Auxin Response Factor Family Research Articles

Towards a better understanding of auxin response factors for improving cereal crops

The auxin response factor (ARF) is the key component of phytohormone auxin signal pathway and is crucial in regulating a plurality of functions throughout plant life cycle. Although ARFs’ structure and function have been well studied in Arabidopsis, such knowledge is far from being sufficient for cereal crops, especially wheat, rice and maize. This review is based on a comprehensive retrospection into the studies on ARFs in the three cereal crops, consisting of four parts: (1) characterization of the domains of 23, 25 and 33 ARF family members in wheat, rice and maize, respectively; (2) revision of nomenclatures for previously reported ARFs to the family numbers based on sequence alignment, and summary of ARFs’ functions including the regulation of agronomic traits and response to biotic/abiotic stresses; (3) highlight of general regulatory models for fundamental physiological and reproductive traits from miRNA-ARFs, IAA-ARF-LBD, IAA-ARF-Auxin response gene, and IAA-ARF-ERF (4) prospects to promising future ARF research for anticipated agronomic traits. Hopefully, the review will facilitate the understanding of ARF functions in the three cereal crops and promote its application in molecular breeding for achieving optimal plant architecture, higher yield, and wide adaptability.

Identification of plant transcriptional activation domains.

Gene expression in Arabidopsis is regulated by more than 1,900 transcription factors (TFs), which have been identified genome-wide by the presence of well-conserved DNA-binding domains. Activator TFs contain activation domains (ADs) that recruit coactivator complexes; however, for nearly all Arabidopsis TFs, we lack knowledge about the presence, location and transcriptional strength of their ADs1. To address this gap, here we use a yeast library approach to experimentally identify Arabidopsis ADs on a proteome-wide scale, and find that more than half of the Arabidopsis TFs contain an AD. We annotate 1,553 ADs, the vast majority of which are, to our knowledge, previously unknown. Using the dataset generated, we develop a neural network to accurately predict ADs and to identify sequence features that are necessary to recruit coactivator complexes. We uncover six distinct combinations of sequence features that result in activation activity, providing a framework to interrogate the subfunctionalization of ADs. Furthermore, we identify ADs in the ancient AUXIN RESPONSE FACTOR family of TFs, revealing that AD positioning is conserved in distinct clades. Our findings provide a deep resource for understanding transcriptional activation, a framework for examining function in intrinsically disordered regions and a predictive model of ADs.

Genome wide characterization and expression insights of Auxin Response Factor (ARF) gene family in cabbage in response to clubroot disease

Auxin response factors (ARFs) regulate genes expression in response to auxin. Auxin performs a significant role in plant development and growth such as vascular tissue differentiation, root initiation and apical dominance. Within plants, auxin promotes the growth of vascular tissue differentiation, apical dominance and at the cellular level, auxin helps cells to divide and differentiate. Members of the ARF family carry a DNA binding domain (DBD) that binds to specific DNA sequences in the promoters of target genes. Despite the crucial role of the ARF gene family, no comprehensive study has been carried out to characterize the comparative ARF in B oleracea and A thaliana. For this reason, an in-silico approach was adopted. During the study, we found 49 genes in B oleracea and 37 genes in A thaliana. Of these, only 17 and 22 genes were studied due to their redundancy. In the phylogenetic analysis, we included B oleracea, B rapa, B napus and A thaliana and divided them into four groups based on conserved domains. Chromosomal mapping identified the positions of the genes on different chromosomes. The diversity of introns in the gene structure varies from 1 to 13 and up to five protein motifs are shown on each gene. Many transcription factors were identified by cis-regulatory elements. Gene expression revealed the involvement of BolARFs in root gall disease. The presence of nine different conserved domains was observed, of which B3, AUX_IAA superfamily together with auxin response were dominantly repeated in maximum number of genes. The overall study provides new insights and directions for further research and analysis in the future.

Unraveling the role of PlARF2 in regulating deed formancy in Paeonia lactiflora.

