Different evolutionary patterns of TIR1/AFBs and AUX/IAAs and their implications for the morphogenesis of land plants
BMC Plant Biology volume 23, Article number: 265 (2023)
The plant hormone auxin is widely involved in plant growth, development, and morphogenesis, and the TIR1/AFB and AUX/IAA proteins are closely linked to rapid auxin response and signal transmission. However, their evolutionary history, historical patterns of expansion and contraction, and changes in interaction relationships are still unknown.
Here, we analyzed the gene duplications, interactions, and expression patterns of TIR1/AFBs and AUX/IAAs to understand their underlying mechanisms of evolution. The ratios of TIR1/AFBs to AUX/IAAs range from 4:2 in Physcomitrium patens to 6:29 in Arabidopsis thaliana and 3:16 in Fragaria vesca. Whole-genome duplication (WGD) and tandem duplication have contributed to the expansion of the AUX/IAA gene family, but numerous TIR1/AFB gene duplicates were lost after WGD. We further analyzed the expression profiles of TIR1/AFBs and AUX/IAAs in different tissue parts of Physcomitrium patens, Selaginella moellendorffii, Arabidopsis thaliana and Fragaria vesca, and found that TIR1/AFBs and AUX/IAAs were highly expressed in all tissues in P. patens, S. moellendorffii. In A. thaliana and F. vesca, TIR1/AFBs maintained the same expression pattern as the ancient plants with high expression in all tissue parts, while AUX/IAAs appeared tissue-specific expression. In F. vesca, 11 AUX/IAAs interacted with TIR1/AFBs with different interaction strengths, and the functional specificity of AUX/IAAs was related to their ability to bind TIR1/AFBs, thus promoting the development of specific higher plant organs. Verification of the interactions among TIR1/AFBs and AUX/IAAs in Marchantia polymorpha and F. vesca also showed that the regulation of AUX/IAA members by TIR1/AFBs became more refined over the course of plant evolution.
Our results indicate that specific interactions and specific gene expression patterns both contributed to the functional diversification of TIR1/AFBs and AUX/IAAs.
Indole-3-acetic acid (IAA) is the most common and first discovered auxin in plants. Auxin is widely distributed in the plant kingdom and participates in almost all aspects of plant growth, development, and morphogenesis [1, 2]. Plants control the function of auxin mainly by coordinating auxin synthesis metabolism [3, 4], polar transport , and signal transduction . The classical auxin signaling pathway in plants refers to the TIR1/AFBs-mediated auxin regulation mechanism [7, 8]. TIR1 was the first auxin receptor protein to be identified [9, 10], and AFBs were subsequently discovered to also belong to this family [11,12,13]. The auxin receptor protein family is a subfamily of the F-Box family, whose members contain a highly conserved F-box domain and leucine repeat domain. The F-box domain of the TIR1/AFB family is an important component of the E3 ubiquitin ligase complex that participates in the degradation of AUX/IAA proteins [14, 15].
The AUX/IAA gene family is one of the key gene families involved in the rapid response to changes in auxin concentration [16, 17]. AUX/IAAs have a very short lifespan in plants, with a half-life ranging from 10 to 60 min that is determined by their domain II [18, 19]. At lower auxin levels, they bind to auxin response factor (ARF) to form a dimer that inhibits ARF and thus regulates auxin-induced gene expression. At high auxin levels, TIR1/AFB binds to AUX/IAA and degrades it through ubiquitination, thereby eliminating the inhibitory effect of AUX/IAA on ARF [20, 21]. The TIR1/AFB-AUX/IAA-ARF pathway thus describes the process of plant perception, transduction, and response to auxin signals. In addition, it was shown that the binding of auxin to TIR1/AFBs requires the co-involvement of AUX/IAAs . In Arabidopsis thaliana, there are six TIR1/AFBs and 29 AUX/IAAs, and there may from numerous coreceptor complexes for diversification of auxin functions . Also, in addition to the domain II of AUX/IAAs, different auxin concentration distributions in plants partially determine the assembly of TIR1/AFBs and AUX/IAAs [22, 23]. Thus, the SCFTIR1/AFB-based auxin mechanism is complex and diverse.
