SQUAMOSA Promoter Binding Protein-like Gene Family Research Articles

Genome-wide identification and analysis of SPL gene family in chickpea (Cicer arietinum L.).

A transcription factor in plants encodes SQUAMOSA promoter binding protein-like (SPL) serves a broad spectrum of important roles for the plant, like, growth, flowering, and signal transduction. A gene family that encodes SPL proteins is documented in various model plant species, including Arabidopsis thaliana and Oryza sativa. Chickpea (Cicer arietinum), a leguminous crop, has not been thoroughly explored with regard to the SPL protein-encoding gene family. Chickpea SPL family genes were located and characterized computationally using a genomic database. Gene data of chickpea were obtained from the phytozome repository and was examined using bioinformatics methods. For investigating the possible roles of SPLs in chickpea, genome-wide characterization, expression, as well as structural analyses of this SPL gene family were performed. Cicer arietinum genome had 19 SPL genes, whereas, according to phylogenetic analysis, the SPLs in chickpea are segregated among four categories: Group-I has 2 introns, Group-II and IV have 1-2 introns (except CaSPL13 and CaSPL15 having 3 introns), and Group-III has 9 introns (except CaSPL1 and CaSPL11 with 1 and 8 introns, respectively). The SBP domain revealed that SPL proteins featured two zinc-binding sites, i.e., C3H and C2HC and one nuclear localization signal. All CaSPL proteins are found to contain highly conserved motifs, i.e., Motifs 1, 2, and 4, except CaSPL10 in which Motifs 1 and 4 were absent. Following analysis, it was found that Motifs 1 and 2 of the chickpea SBP domain are Zinc finger motifs, and Motif 4 includes a nuclear localization signal. All pairs of CaSPL paralogs developed by purifying selection. The CaSPL promoter investigation discovered cis-elements that are responsive to stress, light, and phytohormones. Examination of their expression patterns highlighted major CaSPLs to be evinced primarily among younger pods and flowers. Indicating their involvement in the plant's growth as well as development, along with their capacity to react as per different situations by handling the regulation of target gene's expression, several CaSPL genes are also expressed under certain stress conditions, namely, cold, salt, and drought. The majority of the CaSPL genes are widely expressed and play crucial roles in terms of the plant's growth, development, and responses to the environmental-stress conditions. Our work provides extensive insight into the gene family CaSPL, which might facilitate further studies related to the evolution and functions of the SPL genes for chickpea and other plant species.

Genome-wide investigation of SQUAMOSA promoter binding protein-like genes in Liriodendron and functional characterization of LcSPL2.

The plant-specific SQUAMOSA promoter-binding protein-like (SPL) transcription factors play a pivotal role in various developmental processes, including leaf morphogenesis and vegetative to reproductive phase transition. Liriodendron chinense and Liriodendron tulipifera are widely used in landscaping due to their tulip-like flowers and peculiar leaves. However, the SPL gene family in Liriodendron has not been identified and systematically characterized. We systematically identified and characterized the SPL family members in Liriodendron, including phylogeny, gene structure and syntenic analyses. Subsequently, we quantified the expression patterns of LcSPLs across various tissue sites through transcription-quantitative polymerase chain reaction (RT-qPCR) assays and identified the target gene, LcSPL2. Finally, we characterized the functions of LcSPL2 via ectopic transformation. Altogether, 17 LcSPL and 18 LtSPL genes were genome-widely identified in L. chinense and L. tulipifera, respectively. All the 35 SPLs were grouped into 9 clades. Both species had three SPL gene pairs arising from segmental duplication events, and the LcSPLs displayed high collinearity with the L. tulipifera genome. RT-qPCR assays showed that SPL genes were differentially expressed in different tissues, especially. Because LcSPL2 is highly expressed in pistils and leaves, it was selected to describe the SPL gene family of L. chinense by ectopic expression. We showed that overexpression of LcSPL2 in Arabidopsis thaliana resulted in earlier flowering and fewer rosette leaves. Moreover, we observed that overexpression of LcSPL2 in A. thaliana up-regulated the expression levels of four genes related to flower development. This study identified SPL genes in Liriodendron and characterized the function of LcSPL2 in advancing flower development.