PlARF2 can positively regulate the seed dormancy in Paeonia lactiflora Pall. and bind the RY cis-element. Auxin, a significant phytohormone influencing seed dormancy, has been demonstrated to be regulated by auxin response factors (ARFs), key transcriptional modulators in the auxin signaling pathway. However, the role of this class of transcription factors (TFs) in perennials with complex seed dormancy mechanisms remains largely unexplored. Here, we cloned and characterized an ARF gene from Paeonia lactiflora, named PlARF2, which exhibited differential expression levels in the seeds during the process of seed dormancy release. The deduced amino acid sequence of PlARF2 had high homology with those of other plants and contained typical conserved Auxin_resp domain of the ARF family. Phylogenetic analysis revealed that PlARF2 was closely related to VvARF3 in Vitis vinifera. The subcellular localization and transcriptional activation assay showed that PlARF2 is a nuclear protein possessing transcriptional activation activity. The expression levels of dormancy-related genes in transgenic callus indicated that PlARF2 was positively correlated with the contents of PlABI3 and PlDOG1. The germination assay showed that PlARF2 promoted seed dormancy. Moreover, TF Centered Yeast one-hybrid assay (TF-Centered Y1H), electrophoretic mobility shift assay (EMSA) and dual-luciferase reporter assay analysis (Dual-Luciferase) provided evidence that PlARF2 can bind to the 'CATGCATG' motif. Collectively, our findings suggest that PlARF2, as TF, could be involved in the regulation of seed dormancy and may act as a repressor of germination.

Identification of ARF genes in Juglans Sigillata Dode and analysis of their expression patterns under drought stress.

Auxin response factor (ARF), a transcription factors that controls the expression of genes responsive to auxin, plays a key role in the regulation of plant growth and development. Analyses aimed at identifying ARF family genes and characterizing their functions in Juglans sigillata Dode are lacking. We used bioinformatic approaches to identify members of the J. sigillata ARF gene family and analyze their evolutionary relationships, collinearity, cis-acting elements, and tissue-specific expression patterns. The expression patterns of ARF gene family members under natural drought conditions were also analyzed. The J. sigillata ARF gene family contained 31 members, which were unevenly distributed across 16 chromosomes. We constructed a phylogenetic tree of JsARF genes and other plant ARF genes. Cis-acting elements in the promoters of JsARF were predicted. JsARF28 showed higher expressions in both the roots and leaves. A heat map of the transcriptome data of the cluster analysis under drought stress indicated that JsARF3/9/11/17/20/26 are responsive to drought. The expression of the 11 ARF genes varied under PEG treatment and JsARF18 and JsARF20 were significantly up-regulated. The interactions between abiotic stresses and plant hormones are supported by our cumulative data, which also offers a theoretical groundwork for comprehending the ARF mechanism and drought resistance in J. sigillata.

Large-scale analysis of the ARF and Aux/IAA gene families in 406 horticultural and other plants

The auxin response factor (ARF) and auxin/indole-3-acetic acid (Aux/IAA) family of genes are central components of the auxin signaling pathway and play essential roles in plant growth and development. Their large-scale analysis and evolutionary trajectory of origin are currently not known. Here, we identified the corresponding ARF and Aux/IAA family members and performed a large-scale analysis by scanning 406 plant genomes. The results showed that the ARF and Aux/IAA gene families originated from charophytes. The ARF family sequences were more conserved than the Aux/IAA family sequences. Dispersed duplications were the common expansion mode of ARF and Aux/IAA families in bryophytes, ferns, and gymnosperms; however, whole-genome duplication was the common expansion mode of the ARF and Aux/IAA families in basal angiosperms, magnoliids, monocots, and dicots. Expression and regulatory network analyses revealed that the Arabidopsis thaliana ARF and Aux/IAA families responded to multiple hormone, biotic, and abiotic stresses. The APETALA2 and serum response factor-transcription factor gene families were commonly enriched in the upstream and downstream genes of the ARF and Aux/IAA gene families. Our study provides a comprehensive overview of the evolutionary trajectories, structural functions, expansion mechanisms, expression patterns, and regulatory networks of these two gene families.