With the process of terrestrialization of plants, auxin has played an important role in the evolution of the original unicellular algae to the present morphologically diverse land plants. Auxin signaling genes are rare in algae and the components of signaling are severely missing . For example, TIR1/AFBs were not found in all algae, while AUX/IAAs and ARFs were only found in a small fraction of algae. Until the discovery of TIR1/AFBs in bryophytes represented the emergence of the classical auxin signaling pathway . The TIR1/AFB and AUX/IAA gene families have been extensively characterized in plants [24,25,26].However, TIR1/AFBs and AUX/IAAs exercise their functions by forming the SCF complex, and the evolutionary history of these two gene families cannot be fully understood by the analysis of a single gene family. In addition, previous studies identified auxin response proteins and their subdomains and precursors to reconstructed the origin and evolution of the auxin response system. It was shown that whole-genome duplication was the driving force behind the evolution and subfunctionalization of the TIR1/AFB and AUX/IAA gene families [27, 28]. However, the retention and loss of different branch members of TIR1/AFBs and AUX/IAAs after whole-genome duplication remains unknown.
In this study, TIR1/AFBs and AUX/IAAs were identified from 34 plants and algae, including members of the Rhodophyta, Chlorophyta, Charophyta, Bryophyta, ferns, gymnosperms, basal angiosperms, monocots, and dicots. We then reconstructed the phylogeny, synteny network, and duplication mechanisms of the TIR1/AFBs and AUX/IAAs. We compared the expression patterns of TIR1/AFBs and AUX/IAAs in different tissues of Physcomitrium patens, Selaginella moellendorffii, A. thaliana, and F. vesca, as well as their promoter cis-elements. According to our analysis, Fragaria vesca have a typical number of TIR1/AFB members and have remained undifferentiated during evolution. F. vesca was selected as a representative dicot. Bryophyte is the first taxa in which stable TIR1/AFB and AUX/IAA signaling pathways appear. Also, the smaller number of members in Marchantia polymorpha compared to other plants in the same group is more favorable for studying the evolutionary mode of action of ancient TIR1/AFB and AUX/IAA. In F. vesca and M. polymorpha, we explored the interactions and expression patterns between TIR1/AFBs and AUX/IAAs in detail. We found that expansion and subfunctionalization of the TIR1/AFB and AUX/IAA gene families had promoted functional specificity and the elaboration of regulatory networks.
Genome-wide identification of TIR1/AFB and AUX/IAA gene members in plants
To explore the evolutionary histories of the TIR1/AFB and AUX/IAA gene families, we first used BLAST to identify the gene family members from 34 plants and algae, including Rhodophyta, Chlorophyta, Charophyta, Bryophyta, ferns, gymnosperms, basal angiosperms, monocots, and dicots. We then used HMMER to check whether the sequences contained specific domains: the Transp_inhibit (PF18791) and F-box (PF18511) domains for TIR1/ARFs and the AUX_IAA (PF02309) domain for AUX/IAAs. After filtering, there were 142 TIR1/AFB and 546 AUX/IAA candidate genes in the 34 species (Fig. 1). The results showed that TIR1/AFB and AUX/IAA first appeared in bryophytes, and the two gene families had similar numbers of members in these early plants. Over the course of evolution, there was relatively little change in the number of TIR1/AFB genes, but the AUX/IAA gene family showed marked expansion in angiosperms (Fig. 1).