Comparative analysis of SPL transcription factors from streptophyte algae and embryophytes reveals evolutionary trajectories of SPL family in streptophytes

SQUAMOSA-PROMOTER BINDING PROTEIN-LIKE (SPL) genes encode plant-specific transcription factors which are important regulators of diverse plant developmental processes. We took advantage of available genome sequences of streptophyte algae representatives to investigate the relationships of SPL genes between freshwater green algae and land plants. Our analysis showed that streptophyte algae, hornwort and liverwort genomes encode from one to four SPL genes which is the smallest set, in comparison to other land plants studied to date. Based on the phylogenetic analysis, four major SPL phylogenetic groups were distinguished with Group 3 and 4 being sister to Group 1 and 2. Comparative motif analysis revealed conserved protein motifs within each phylogenetic group and unique bryophyte-specific motifs within Group 1 which suggests lineage-specific protein speciation processes. Moreover, the gene structure analysis also indicated the specificity of each by identifying differences in exon–intron structures between the phylogenetic groups, suggesting their evolutionary divergence. Since current understanding of SPL genes mostly arises from seed plants, the presented comparative and phylogenetic analyzes from freshwater green algae and land plants provide new insights on the evolutionary trajectories of the SPL gene family in different classes of streptophytes.

Genome-Wide Identification and Expression Analysis of the SQUAMOSA Promoter-Binding Protein-like (SPL) Transcription Factor Family in Catalpabungei.

As a plant-specific transcription factor, the SPL gene family plays a critical role in plant growth and development. Although the SPL gene family has been identified in diverse plant species, there have been no genome-wide identification or systematic study reports on the SPL gene family in Catalpa bungei. In this study, we identified 19 putative SPL gene family members in the C. bungei genome. According to the phylogenetic relationship, they can be divided into eight groups, and the genes in the same group have a similar gene structure and conserved motifs. Synteny analysis showed that fragment duplication played an important role in the expansion of the CbuSPL gene family. At the same time, CbuSPL genes have cis-acting elements and functions related to light response, hormone response, growth and development, and stress response. Tissue-specific expression and developmental period-specific expression analysis showed that CbuSPL may be involved in flowering initiation and development, flowering transition, and leaf development. In addition, the ectopic expression of CbuSPL4 in Arabidopsis confirmed that it can promote early flowering and induce the expression of related flowering genes. These systematic research results will lay a foundation for further study on the functional analysis of SPL genes in C. bungei.

Genome-Wide Analysis of SPL/miR156 Module and Its Expression Analysis in Vegetative and Reproductive Organs of Oil Palm (Elaeis guineensis).

The SPL (SQUAMOSA-promoter binding protein-like) gene family is one of the largest plant transcription factors and is known to be involved in the regulation of plant growth, development, and stress responses. The genome-wide analysis of SPL gene members in a diverse range of crops has been elucidated. However, none of the genome-wide studies on the SPL gene family have been carried out for oil palm, an important oil-yielding plant. In this research, a total of 24 EgSPL genes were identified via a genome-wide approach. Phylogenetic analysis revealed that most of the EgSPLs are closely related to the Arabidopsis and rice SPL gene members. EgSPL genes were mapped onto the only nine chromosomes of the oil palm genome. Motif analysis revealed conservation of the SBP domain and the occurrence of 1-10 motifs in EgSPL gene members. Gene duplication analysis demonstrated the tandem duplication of SPL members in the oil palm genome. Heatmap analysis indicated the significant expression of SPL genes in shoot and flower organs of oil palm plants. Among the identified EgSPL genes, a total 14 EgSPLs were shown to be targets of miR156. Real-time PCR analysis of 14 SPL genes showed that most of the EgSPL genes were more highly expressed in female and male inflorescences of oil palm plants than in vegetative tissues. Altogether, the present study revealed the significant role of EgSPL genes in inflorescence development.