Identification of ARF transcription factor gene family and its defense responses to bacterial infection and salicylic acid treatment in sugarcane.

Auxin response factor (ARF) is a critical regulator in the auxin signaling pathway, involved in a variety of plant biological processes. Here, gene members of 24 SpapARFs and 39 SpnpARFs were identified in two genomes of Saccharum spontaneum clones AP85-441 and Np-X, respectively. Phylogenetic analysis showed that all ARF genes were clustered into four clades, which is identical to those ARF genes in maize (Zea mays) and sorghum (Sorghum bicolor). The gene structure and domain composition of this ARF family are conserved to a large degree across plant species. The SpapARF and SpnpARF genes were unevenly distributed on chromosomes 1-8 and 1-10 in the two genomes of AP85-441 and Np-X, respectively. Segmental duplication events may also contribute to this gene family expansion in S. spontaneum. The post-transcriptional regulation of ARF genes likely involves sugarcane against various stressors through a miRNA-medicated pathway. Expression levels of six representative ShARF genes were analyzed by qRT-PCR assays on two sugarcane cultivars [LCP85-384 (resistant to leaf scald) and ROC20 (susceptible to leaf scald)] triggered by Acidovorax avenae subsp. avenae (Aaa) and Xanthomonas albilineans (Xa) infections and salicylic acid (SA) treatment. ShARF04 functioned as a positive regulator under Xa and Aaa stress, whereas it was a negative regulator under SA treatment. ShARF07/17 genes played positive roles against both pathogenic bacteria and SA stresses. Additionally, ShARF22 was negatively modulated by Xa and Aaa stimuli in both cultivars, particularly LCP85-384. These findings imply that sugarcane ARFs exhibit functional redundancy and divergence against stressful conditions. This work lays the foundation for further research on ARF gene functions in sugarcane against diverse environmental stressors.

Genome-Wide Identification and Expression Analysis of Auxin Response Factor Gene Family in Linum usitatissimum.

Auxin response factors (ARFs) are critical components of the auxin signaling pathway, and are involved in diverse plant biological processes. However, ARF genes have not been investigated in flax (Linum usitatissimum L.), an important oilseed and fiber crop. In this study, we comprehensively analyzed the ARF gene family and identified 33 LuARF genes unevenly distributed on the 13 chromosomes of Longya-10, an oil-use flax variety. Detailed analysis revealed wide variation among the ARF family members and predicted nuclear localization for all proteins. Nineteen LuARFs contained a complete ARF structure, including DBD, MR, and CTD, whereas the other fourteen lacked the CTD. Phylogenetic analysis grouped the LuARFs into four (I-V) clades. Combined with sequence analysis, the LuARFs from the same clade showed structural conservation, implying functional redundancy. Duplication analysis identified twenty-seven whole-genome-duplicated LuARF genes and four tandem-duplicated LuARF genes. These duplicated gene pairs' Ka/Ks ratios suggested a strong purifying selection pressure on the LuARF genes. Collinearity analysis revealed that about half of the LuARF genes had homologs in other species, indicating a relatively conserved nature of the ARFs. The promoter analysis identified numerous hormone- and stress-related elements, and the qRT-PCR experiment revealed that all LuARF genes were responsive to phytohormone (IAA, GA3, and NAA) and stress (PEG, NaCl, cold, and heat) treatments. Finally, expression profiling of LuARF genes in different tissues by qRT-PCR indicated their specific functions in stem or capsule growth. Thus, our findings suggest the potential functions of LuARFs in flax growth and response to an exogenous stimulus, providing a basis for further functional studies on these genes.