To further explore their evolutionary relationships, we constructed phylogenetic trees of TIR1/AFBs and AUX/IAAs by the maximum likelihood method. The TIR1/AFB tree consisted of three main branches and could be further divided into four groups (Fig. 2A). There were no monocotyledons in clade I, and clade II contained all lineages of seed plants. TIR1/AFB gene families expanded in ferns, and functional differentiation of TIR1/AFBs occurred in monocotyledons and dicotyledons (Fig. 2A, Fig. S1). For the AUX/IAA gene family, there were four ancient clades and twelve groups (Fig. 2B). Functional differentiation and rapid expansion of the AUX/IAA gene family occurred in the gymnosperms and basal angiosperms, respectively (Fig. 2B, Fig. S1). The AUX/IAA gene family appeared to have a more complicated evolutionary history than the TIR1/AFB family.
Phylogenomic and synteny network analyses of TIR1/AFBs and AUX/IAAs
To understand the obvious differences in the evolutionary histories of plant TIR1/AFBs and AUX/IAAs, we constructed a collinearity network based on the obtained TIR1/AFB and AUX/IAA phylogenetic trees and a collinearity analysis of 19 angiosperm species. Genome-level analysis divided the TIR1/AFB gene family into four clusters and the AUX/IAA gene family into eleven clusters (Fig. 3). In the TIR1/AFBs, cluster 1 was specific to dicots and basal angiosperms, indicating that this group was lost in monocots. Clusters 2 and 4 were conserved in angiosperms. Cluster 3 was a new group present only in monocots and dicots (Fig. 3). In the AUX/IAAs, clusters 3, 5, and 8 were specific to dicots, and only cluster 4 was specific to monocots, indicating that the AUX/IAAs of dicots were more diverse than those of monocots. The remaining clusters contained representatives from all the angiosperms (Fig. 3).
Based on phylogenetic collinear network analysis, we found 747 gene pairs and 3 tandem duplications among the TIR1/AFBs. These tandem duplications were observed only in Theobroma cacao, Prunus persica, and Brachypodium distachyon (Fig. 4A). We further counted the intron numbers of the TIR1/AFBs and calculated the Ks (synonymous substitution) values of collinear gene pairs. Four groups of TIR1/AFBs shared the same intron numbers and distribution of Ks values (Fig. S2, Fig. S4A), indicating that whole-genome duplication was the primary driving force for the evolution of the TIR1/AFB gene family. For AUX/IAAs, there were 1465 gene pairs and 59 tandem duplications. One tandem duplication occurred in group C, and the rest occurred in Groups I, J, K, and L (Fig. 4C). The Ks distribution of collinear gene pairs and tandemly duplicated gene pairs showed that the AUX/IAAs expanded through two WGD events and one tandem duplication event, and the tandem duplication event happened between the two WGD events (Fig. S4B). Moreover, the number of introns in groups K and L decreased after the tandem duplication event (Fig. S3). The numbers of introns in group E genes were also significantly reduced (Fig. S3). We therefore speculated that group E may have experienced tandem duplication independently after the second WGD event and the original branch was lost in subsequent evolution.
According to these results, we constructed a possible evolutionary history of TIR1/AFBs and AUX/IAAs. In the TIR1/AFB family, we hypothesized that there were already three TIR1/AFB members in the ancestors of seed plants, and the TIR1/AFB family expanded further through two WGD events (Fig. 4B). In the AUX/IAA family, there were four members in the common ancestor of seed plants. The four ancestral clades expanded through the two WGD events and two additional tandem duplication events in seed plants (Fig. 4D). Therefore, the evolution and expansion of the AUX/IAA gene family were driven by tandem duplication and WGD.