Genome-Wide Analysis of SPL Gene Families Illuminate the Evolution Patterns in Three Rubber-Producing Plants

Transcription factors SQUAMOSA Promoter-binding Protein-like (SPL) play a crucial role in regulating plant response to stress, root development, and flower production. However, analysis of SPL gene families in the three rubber-producing plants Taraxacum kok-saghyz, Hevea brasiliensis, and Eucommia ulmoides, renowned for their natural rubber production, has not yet been conducted. In this study, we utilized reference genomes to perform genome-wide analysis, and obtained new insights on the evolution of SPL gene families in these three rubber-producing plants. Our results revealed the following: (1) T. kok-saghyz, H. brasiliensis, and E. ulmoides harbored 25, 16, and 13 SPL genes, respectively, containing conserved structural domains of SBP. (2) A phylogenetic analysis categorized 90 SPL proteins from 25 TkSPLs, 16 HbSPLs, 13 EuSPLs, 17 AtSPLs, and 19 OsSPLs into eight groups. (3) Analysis of cis-acting elements demonstrated that the promoters of EuSPLs contained a significant number of light response elements, hormone regulatory elements, and stress response elements. (4) Transcriptome data analysis revealed that the EuSPL8 gene had strong expression in bark, as well as TkSPL4 and TkSPL8 exhibit high expression levels specifically in roots and latex. This study provides valuable insights into the biological functions of the SPL gene family in the three rubber plants and might serve as a reference for identifying efficient genes.

Genome-Wide Identification of the SQUAMOSA Promoter-Binding Protein-like (SPL) Transcription Factor Family in Sweet Cherry Fruit.

Plant-specific SQUAMOSA promoter-binding protein-like (SPL) transcription factors play important regulatory roles during plant growth and development, fruit ripening, inflorescence branching, and biotic and abiotic stresses. However, there have been no identification or systematic studies of the SPL gene family in the sweet cherry. In this study, 12 SPL genes were identified in the sweet cherry reference genome, which were distributed over 6 chromosomes and classified into six groups according to phylogenetic relationships with other SPL gene families. Nine PavSPLs were highly expressed at green fruit stages and dramatically decreased at the onset of fruit ripening, which implied that they were important regulators during fruit development and ripening. The expression patterns of PavSPL genes under ABA, GA, and MeJA treatments showed that the PavSPLs were involved in the process of fruit ripening. A subcellular localization experiment proved that PavSPL4 and PavSPL7 proteins were localized in the nucleus. The genome-wide identification of the SPL gene family provided new insights while establishing an important foundation for sweet cherry studies.

Genome-Wide Analysis of the SPL Gene Family and Expression Analysis during Flowering Induction in Prunus × yedoensis 'Somei-yoshino'.

SQUAMOSA Promoter-Binding Protein-Like (SPL) genes encode plant-specific transcription factors which bind to the SQUAMOSA promoter of the MADS-box genes to regulate its expression. It plays important regulatory roles in floral induction and development, fertility, light signals and hormonal transduction, and stress response in plants. In this study, 32 PySPL genes with complete SBP (squamosa promoter binding protein) conserved domain were identified from the genome of Prunus × yedoensis ‘Somei-yoshino’ and analyzed by bioinformatics. 32 PySPLs were distributed on 13 chromosomes, encoding 32 PySPL proteins with different physical and chemical properties. The phylogenetic tree constructed with Arabidopsis thaliana and Oryza sativa can be divided into 10 subtribes, indicating PySPLs of different clusters have different biological functions. The conserved motif prediction showed that the number and distribution of motifs on each PySPL is varied. The gene structure analysis revealed that PySPLs harbored exons ranging from 2 to 10. The predictive analysis of acting elements showed that the promoter of PySPLs contain a large number of light-responsive elements, as well as response elements related to hormone response, growth and development and stress response. The analysis of the PySPLs expressions in flower induction and flower organs based on qRT-PCR showed that PySPL06/22 may be the key genes of flower development, PySPL01/06 and PySPL22 may play a role in the development of sepal and pistil, respectively. The results provide a foundation for the study of SPL transcription factors of Prunus × yedoensis ‘Somei-yoshino’ and provide more reference information of the function of SPL gene in flowering.