Identification and transcriptome data analysis of ARF family genes in five Orchidaceae species

The Orchidaceae is a large family of perennial herbs especially noted for the exceptional diversity of specialized flowers. Elucidating the genetic regulation of flowering and seed development of orchids is an important research goal with potential utility in orchid breeding programs. Auxin Response Factor (ARF) genes encode auxin-responsive transcription factors, which are involved in the regulation of diverse morphogenetic processes, including flowering and seed development. However, limited information on the ARF gene family in the Orchidaceae is available. In this study, 112 ARF genes were identified in the genomes of 5 orchid species (Apostasia shenzhenica, Dendrobium catenatum, Phalaenopsis aphrodite, Phalaenopsis equestris and Vanilla planifolia,). These genes were grouped into 7 subfamilies based on their phylogenetic relationships. Compared with the ARF family in model plants, such as Arabidopsis thaliana and Oryza sativa, one group of ARF genes involved in pollen wall synthesis has been lost during evolution of the Orchidaceae. This loss corresponds with absence of the exine in the pollinia. Through mining of the published genomic and transcriptomic data for the 5 orchid species: the ARF genes of subfamily 4 may play an important role in flower formation and plant growth, whereas those of subfamily 3 are potentially involved in pollen wall development. the study results provide novel insights into the genetic regulation of unique morphogenetic phenomena of orchids, which lay a foundation for further analysis of the regulatory mechanisms and functions of sexual reproduction-related genes in orchids.

ARF19 Condensation in the Arabidopsis Stomatal Lineage.

The phytohormone auxin regulates nearly every aspect of plant development. Transcriptional responses to auxin are driven by the activities of the AUXIN RESPONSE FACTOR family of transcription factors. ARF19 (AT1G19220) is critical in the auxin signaling pathway and has previously been shown to undergo protein condensation to tune auxin responses in the root. However, ARF19 condensation dynamics in other organs has not yet been described. In the Arabidopsis stomatal lineage, we found that ARF19 cytoplasmic condensates are enriched in guard cells and pavement cells, terminally differentiated cells in the leaf epidermis. This result is consistent with previous studies showing ARF19 condensation in mature root tissues. Our data reveal that the sequestration of ARF19 into cytoplasmic condensation in differentiated leaf epidermal cells is similar to root-specific condensation patterns.

FveARF2 negatively regulates fruit ripening and quality in strawberry.

Auxin response factors (ARFs) are transcription factors that play important roles in plants. ARF2 is a member of the ARF family and participates in many plant growth and developmental processes. However, the role of ARF2 in strawberry fruit quality remains unclear. In this study, FveARF2 was isolated from the woodland strawberry 'Ruegen' using reverse transcription-polymerase chain reaction (RT-PCR), which showed that FveARF2 expression levels were higher in the stem than in other organs of the 'Ruegen' strawberry. Moreover, FaARF2 was higher in the white fruit stage of cultivated strawberry fruit than in other stage. Subcellular localization analysis showed that FveARF2 is located in the nucleus, while transcriptional activation assays showed that FveARF2 inhibited transcription in yeast. Silencing FveARF2 in cultivated strawberry fruit revealed earlier coloration and higher soluble solid, sugar, and anthocyanin content in the transgenic fruit than in the control fruit, overexpression of FveARF2 in strawberry fruit delayed ripening and lower soluble solid, sugar, and anthocyanin content compared to the control fruit. Gene expression analysis indicated that the transcription levels of the fruit ripening genes FaSUT1, FaOMT, and FaCHS increased in FveARF2-RNAi fruit and decreased in FveARF2-OE fruit, when compared with the control. Furthermore, yeast one-hybrid (Y1H) and GUS activity experiments showed that FveARF2 can directly bind to the AuxRE (TGTCTC) element in the FaSUT1, FaOMT, and FaCHS promoters in vitro and in vivo. Potassium ion supplementation improved the quality of strawberry fruit, while silencing FveARF2 increased potassium ion content in transgenic fruit. The Y1H and GUS activity experiments also confirmed that FveARF2 could directly bind to the promoter of FveKT12, a potassium transporter gene, and inhibited its expression. Taken together, we found that FveARF2 can negatively regulate strawberry fruit ripening and quality, which provides new insight for further study of the molecular mechanism of strawberry fruit ripening.

Genome-Wide Identification and Co-Expression Analysis of ARF and IAA Family Genes in Euscaphis konishii: Potential Regulators of Triterpenoids and Anthocyanin Biosynthesis.