Functionally specialized AUX/IAAs show enhanced interaction ability with TIR1/AFBs
To investigate the functional differentiation of TIR1/AFB and AUX/IAA members in plants, we constructed heat maps showing their expression in different tissues of P. patens, S. moellendorffii, A. thaliana, and F. vesca. All TIR1/AFBs were highly expressed in different tissues of the four species (Fig. S5-S8), demonstrating that TIR1/AFB members were functionally conserved and extensively involved in plant morphogenesis. All AUX/IAAs were highly expressed in different tissues of P. patens and S. moellendorffii (Fig. S5, S6). These results suggest that when the TIR1/AFB-AUX/IAA signaling pathway appeared in ancient plants, it was likely to have been involved in all aspects of plant development. Some groups of AUX/IAAs maintained the same expression patterns in A. thaliana and F. vesca as in P. patens and S. moellendorffii (groups A, F, G, H, I, and L in A. thaliana and groups A, D, G, and H in F. vesca) (Fig. S5–S8). However, AUX/IAAs in the remainder of the groups appeared to have begun functional differentiation and were highly expressed in only one of the vegetative or reproductive organs; these included groups B and D of A. thaliana, focused on seed development, and group F of F. vesca, also focused on seed development (Fig. S7, S8). In addition, the functional differentiation of AUX/IAAs was more clearly in F. vesca than in A. thaliana.
In addition, clade IV appeared to have experienced the most complex duplication history (Fig. 4D). Therefore, we constructed the evolutionary trajectories of TIR1/AFBs and AUX/IAAs in F. vesca and M. polymorpha and verified their interaction patterns (Fig. 5, Fig. S12). The results showed that FvTIR1, FvAFB2, and FvAFB5 could interact with 7, 2, and 4 FvAUX/IAA members, respectively (Fig. 5, Fig. S10). Furthermore, we found that FvIAA14a strongly interacted with all FvTIR1/AFBs, and FvIAA14b and FvIAA6 strongly interacted with FvTIR1 and FvAFB5, respectively (Fig. 5, Fig. S11). Interestingly, AUX/IAAs in groups G and H showed little interaction with the TIR1/AFBs, even though they shared a common ancestor with groups I, J, K, and L (Fig. 5, Fig. S11). AUX/IAAs in groups G and H appeared to be involved in the development of all tissues. However, AUX/IAAs in groups I, J, K, and L (except FvIAA16) showed tissue-specific expression (Fig. S8). Therefore, we speculated that the enhanced binding ability of AUX/IAAs for TIR1/AFBs may have promoted the development of functional specificity. In clade IV, the functional specificity of AUX/IAAs in reproductive organs was related to their ability to interact with TIR1/AFBs. The binding ability of FvIAA3 and FvIAA4b with FvTIR1/AFBs was weakened after tandem duplication events, and their expression in reproductive organs also disappeared (Fig. 5, Fig. S8). In addition, we comprehensively analyzed the promoter of TIR1/AFB and AUX/IAA gene families in P. patens, S. moellendorfii, A. thaliana, and F. vesca. The results showed that a large number of hormone responsive elements (Abscisic acid, Gibberellin, Methyl jasmonate, Salicylic acid., Auxin) present in the promoter. Furthermore, cis-elements related to growth and development are also specifically present in different TIR1/AFB and AUX/IAA member promoters (Fig. S9). These results suggested that changes in interaction modes after duplication events and regulatory elements of the promoter may have helped to promote the specific development of higher plant organs, at least to some extent.
To further investigate these results, we confirmed the interaction patterns of TIR1/AFBs and AUX/IAAs between F. vesca and M. polymorpha. As shown in Fig. 5, MpTIR1/AFB interacted with eight FvAUX/IAAs, and MpAUX/IAA could interact with all TIR1/AFBs of F. vesca and M. polymorpha. This result showed that ancient TIR1/AFB and AUX/IAA had broad binding capacity but lacked the ability to be precisely regulated. Compared with M. polymorpha, strawberry displayed a more elaborate regulatory network formed by the subfunctionalization of genes that arose through the expansion of the TIR1/AFB and AUX/IAA gene families.
The terrestrialization of aquatic plants was an important event in plant evolution. However, the time at which the auxin signal appeared is still unknown TIR1/AFBs and AUX/IAAs have been found in a large number of land plants, ranging from lower plants such as mosses to higher plants. Thus, the TIR1/AFBs-AUX/IAAs signaling pathway is conserved in land plants [24, 30,31,32]. In this study, TIR1/AFBs was identified in many mosses and ferns, but no homologous genes were found in the surveyed algae. Although auxin signals and AUX/IAAs had already emerged in algae [16, 33, 34], the absence of classical auxin signaling pathway elements indicated that this pathway first appeared in bryophytes. It has been suggested that this phenomenon may have been caused by a massive loss of ubiquitinated components in algae .
