Gao et al. Plant Methods (2020) 16:70 https://doi.org/10.1186/s13007-020-00609-1 Plant Methods RESEARCH Open Access Phylogenomic synteny network analyses reveal ancestral transpositions of auxin response factor genes in plants Bei Gao1, Liuqiang Wang2, Melvin Oliver3, Moxian Chen4* and Jianhua Zhang1,5* Abstract Background: Auxin response factors (ARFs) have long been a research focus and represent a class of key regulators of plant growth and development. Integrated phylogenomic synteny network analyses were able to provide novel insights into the evolution of the ARF gene family. Results: Here, more than 3500 ARFs collected from plant genomes and transcriptomes covering major streptophyte lineages were used to reconstruct the broad-scale family phylogeny, where the early origin and diversifcation of ARF in charophytes was delineated. Based on the family phylogeny, we proposed a unifed six-group classifcation system for angiosperm ARFs. Phylogenomic synteny network analyses revealed the deeply conserved genomic syntenies within each of the six ARF groups and the interlocking syntenic relationships connecting distinct groups. Recurrent duplication events, such as those that occurred in seed plants, angiosperms, core eudicots and grasses contributed to the expansion of ARF genes which facilitated functional diversifcation. Ancestral transposition activities in important plant families, including crucifers, legumes and grasses, were unveiled by synteny network analyses. Ancestral gene duplications along with transpositions have profound evolutionary signifcance which may have accelerated the func- tional diversifcation process of paralogues. Conclusions: The broad-scale family phylogeny in combination with the state-of-art phylogenomic synteny net- work analyses not only allowed us to infer the evolutionary trajectory of ARF genes across distinct plant lineages, but also facilitated to generate a more robust classifcation regime for this transcription factor family. Our study provides insights into the evolution of ARFs which will enhance our current understanding of this important transcription fac- tor family. Keywords: Auxin, ARF, Transcription factor, Gene duplication, Genomic synteny Background gravitropism as reviewed previously [1, 2]. Completion Te plant hormone auxin, or indole-3-acetic acid, con- of the genomes of the moss Physcomitrella patens, [3, trols many physiological and developmental processes 4], the liverwort Marchantia polymorpha [5] and the in land plants including but not limited to organo- lycophyte Selaginella moellendorfi [6] revealed that genesis, tissue diferentiation, apical dominance, many core functional proteins required for auxin bio- synthesis, perception, and signaling were present in *Correspondence: [email protected]; [email protected] the early-diverging land plant lineages. Comprehen- 1 State Key Laboratory of Agrobiotechnology, The Chinese University of Hong Kong, Hong Kong, China sive evolutionary studies also suggested that the auxin 4 CAS Key Laboratory of Quantitative Engineering Biology, Shenzhen molecular regulatory network evolved at the latest Institute of Synthetic Biology, Shenzhen Institutes of Advanced in the common ancestor of embryophytes [7, 8] and Technology, Chinese Academy of Sciences, Shenzhen 518055, China Full list of author information is available at the end of the article the auxin response factor (ARF) genes evolved from © The Author(s) 2020. This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. 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A recent review dem- suggesting an earlier origin of ARF than that proposed by onstrated that with the exception of the PGP/ABCB Finet and colleagues [13]. auxin transporters, homologues of all the other core Compared to conventional gene family studies that components for hormonal control of physiology and focus on one or a limited number of species of inter- development by auxin could be identifed in P. patens est [25–27], phylogenetic studies on a broader scale [10]. Changes in auxin perception and signaling that that include multiple plant lineages were able to gener- occurred through evolution could have generated the ate more robust insights into the evolutionary process diversifcation of plant forms that occurred during the that gave rise to the modern assemblage of a target gene past ~ 474–515 million-year history of the land plants family [13, 28]. Te inclusion of genomic synteny data [11], which eventually led to the complex vegetative provides important information that impacts the deter- innovations that shape the modern terrestrial and mination of the evolutionary past of a gene family, espe- freshwater ecosystems [2]. cially when the gene family of interest evolved in parallel Auxin response factors, as core components in auxin with ancestral genome duplication events [29]. Te con- signaling, have long been a focus of plant signaling ventional genomic block alignment that connects ortho- research [12]. Te 23 ARFs identifed in the Arabidopsis logues, retained on genomic syntenic blocks, worked well thaliana genome were phylogenetically clustered into for a limited number of species [29, 30], but a network three subfamilies (Clades A, B and C) which were subse- approach was more efective when multiple genomes quently divided into seven groups (ARF9, ARF1, ARF2, were included in the synteny analyses [31, 32]. A com- ARF3/4, ARF6/8, ARF5/7 and ARF10/16/17), a classif- prehensive genomic synteny network can be constructed cation that was well supported by ARF genes from other using nodes to represent the target genes and associated angiosperms and representative non-fowering lineages adjacent genomic blocks and the network edges (con- [13]. Generally, ARF proteins can be functionally divided necting lines) to represent syntenic relationships [29, into transcriptional activators (ARF5-8 and 19 in A. thali- 32]. Te recently established phylogenomic synteny net- ana) and repressors (remaining ARFs in A. thaliana) with work methodology was able to integrate and summa- well-characterized functional domain architectures [13, rize genomic synteny relationships to uncover and place 14]. ARFs bind to the auxin response elements (AuxRE: genomic events (e.g. ancient tandem duplications, line- TGT CTC ) in the promoter region of downstream auxin- age-specifc transposition activities) into the evolutionary inducible genes [15] and function in combination with past of a target gene family [31, 32]. Aux/IAA repressors, which dimerize with ARF activa- In this study, we collected more than 3500 ARF mem- tors in an auxin-regulated manner [14, 16]. Unlike ARF bers to generate a comprehensive gene-family phylogeny activators, few reports have demonstrated that ARF with the aim of flling evolutionary gaps in the non-fow- repressors are able to interact with other ARF proteins ering plants and splitting the long branches present in the or Aux/IAA proteins [17]. Recent work revealed a newly current phylogeny [13]. We propose an updated model identifed mechanism whereby the IAA32 and IAA34 for the evolution of ARF family that covered the major transcriptional repressors are stabilized by the trans- streptophytic clades that was based on the six-group membrane kinase 1 (TMK1) at the concave side of the classifcation system we proposed for the ARF genes in apical hook of the kinase to regulate ARF gene expression angiosperms. Phylogenomic synteny network analyses of and ultimately inhibit growth [18]. angiosperm genomes revealed the deep positional con- In most of the well-established transcription fac- servation of ARF members within each of the six groups. tor annotation procedures, such as those implemented Detailed individual synteny network analyses together by the PlnTFDB [19], PlantTFDB [20], iTAK [21] and with phylogenetic reconstructions for the six ARF groups TAPScan [22], ARFs were identifed using two sig- revealed their distinctive evolutionary histories. Ances- nature domains: the B3 (PF02362) domain and the tral duplication events in angiosperms, and subsequent auxin-response (PF06507) domain, although some ARF WGDs in eudicots and monocots have contributed to proteins (e.g. ARF23 in A. thaliana) may be truncated the expansion of ARF members. Ancestral lineage-spe- and lack the auxin-response domain [13, 14, 23]. Finet cifc transpositions in important angiosperm families et al. [13] established a robust and comprehensive phy- such as cucifers, legumes and grasses were also unveiled. logenetic framework