Euscaphis konishii is an evergreen plant that is widely planted as an industrial crop in Southern China. It produces red fruits with abundant secondary metabolites, giving E. konishii high medicinal and ornamental value. Auxin signaling mediated by members of the AUXIN RESPONSE FACTOR (ARF) and auxin/indole-3-acetic acid (Aux/IAA) protein families plays important roles during plant growth and development. Aux/IAA and ARF genes have been described in many plants but have not yet been described in E. konishii. In this study, we identified 34 EkIAA and 29 EkARF proteins encoded by the E. konishii genome through database searching using HMMER. We also performed a bioinformatic characterization of EkIAA and EkARF genes, including their phylogenetic relationships, gene structures, chromosomal distribution, and cis-element analysis, as well as conserved motifs in the proteins. Our results suggest that EkIAA and EkARF genes have been relatively conserved over evolutionary history. Furthermore, we conducted expression and co-expression analyses of EkIAA and EkARF genes in leaves, branches, and fruits, which identified a subset of seven EkARF genes as potential regulators of triterpenoids and anthocyanin biosynthesis. RT-qPCR, yeast one-hybrid, and transient expression analyses showed that EkARF5.1 can directly interact with auxin response elements and regulate downstream gene expression. Our results may pave the way to elucidating the function of EkIAA and EkARF gene families in E. konishii, laying a foundation for further research on high-yielding industrial products and E. konishii breeding.

Genome-wide identification of auxin response factor (ARF) gene family and the miR160-ARF18-mediated response to salt stress in peanut (Arachis hypogaea L.)

Auxin response factors (ARFs) are transcription factors that regulate the transcription of auxin-responsive genes during plant growth and development. In this study, 29 and 30 ARF members were identified from the two wild peanut species, A. duranensis and A. ipaensis, respectively. The ARFs, including their classifications, conserved domains and evolutionary relationships were characterized. RNA-seq analyses revealed that some of the ARF genes were responsive to abiotic stress, particularly high salinity. In addition to abiotic stress, the expression of 2 ARF members was also regulated by biotic stress, specifically Bradyrhizobium infection in A. duranensis. The ARF gene Arahy.7DXUOK was predicted to be a potential target of miR160. Overexpression of miR160 could cause degradation of the Arahy.7DXUOK target gene transcript and increased salt tolerance in miR160OX transgenic plants. Therefore, these molecular characterization and expression profile analyses provide comprehensive information on ARF family members and will help to elucidate their functions to facilitate further research on peanuts.

Key auxin response factor (ARF) genes constraining wheat tillering of mutant dmc.

Tillering ability is a key agronomy trait for wheat (Triticum aestivum L.) production. Studies on a dwarf monoculm wheat mutant (dmc) showed that ARF11 played an important role in tillering of wheat. In this study, a total of 67 ARF family members were identified and clustered to two main classes with four subgroups based on their protein structures. The promoter regions of T. aestivum ARF (TaARF) genes contain a large number of cis-acting elements closely related to plant growth and development, and hormone response. The segmental duplication events occurred commonly and played a major role in the expansion of TaARFs. The gene collinearity degrees of the ARFs between wheat and other grasses, rice and maize, were significantly high. The evolution distances among TaARFs determine their expression profiles, such as homoeologous genes have similar expression profiles, like TaARF4-3A-1, TaARF4-3A-2 and their homoeologous genes. The expression profiles of TaARFs in various tissues or organs indicated TaARF3, TaARF4, TaARF9 and TaARF22 and their homoeologous genes played basic roles during wheat development. TaARF4, TaARF9, TaARF12, TaARF15, TaARF17, TaARF21, TaARF25 and their homoeologous genes probably played basic roles in tiller development. qRT-PCR analyses of 20 representative TaARF genes revealed that the abnormal expressions of TaARF11 and TaARF14 were major causes constraining the tillering of dmc. Indole-3-acetic acid (IAA) contents in dmc were significantly less than that in Guomai 301 at key tillering stages. Exogenous IAA application significantly promoted wheat tillering, and affected the transcriptions of TaARFs. These data suggested that TaARFs as well as IAA signaling were involved in controlling wheat tillering. This study provided valuable clues for functional characterization of ARFs in wheat.