Whole-genome duplication doubles the entire genome of the plant and is thus a major mechanism of gene family expansion. In addition, tandem duplication also promotes the expansion of gene families [35, 36]. However, a large number of gene copies are lost during the evolutionary process [37, 38], and the retained genes promote better adaptation of plant growth and development [39,40,41,42]. TIR1/AFBs have experienced two WGDs in land plants, but the number of TIR1/AFBs has not expanded substantially, and they are divided into only three different types . The AUX/IAAs not only experienced the same WGD events as the TIR1/AFBs but also experienced two large-scale tandem duplication events in angiosperms . Although the evolutionary patterns of TIR1/AFBs and AUX/IAAs have been explored previously, this paper describes their retention and loss after duplication event in more detail. Also, two tandem duplication events that promote AUX/IAAs expansion were identified. Large numbers of AUX/IAAs were retained after duplication. Thus, the number of AUX/IAAs is greater than the number of TIR1/AFBs in seed plants. In Bryophyta, the ratio of TIR1/AFBs and AUX/IAAs was 1 or more. However, the rapid expansion of AUX/IAAs altered the balance between TIR1/AFBs and AUX/IAAs.
Gene subfunctionalization is a key mechanism for duplicate gene retention, and the main phenomenon of subfunctionalization is a temporal and/or spatial differentiation of gene expression . The TIR1/AFBs were all highly expressed in multiple tissues of P. patens, S. moellendorffii, A. thaliana, and F. vesca. Thus, a large number of TIR1/AFB duplicates were not retained after WGDs. By contrast, from lower land plants to higher plants, duplicated AUX/IAA genes were retained because of gene expression divergence. The functional differentiation of AUX/IAAs was most obvious in F. vesca. Thus, subfunctionalization and duplicate retention explained the marked expansion of AUX/IAAs in higher plants.
The expansion of AUX/IAAs would result in AUX/IAAs competing with one another for binding to TIR1/AFBs. Therefore, the interaction relationships between TIR1/AFBs and AUX/IAAs might be retained, lost, or gained over evolutionary history. We focused on clade IV, which was the most complex branch in AUX/IAA evolutionary history, and we analyzed the interaction relationships among AUX/IAAs and TIR1/AFBs of F. vesca. We found that the binding ability between F. vesca TIR1/AFBs and AUX/IAAs was closely related to the functional specificity of the F. vesca AUX/IAAs. AUX/IAAs with stronger functional specificity were more stringently regulated by TIR1/AFBs. This was also demonstrated by the binding relationships among TIR1/AFBs and AUX/IAAs from M. polymorpha and F. vesca. In addition, the analysis of the strong and weak binding ability of TIR1/AFBs and AUX/IAAs had been similarly studied in previous studies. In A. thaliana, AtIAA7 had the strongest binding capacity to AtTIR1/AFBs. It can bind to AtTIR1 and AtAFB2 in the absence of auxin and to all AtTIR1/AFBs in the presence of trace amounts of auxin. In contrast, its tandem duplication gene AtIAA3 only bound to AtAFB1 in the presence of trace auxin and fails to bind to AtAFB5 at high auxin concentrations . In addition, the functions of AtIAA7 and AtIAA3 in plant development and root growth were similar [44,45,46,47,48,49,50,51,52,53,54,55], and AtIAA7 was also able to regulate flowering time in A. thaliana [56, 57] while the function of AtIAA3 in flower development had not been reported yet. This was identical to the expression profiles of AtIAA7 and AtIAA3 in different tissue parts of A. thaliana. The expression of IAA3 was significantly lower in reproductive organs than in nutritional organs. Thus, we hypothesize that the conserved functions of Arabidopsis and strawberry in Vegetative tissue was its origin from their ancestors, but as plants evolved, organ specificity led to the emergence of more refined regulation. This phenomenon could strongly promote accurate regulation of the development of diverse organs in higher plants by auxin.
