MYB5 during seed oil accumulation in Arabidopsis. Together, our findings provide the transcriptional regulatory network antagonistically orchestrated by STK and SEP3, which fine tunes oil accumulation in seeds.

Functional analysis of a lily SHORT VEGETATIVE PHASE ortholog in flowering transition and floral development.

Lilium is a commercially important genus of bulbous flowers, investigating the flowering molecular mechanisms is important for flowering regulation of lily. MADS-box SHORT VEGETATIVE PHASE (SVP) orthologs are involved in the flowering transition and floral organ differentiation in many plants. In this study, we identified an SVP ortholog from L. × formolongi (LfSVP), which was closely related to Arabidopsis SVP according to phylogenetic analysis. Tissue-specific expression patterns indicated that LfSVP expression levels peaked in the leaves and showed low expression levels in flowering tepals. Stage-dependent expression patterns of LfSVP showed high transcription level in the flowering induction stage under different photoperiods and exhibited transcription peak in the floral budding development stage under long days. Overexpressed LfSVP led to delayed flowering and floral organ defects in Arabidopsis independent of photoperiod. Tobacco rattle virus -induced gene silencing of LfSVP caused a strongly earlier flowering time and floral organ defects of L. × formolongi. Moreover, LfSVP can interact with L. × formolongi APETALA1 (AP1) in both yeast and tobacco cells, and the two may interact to regulate floral organ differentiation. In conclusion, LfSVP is a flowering repressor and may be involved in the regulation of floral organ differentiation. This study will be helpful for the molecular breeding of short-life-period and rich floral patterns lily varieties.

Arabidopsis SEC13B Interacts with Suppressor of Frigida 4 to Repress Flowering.

SECRETORY13 (SEC13) is an essential member of the coat protein complex II (COPII), which was reported to mediate vesicular-specific transport from the endoplasmic reticulum (ER) to the Golgi apparatus and plays a crucial role in early secretory pathways. In Arabidopsis, there are two homologous proteins of SEC13: SEC13A and SEC13B. SUPPRESSOR OF FRIGIDA 4 (SUF4) encodes a C2H2-type zinc finger protein that inhibits flowering by transcriptionally activating the FLOWERING LOCUS C (FLC) through the FRIGIDA (FRI) pathway in Arabidopsis. However, it remains unclear whether SEC13 proteins are involved in Arabidopsis flowering. In this study, we first identified that the sec13b mutant exhibited early flowering under both long-day and short-day conditions. Quantitative real-time PCR (qRT-PCR) analysis showed that both SEC13A and SEC13B were expressed in all the checked tissues, and transient expression assays indicated that SEC13A and SEC13B were localized not only in the ER but also in the nucleus. Then, we identified that SEC13A and SEC13B could interact with SUF4 in vitro and in vivo. Interestingly, both sec13b and suf4 single mutants flowered earlier than the wild type (Col-0), whereas the sec13b suf4 double mutant flowered even earlier than all the others. In addition, the expression of flowering inhibitor FLC was down-regulated, and the expressions of flowering activator FLOWERING LOCUS T (FT), CONSTANS (CO), and SUPPRESSOR OF OVEREXPRESSION OF CO 1 (SOC1) were up-regulated in sec13b, suf4, and sec13b suf4 mutants, compared with Col-0. Taken together, our results indicated that SEC13B interacted with SUF4, and they may co-regulate the same genes in flowering-regulation pathways. These results also suggested that the COPII component could function in flowering in Arabidopsis.