Genome-Wide Identification of ARF Transcription Factor Gene Family and Their Expression Analysis in Sweet Potato.

Auxin response factors (ARFs) are a family of transcription factors that play an important role of auxin regulation through their binding with auxin response elements. ARF genes are represented by a large multigene family in plants; however, to our knowledge, the ARF gene family has not been well studied and characterized in sweet potatoes. In this study, a total of 25 ARF genes were identified in Ipomea trifida. The identified ItrARF genes’ conserved motifs, chromosomal locations, phylogenetic relationships, and their protein characteristics were systemically investigated using different bioinformatics tools. The expression patterns of ItfARF genes were analyzed within the storage roots and normal roots at an early stage of development. ItfARF16b and ItfARF16c were both highly expressed in the storage root, with minimal to no expression in the normal root. ItfARF6a and ItfARF10a exhibited higher expression in the normal root but not in the storage root. Subsequently, ItfARF1a, ItfARF2b, ItfARF3a, ItfARF6b, ItfARF8a, ItfARF8b, and ItfARF10b were expressed in both root types with moderate to high expression for each. All ten of these ARF genes and their prominent expression signify their importance within the development of each respective root type. This study provides comprehensive information regarding the ARF family in sweet potatoes, which will be useful for future research to discover further functional verification of these ItfARF genes.

Knockout of Auxin Response Factor SlARF4 Improves Tomato Resistance to Water Deficit.

Auxin response factors (ARFs) play important roles in various plant physiological processes; however, knowledge of the exact role of ARFs in plant responses to water deficit is limited. In this study, SlARF4, a member of the ARF family, was functionally characterized under water deficit. Real-time fluorescence quantitative polymerase chain reaction (PCR) and β-glucuronidase (GUS) staining showed that water deficit and abscisic acid (ABA) treatment reduced the expression of SlARF4. SlARF4 was expressed in the vascular bundles and guard cells of tomato stomata. Loss of function of SlARF4 (arf4) by using Clustered Regularly Interspaced Short Palindromic Repeats/Cas 9 (CRISPR/Cas 9) technology enhanced plant resistance to water stress and rehydration ability. The arf4 mutant plants exhibited curly leaves and a thick stem. Malondialdehyde content was significantly lower in arf4 mutants than in wildtype plants under water stress; furthermore, arf4 mutants showed higher content of antioxidant substances, superoxide dismutase, actual photochemical efficiency of photosystem II (PSII), and catalase activities. Stomatal and vascular bundle morphology was changed in arf4 mutants. We identified 628 differentially expressed genes specifically expressed under water deficit in arf4 mutants; six of these genes, including ABA signaling pathway-related genes, were differentially expressed between the wildtype and arf4 mutants under water deficit and unlimited water supply. Auxin responsive element (AuxRE) elements were found in these genes’ promoters indicating that SlARF4 participates in ABA signaling pathways by regulating the expression of SlABI5/ABF and SCL3, thereby influencing stomatal morphology and vascular bundle development and ultimately improving plant resistance to water deficit.

Molecular Evolution and Local Root Heterogeneous Expression of the Chenopodium quinoa ARF Genes Provide Insights into the Adaptive Domestication of Crops in Complex Environments.

Auxin response factors (ARFs) influence plant growth and development via the coupling of basic biological processes. However, the evolution, expansion, and regulatory mechanisms of ARFs in the domesticated crop quinoa after artificial selection remain elusive. In this study, we systematically identified 30 Chenopodium quinoa ARFs (CqARFs). In this typical domesticated crop, ARFs divided into three subfamilies are subjected to strong purification selection and have a highly conserved evolutionary pattern. Polyploidy is the primary reason for the expansion of the ARF family after quinoa domestication. The expression patterns of CqARFs in different tissues have been differentiated, and CqARF2, 5, 9 and 10 from class A have the characteristics of local heterogeneous expression in different regions of roots, which may be the key factors for crops to respond in complex environments. Overall, we examined the evolution and expansion of ARFs in representative domesticated crops using the genome, transcriptome, and molecular biology and discovered a class A ARF-centered heterogeneous expression network that played an important role in auxin signaling and environmental responses. We provide new insights into how ARFs promote domesticated crop adaptation to artificial selection by polyploid expansion.