In this study, TIR1/AFBs and AUX/IAAs were identified from 34 plants and algae, including members of the Rhodophyta, Chlorophyta, Charophyta, Bryophyta, ferns, gymnosperms, basal angiosperms, monocots, and dicots. We found that members of TIR1/AFBs and AUX/IAAs were incomplete in basal plants. Their appearance was associated with the terrestrialization of plants and appears intact for the first time in bryophytes. Meanwhile, whole genome duplication (WGD) and tandem duplication promoted the expansion of the AUX/IAA gene family, but many TIR1/AFB gene duplications were lost after WGD. In the expression profiles of different plants, TIR1/AFBs were found to be highly expressed in all tissue sites, whereas AUX/IAAs were highly expressed in all tissue sites in mosses and ferns, but tissue-specific expression was observed in higher plants. Also, further binding experiments showed that the functional specificity of AUX/IAAs was related to their ability to bind TIR1/AFBs. Our results indicate that specific interactions and specific gene expression patterns both contributed to the functional diversification of TIR1/AFBs and AUX/IAAs.
Identification of TIR1/AFBs and AUX/IAAs.
Thirty-four plant genome sequences were downloaded from Phytozome v13 (https://phytozome-next.jgi.doe.gov/) and other websites (Table S1). The 6 TIR1/AFBs and 29 AUX/IAAs of A. thaliana were used as query sequences, and BLASTP v2.10.0  was performed against the 33 other plant proteins with E-value < 1E − 10. We further confirmed the BLAST hits using HMMER v3.3.2 . TIR1/AFB proteins were required to contain the Transp_inhibit (PF18791) and F-box (PF18511) domains, and the AUX/IAA proteins were required to contain the AUX_IAA (PF02309) domain. Finally, we constructed evolutionary trees using FastTree v2.10.0  and manually filtered extremely long branches.
Reconstructed phylogenies of TIR1/AFBs and AUX/IAAs.
The protein sequences of the TIR1/AFBs and AUX/IAAs were aligned using MUSCLE v3.8.1551 , and phylogenetic trees were reconstructed using RAxML v8.2.12  with the GTRGAMMA model and 100 bootstrap replicates. Finally, we visualized the phylogenetic trees using the ITOL website .
Synteny, duplication, and gene-pair Ks analysis.
Collinearity networks were identified using the python version of MCScan (JCVI v1.1.7)  by comparing coding sequences to coding sequences. Then, the DupGen_finder pipeline  was performed to investigate potential duplication events. The Ks values of all gene pairs were calculated using KaKs_Calculator v2.0 .
Gene expression analysis.
The expression profile of P. patens was downloaded from the Physcomitrium eFP Browser (http://bar.utoronto.ca/efp_physcomitrella/cgi-bin/efpWeb.cgi). The expression profile of S. moellendorffii was downloaded from the Selaginella eFP Browser (http://bar.utoronto.ca/efp_selaginella/cgi-bin/efpWeb.cgi), and RNA sequencing (RNA-seq) data from spores were downloaded from NCBI (PRJNA326972). We obtained expression data for A. thaliana from the Arabidopsis eFP Browser (http://bar.utoronto.ca/efp/cgi-bin/efpWeb.cgi). Expression data for F. vesca were downloaded from the Strawberry eFP Browser (http://bar.utoronto.ca/efp_strawberry/cgi-bin/efpWeb.cgi), and RNA-seq data from old leaves, roots and shoots of F. vesca were downloaded from NCBI (PRJNA695578). We mapped all RNAseq data to the genome using HISAT2 v2.2.1  and SAMtools v1.7.1 . The genome version used was listed in Table S1. Then, the FPKM (Fragments Per Kilobase of exon model per Million mapped fragments) value were generated with Subread v2.0.1  and Trinity v2.13.2 . Finally, we visualized the expression data using TBtools .