The Mulberry SPL Gene Family and the Response of MnSPL7 to Silkworm Herbivory through Activating the Transcription of MnTT2L2 in the Catechin Biosynthesis Pathway.

SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPL) genes, as unique plant transcription factors, play important roles in plant developmental regulation and stress response adaptation. Although mulberry is a commercially valuable tree species, there have been few systematic studies on SPL genes. In this work, we identified 15 full-length SPL genes in the mulberry genome, which were distributed on 4 Morus notabilis chromosomes. Phylogenetic analysis clustered the SPL genes from five plants (Malus × domestica Borkh, Populus trichocarpa, M. notabilis, Arabidopsis thaliana, and Oryza sativa) into five groups. Two zinc fingers (Zn1 and Zn2) were found in the conserved SBP domain in all of the MnSPLs. Comparative analyses of gene structures and conserved motifs revealed the conservation of MnSPLs within a group, whereas there were significant structure differences among groups. Gene quantitative analysis showed that the expression of MnSPLs had tissue specificity, and MnSPLs had much higher expression levels in older mulberry leaves. Furthermore, transcriptome data showed that the expression levels of MnSPL7 and MnSPL14 were significantly increased under silkworm herbivory. Molecular experiments revealed that MnSPL7 responded to herbivory treatment through promoting the transcription of MnTT2L2 and further upregulating the expression levels of catechin synthesis genes (F3′H, DFR, and LAR).

Characterization and expression analysis of the SPL gene family during floral development and abiotic stress in pecan (Carya illinoinensis).

SQUAMOSA promoter binding protein-like (SPL) genes are a type of plant-specific transcription factors that play crucial roles in the regulation of phase transition, floral transformation, fruit development, and various stresses. Although SPLs have been characterized in several model species, no systematic analysis has been studied in pecans, an important woody oil tree species. In this study, a total of 32 SPL genes (CiSPLs) were identified in the pecan genome. After conducting phylogenetic analysis of the conserved SBP proteins from Arabidopsis, rice, and poplar, the CiSPLs were separated into eight subgroups. The CiSPL genes within the same subgroup contained very similar exon-intron structures and conserved motifs. Nine segmentally duplicated gene pairs in the pecan genome and 16 collinear gene pairs between the CiSPL and AtSPL genes were identified. Cis-element analysis showed that CiSPL genes may regulate plant meristem differentiation and seed development, participate in various biological processes, and respond to plant hormones and environmental stresses. Therefore, we focused our study on the expression profiles of CiSPL genes during flower and fruit development. Most of the CiSPL genes were predominantly expressed in buds and/or female flowers. Additionally, quantitative real time PCR (qRT-PCR) analyses confirmed that CiSPL genes showed distinct spatiotemporal expression patterns in response to drought and salt treatments. The study provides foundation for the further exploration of the function and evolution of SPL genes in pecan.

Genome-Wide Characterization of SPL Gene Family in Codonopsis pilosula Reveals the Functions of CpSPL2 and CpSPL10 in Promoting the Accumulation of Secondary Metabolites and Growth of C. pilosula Hairy Root.

SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPL) transcription factors play critical roles in regulating diverse aspects of plant growth and development, including vegetative phase change, plant architecture, anthocyanin accumulation, lateral root growth, etc. In the present study, 15 SPL genes were identified based on the genome data of Codonopsis pilosula, a well-known medicinal plant. Phylogenetic analysis clustered CpSPLs into eight groups (G1-G8) along with SPLs from Arabidopsis thaliana, Solanum lycopersicum, Oryza sativa and Physcomitrella patens. CpSPLs in the same group share similar gene structure and conserved motif composition. Cis-acting elements responding to light, stress and phytohormone widely exist in their promoter regions. Our qRT-PCR results indicated that 15 CpSPLs were differentially expressed in different tissues (root, stem, leaf, flower and calyx), different developmental periods (1, 2 and 3 months after germination) and various conditions (NaCl, MeJA and ABA treatment). Compared with the control, overexpression of CpSPL2 or CpSPL10 significantly promoted not only the growth of hairy roots, but also the accumulation of total saponins and lobetyolin. Our results established a foundation for further investigation of CpSPLs and provided novel insights into their biological functions. As far as we know, this is the first experimental research on gene function in C. pilosula.

Genome-wide identification, gene cloning, subcellular location and expression analysis of SPL gene family in P. granatum L

BackgroundsPomegranate is an excellent tree species with nutritional, medicinal, ornamental and ecological values. Studies have confirmed that SPL factors play an important role in floral transition and flower development.ResultsUsed bioinformatics methods, 15 SPL (SQUAMOSA promoter-binding protein-like) genes were identified and analyzed from the ‘Taishanhong’ pomegranate (P. granatum L.) genome. Phylogenetic analysis showed that PgSPLs were divided into six subfamilies (G1 ~ G6). PgSPL promoter sequences contained multiple cis-acting elements associated with abiotic stress or hormonal response. Based on the transcriptome data, expression profiles of different tissues and different developmental stages showed that PgSPL genes had distinct temporal and spatial expression characteristics. The expression analysis of miR156 in small RNA sequencing results showed that miR156 negatively regulated the expression of target genes. qRT-PCR analysis showed that the expression levels of PgSPL2, PgSPL3, PgSPL6, PgSPL11 and PgSPL14 in leaves were significantly higher than those in buds and stems (p < 0.05). The expression levels of PgSPL5, PgSPL12 and PgSPL13 in flower buds were significantly higher than that in leaves and stems (p < 0.05). The full-length of coding sequence of PgSPL5 and PgSPL13 were obtained by homologous cloning technology. The full length of PgSPL5 is 1020 bp, and PgSPL13 is 489 bp, which encodes 339 and 162 amino acids, respectively. Further investigation revealed that PgSPL5 and PgSPL13 proteins were located in the nucleus. Exogenous plant growth regulator induction experiments showed that PgSPL5 was up-regulated in leaves and stems. PgSPL13 was up-regulated in leaves and down-regulated in stems. When sprayed with 6-BA, IBA and PP333 respectively, PgSPL5 and PgSPL13 were up-regulated most significantly at P2 (bud vertical diameter was 5.1 ~ 12.0 mm) stage of bisexual and functional male flowers.ConclusionsOur findings suggested that PgSPL2, PgSPL3, PgSPL6, PgSPL11 and PgSPL14 played roles in leaves development of pomegranate. PgSPL5, PgSPL12 and PgSPL13 played roles in pomegranate flower development. PgSPL5 and PgSPL13 were involved in the response process of different plant hormone signal transduction in pomegranate development. This study provided a robust basis for further functional analyses of SPL genes in pomegranate.

Genome-wide identification, phylogenetic analysis, and expression analysis of the SPL gene family in orchardgrass (Dactylis glomerata L.)