Genome-Wide Identification Analysis of the Auxin Response Factors Family in Nicotiana tabacum and the function of NtARF10 in Leaf Size Regulation

Auxin is well recognized for its involvement in several developmental processes like floral and leaf development and shoot elongation. The transcriptional regulation of auxin-responsive genes is mediated via auxin response factors (ARF). In this study, we identified 46 ARF genes in Nicotiana tabacum, performed phylogenetic analysis and investigated their structure, conserved domains, and motifs. Our results demonstrate that some of NtARF genes are regulated by mi-RNAs and expression in multiple tobacco tissues. Additionally, the leaf NtARFs display a diverse expression pattern in vein and shoot apical meristem in response to exogenous auxin stimulus. Transgenic NtARF10-overexpressing Arabidopsis plants exhibit larger leave areas, cell area, and more numerous cell numbers compared to wild-type plants and include several upregulated genes involved in cell division and expansion, including AtCYCD3, AtTCP1, AtTCP20, AtXTH33, and AtARGOS. This suggests NtARF10 might play a role in the regulation of leaf size. Our study contributes to a better understanding of the characteristics of the ARF family in tobacco and provides a basis for further functional research into NtARFs.

Genome-Wide Identification and Comparative Analysis of ARF Family Genes in Three Apiaceae Species.

The family Apiaceae includes many important vegetables and medicinal plants. Auxin response factors (ARFs) play critical roles in regulating plant growth and development. Here, we performed a comprehensive analysis of the ARF gene family in three Apiaceae species, celery, coriander, and carrot, and compared the results with the ARF gene family of lettuce, Arabidopsis, and grape. We identified 156 ARF genes in all six species and 89 genes in the three Apiaceae species, including 28, 34, and 27 in celery, coriander, and carrot, respectively. The paralogous gene number in coriander was far greater than that in carrot and celery. Our analysis revealed that ARF genes of the three Apiaceae species in 34 branches of the phylogenetic tree underwent significant positive selection. Additionally, our findings indicated that whole-genome duplication played an important role in ARF gene family expansion. Coriander contained a greater number of ARF genes than celery and carrot because of more gene duplications and less gene losses. We also analyzed the expression of ARF genes in three tissues by RNA-seq and verified the results by quantitative real-time PCR. Furthermore, we found that several paralogous genes exhibited divergent expression patterns. Overall, this study provides a valuable resource for exploring how ARF family genes regulate plant growth and development in other plants. Since this is the first report of the ARF gene family in Apiaceae, our results will serve as a guide for comparative and functional analyses of ARF and other gene families in Apiaceae.

Genome-wide analysis of auxin response factor (ARF) genes and functional identification of MdARF2 reveals the involvement in the regulation of anthocyanin accumulation in apple

ABSTRACT Auxin response factors (ARFs) mediate auxin response gene transcription by directly binding with AuxREs. However, whether ARF family genes involved in anthocyanin accumulation in apple is largely unexplored. In this study, thirty-three MdARF family members were identified in the apple (Malus × domestica) genome (HFTH1 Whole Genome v1.0) based on the sequence similarity with ARF family members of Arabidopsis. Phylogenetic analysis revealed that thirty-three MdARF members were clustered into Class I, Class II and Class III. Functional domain analysis showed that MdARF proteins have some typical domains such as B3-like DNA-binding domain and Auxin_resp domain, as well as motif III and motif IV. Chromosome localisation analysis demonstrated that apple ARF members were widely distributed in the genome. Among these MdARF genes, MdARF2 was significantly correlated with anthocyanin variation according to extracting expression profiles from published RNA-seq data. Further functional identification in apple calli suggested that MdARF2 functions as a transcriptional repressor of anthocyanin biosynthesis. These findings provide evidence that MdARF family members play a role in the regulation of anthocyanin accumulation in apple.