Promoter cis-element analysis.
The promoter (2000 bp upstream) of TIR1/AFB and AUX/IAA gene families in P. patens, S. moellendorfii, A. thaliana, and F. vesca were extracted. Then, the cis-elements were predicted by online website PlantCARE (http://bioinformatics.psb.ugent.be/webtools/plantcare/html/). Finally, we visualized the cis-elements by ITOL website .
RNA extraction, complementary DNA preparation, and vector construction.
Total RNA was extracted from Fragaria vesca ‘Hawaii-4’ seedlings and M. polymorpha mature plant using a Plant Total RNA Isolation Kit (FOREGENE, Chengdu, China). The plant materials used in this experiment were obtained from the Fruit Tree Phylogenetic Laboratory of Nanjing Agricultural University. According to the instructions of the PrimeScript RT reagent kit (TaKaRa, Beijing, China), we synthesized the complementary DNA for gene cloning. Then, we cloned full-length TIR1/AFBs and AUX/IAAs using PrimeSTAR HS DNA Polymerase (TaKaRa, Beijing, China). Finally, the full-length genes were ligated into the pGADT7 and pGBKT7 vectors with the GenRec Assembly Master Mix Kit (GENERAL BIOL, Anhui, China). The primers used in this study are listed in Table S2 and Information of all amplified genes was listed in Table S3.
Yeast two-hybrid interaction assays.
Yeast two-hybrid interaction assays were performed to investigate the interactions of TIR1/AFBs with AUX/IAAs. Twelve AUX/IAA genes were ligated into the pGADT7 vector as prey, and four TIR1/AFB genes were ligated into the pGBKT7 vector as bait. Then, all constructed vectors and control vectors were transformed into the yeast strain Y2H Gold by the modified lithium acetate method. Finally, the transformed yeasts were cultured on SD-Leu-Trp medium, SD-Leu-Trp-His + X-α-Gal medium, and SD-Leu-Trp-His-Ade + X-α-Gal medium. For interactions, we classified them as interaction and strong interaction. Interactions: yeast could grow in SD-Leu-Trp-His + X-α-Gal medium and turn blue; strong interaction: yeast could grow in SD-Leu-Trp-His-Ade + X-α-Gal medium and turn blue. In addition, the interaction of TIR1/AFBs and AUX/IAAs may be influenced by auxin. Therefore, we added different concentrations of indole-3-acetic acid (0, 0.1, 1, 10 µM) to the SD-Leu-Trp-His + X-α-Gal medium, and SD-Leu-Trp-His-Ade + X-α-Gal medium.
The datasets analysed during the current study are available in the Physcomitrium eFP Browser (http://bar.utoronto.ca/efp_physcomitrella/cgi-bin/efpWeb.cgi), the Selaginella eFP Browser (http://bar.utoronto.ca/efp_selaginella/cgi-bin/efpWeb.cgi), the Arabidopsis eFP Browser (http://bar.utoronto.ca/efp/cgi-bin/efpWeb.cgi) and the Strawberry eFP Browser (http://bar.utoronto.ca/efp_strawberry/cgi-bin/efpWeb.cgi) repository. The raw RNA-seq data used in this study can obtained at NCBI website (PRJNA326972 and PRJNA695578).
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This research was supported in part by the Priority Academic Program Development of Jiangsu Higher Education Institutions and the State Key Laboratory of Crop Genetics and Germplasm Enhancement (ZW201813). This study was supported by the high-performance computing platform of Bioinformatics Center, Nanjing Agricultural University.
The authors declare that they have no competing interests.
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Su, L., Zhang, T., Yang, B. et al. Different evolutionary patterns of TIR1/AFBs and AUX/IAAs and their implications for the morphogenesis of land plants. BMC Plant Biol 23, 265 (2023). https://doi.org/10.1186/s12870-023-04253-4