SPL (SQUAMOSA promoter binding protein-like) is a plant-specific transcription factor family that contains the conserved SBP domain, which plays a vital role in the vegetative-to-reproductive phase transition, flowering development and regulation, tillering/branching, and stress responses. Although the SPL family has been identified and characterized in various plant species, limited information about it has been obtained in orchardgrass, which is a critical forage crop worldwide. In this study, 17 putative DgSPL genes were identified among seven chromosomes, and seven groups that share similar gene structures and conserved motifs were determined by phylogenetic analysis. Of these, eight genes have potential target sites for miR156. cis-Element and gene ontology annotation analysis indicated DgSPLs may be involved in regulating development and abiotic stress responses. The expression patterns of eight DgSPL genes at five developmental stages, in five tissues, and under three stress conditions were determined by RNA-seq and qRT-PCR. These assays indicated DgSPLs are involved in vegetative-to-reproductive phase transition, floral development, and stress responses. The transient expression analysis in tobacco and heterologous expression assays in yeast indicated that miR156-targeted DG1G01828.1 and DG0G01071.1 are nucleus-localized proteins, that may respond to drought, salt, and heat stress. Our study represents the first systematic analysis of the SPL family in orchardgrass. This research provides a comprehensive assessment of the DgSPL family, which lays the foundation for further examination of the role of miR156/DgSPL in regulating development and stress responses in forages grasses.

Genome-wide identification and expression profiling of the SPL family genes in wheat

The SQUAMOSA promoter-binding protein-like (SPL) proteins constitute a family of plant-specific transcription factors that play a vital role in plant development. Wheat (Triticum aestivum, AABBDD) is universally well-known as a cash crop; however, the SPLs of this important crop have not been systematically investigated as yet. In this study, we conducted a genome-wide survey in wheat and found 56 SPL genes belonging to 19 homologous groups. The SPL genes were divided into seven classes using phylogenetic tree analyses. We mapped these genes onto the wheat chromosomes and examined their structures and conserved motifs. Moreover, we performed a synteny analysis on wheat, and summarized the SPL family as well as the evolutionary relationships between SPLs. Thereafter, we compared the expression patterns of wheat SPLs under different conditions, confirming that SPLs play an important role in spike development. To conclude, the SPLs in triplets have analogous structures and similar expression patterns. The three-pair triplet response to jasmonic acid (JA) and abscisic acid (ABA) was determined by quantitative real-time polymerase chain reaction (qRT-PCR). This work provides a comprehensive understanding of the SPL gene family in wheat. Our investigation of the wheat SPL gene family provides a starting point for additional functional studies of these significant transcription factors in wheat.

SQUAMOSA Promoter Binding Protein-Like (SPL) Gene Family: TRANSCRIPTOME-Wide Identification, Phylogenetic Relationship, Expression Patterns and Network Interaction Analysis in Panax ginseng C. A. Meyer.

SPL (SQUAMOSA promoter binding protein-like) gene family is specific transcription factor in the plant that have an important function for plant growth and development. Although the SPL gene family has been widely studied and reported in many various plant species from gymnosperm to angiosperm, there are no systematic studies and reports about the SPL gene family in Panax ginseng C. A. Meyer. In this study, we conducted transcriptome-wide identification, evolutionary analysis, structure analysis, and expression characteristics analysis of SPL gene family in Panax ginseng by bioinformatics. We annotated the PgSPL gene family and found that they might involve in multiple functions including encoding structural proteins, but the main function were still focused on the binding function. The result showed that 106 PgSPL transcripts were classified into two clades - A and B, both of which respectively consisted of three groups. Besides, we profiled PgSPL transcripts’ genotypic, temporal, and spatial expression characteristics. Furthermore, we calculated the correlation of PgSPL transcripts in the 14 tissues of a 4 years old ginseng and 42 farmers’ cultivars farmers’ cultivars of 4 years old ginsengs’ roots with both results showing that SPL transcripts formed a single network, which indicated that PgSPLs inter-coordinated when performing their functions. What’s more, we found that most PgSPL transcripts tended to express in older ginseng instead of younger ginseng, which was not only reflected in the expression of more types of SPL transcripts in older ginseng, but also in the higher expression of SPL transcripts in older ginseng. Additionally, we found that four PgSPL transcripts were only massively expressed in roots. According to PgSPL transcripts’ expression characteristics, we found that PgSPL23-35 and PgSPL24-09 were most proper two transcripts to further study as ginseng age’s molecular marker. These results provide the basis for further elucidation of the PgSPL transcripts’ biological function in ginseng and ginseng genetics improvement and gene breeding in the future.

Evolutionary Analyses Reveal Diverged Patterns of SQUAMOSA Promoter Binding Protein-Like (SPL) Gene Family in Oryza Genus.

The SPL (SQUAMOSA promoter binding protein-like) gene family is one of the plant-specific transcription factor families and controls a considerable number of biological functions, including floral development, phytohormone signaling, and toxin resistance. However, the evolutionary patterns and driving forces of SPL genes in the Oryza genus are still not well-characterized. In this study, we investigated a total of 105 SPL genes from six AA genome Oryza representative species (O. barthii, O. glumipatula, O. nivara, O. rufipogon, O. glaberrima, and O. sativa). Phylogenetic and motif analyses indicated that SPL proteins could be divided into two distinct lineages (I and II), and further studies showed lineage II consisted of three clades (IIA, IIB, and IIC). We found that clade I had comparable structural features with clade IIA, whereas genes in clade IIC displayed intrinsic differences, such as lower exon numbers and the presence of miR156 regulation elements. Nineteen orthologous groups of OsSPLs in Oryza were also identified, and most exons within those genes maintained constant length, whereas length of intron changed relatively. All groups were constrained by stronger purifying selection and diversified continually including alterative gene number, intron length, and miR156 regulation. Subsequently, cis-acting element analyses revealed the potential role of SPLs in wild rice, which might participate in light-responsive, phytohormone response, and plant growth and development. Our results shed light on that different evolutionary rates and duplication events might result in divergent evolutionary patterns in each lineage of SPL genes, providing a guide in exploring diverse function in the rice gene family among six closely related Oryza species.

Selection pressure causes differentiation of the SPL gene family in the Juglandaceae

The SQUAMOSA promoter-binding protein-like (SPL) is a plant-specific transcription factor that influences flowering and vegetative development. Although the SPL genes have been functionally analyzed in many species, studies on the evolutionary history of the whole gene family, and in the Juglandaceae specifically, have been limited. Here, we conducted a phylogenetic relationship analysis of the Juglandaceae SPL gene family compared with other land plant species. Our results showed that the SPL genes were divided into three major clades, all of which were further divided into ten small clades. Selection pressure analysis suggested that all SPL genes were exposed to purifying selection pressure during evolution. The purifying selection was smaller for the Juglandaceae SPL genes than for other angiosperms, indicating a greater susceptibility to functional differentiation in the Juglandaceae. The SPL proteins encoded by Clade 1 contained a branch-specific transmembrane structure and many conserved motif combinations at the C-terminal. We also detected many selection sites in these motif combinations. Expression analysis showed that Clade 1 genes had spatial and temporal differences and were highly expressed in various organs. The expression profile was closely related to the selection sites and motif combinations at the C-terminal. These observations represent essential entry points for revealing the functional differentiation of the SPL gene family. Our data presented here may provide a basis for future investigations of SPL genes in the Juglandaceae, especially for flower development and perhaps crop yield improvement.

Roles of the GA-mediated SPL Gene Family and miR156 in the Floral Development of Chinese Chestnut (Castanea mollissima).

Chestnut (Castanea mollissima) is a deciduous tree species with major economic and ecological value that is widely used in the study of floral development in woody plants due its monoecious and out-of-proportion characteristics. Squamosa promoter-binding protein-like (SPL) is a plant-specific transcription factor that plays an important role in floral development. In this study, a total of 18 SPL genes were identified in the chestnut genome, of which 10 SPL genes have complementary regions of CmmiR156. An analysis of the phylogenetic tree of the squamosa promoter-binding protein (SBP) domains of the SPL genes of Arabidopsis thaliana, Populus trichocarpa, and C. mollissima divided these SPL genes into eight groups. The evolutionary relationship between poplar and chestnut in the same group was similar. A structural analysis of the protein-coding regions (CDSs) showed that the domains have the main function of SBP domains and that other domains also play an important role in determining gene function. The expression patterns of CmmiR156 and CmSPLs in different floral organs of chestnut were analyzed by real-time quantitative PCR. Some CmSPLs with similar structural patterns showed similar expression patterns, indicating that the gene structures determine the synergy of the gene functions. The application of gibberellin (GA) and its inhibitor (Paclobutrazol, PP333) to chestnut trees revealed that these exert a significant effect on the number and length of the male and female chestnut flowers. GA treatment significantly increased CmmiR156 expression and thus significantly decreased the expression of its target gene, CmSPL6/CmSPL9/CmSPL16, during floral bud development. This finding indicates that GA might indirectly affect the expression of some of the SPL target genes through miR156. In addition, RNA ligase-mediated rapid amplification of the 5′ cDNA ends (RLM-RACE) experiments revealed that CmmiR156 cleaves CmSPL9 and CmSPL16 at the 10th and 12th bases of the complementary region. These results laid an important foundation for further study of the biological function of CmSPLs in the floral development of C. mollissima.

Genome-wide identification of the SPL gene family in Dichanthelium oligosanthes.

SQUAMOSA promoter-binding protein-like (SPL) transcription factors play vital roles in various plant physiological processes. Although, the identification of the SPL gene family has been done in C4 grass plants, including rice and maize, the same has not been characterized in the C3 grass species Dichanthelium oligosanthes. In this study, 14 SPL genes were identified in the genome of D. oligosanthes. Gene structure analysis of the identified DoSPLs revealed the similarity and redundancy in their exon/intron organizations. Sequence comparisons within the DoSPLs and along with rice SPLs revealed the putative paralogs and orthologs in D. oligosanthes SPL genes. Phylogenetic analysis clustered the DoSPLs into eight groups along with other plant SPLs. Identification of the conserved SBP motifs in all 14 DoSPLs suggested them to be putative SPLs. In addition, the prediction of sub-cellular localization and associated functions for DoSPLs further supported to be SPL genes. The outcome of this study can serve as a framework for the isolation and functional validation of SPL genes in D. oligosanthes

Identification and Functional Characterization of Squamosa Promoter Binding Protein-Like Gene TaSPL16 in Wheat (Triticum aestivum L.).

Wheat (Triticum aestivum L.) is one of the most important crops in the world. Squamosa promoter binding protein-like (SPL) proteins are plant-specific transcript factors and play critical roles in plant growth and development. The functions of many SPL gene family members were well characterized in Arabidopsis and rice, in contrast, research on wheat SPL genes is lagging behind. In this study, we cloned and characterized TaSPL16, an orthologous gene of rice OsSPL16, in wheat. Three TaSPL16 homoeologs are located on the short arms of chromosome 7A, 7B, and 7D, and share more than 96% sequence identity with each other. All the TaSPL16 homoeologs have three exons and two introns, with a miR156 binding site in their last exons. They encode putative proteins of 407, 409, and 414 amino acid residues, respectively. Subcellular localization showed TaSPL16 distribution in the cell nucleus, and transcription activity of TaSPL16 was validated in yeast. Analysis of the spatiotemporal expression profile showed that TaSPL16 is highly expressed in young developing panicles, lowly expressed in developing seeds and almost undetectable in vegetative tissues. Ectopic expression of TaSPL16 in Arabidopsis causes a delay in the emergence of vegetative leaves (3–4 days late), promotes early flowering (5–7 days early), increases organ size, and affects yield-related traits. These results demonstrated the regulatory roles of TaSPL16 in plant growth and development as well as seed yield. Our findings enrich the existing knowledge on SPL genes in wheat and provide valuable information for further investigating the effects of TaSPL16 on plant architecture and yield-related traits of wheat.