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The evolution of auxin transport and its contribution to plant morphology
2024 年 36 巻 1 号 p. 5-19
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2024 年 36 巻 1 号 p. 5-19
Directional transport of the phytohormone auxin causes differential auxin distribution within tissues. This gradient is essential for various aspects of plant morphogenesis, such as embryogenesis, tissue patterning, and tropism. The auxin efflux carrier PIN FORMED (PIN) is responsible for such transport by being localized to a specific side of the plasma membrane. PIN exists in organisms throughout the Streptophyta, and polar auxin transport is detected in the Charophyceae algae Chara braunii. This suggests that the importance of auxin transport is conserved among land plants and charophytes. However, the morphology of land plants and charophytes is highly diversified, from complex angiosperms to simple algae without any differentiated cells. Therefore, the role of PIN and auxin transport may be different between these plants. Here, we review the current findings on the function of PIN and auxin transport reported in Charophyte, Bryophyte, and Fern. Based on these reports, we introduce three different types of auxin transport, “cell-to-environment auxin transport”, “non-canalized intercellular auxin transport”, and “canalized auxin transport”. Each of these is assumed to contribute to plant development differently. Finally, we discuss how such types of auxin transport are involved in plant morphogenesis and their relation to plant evolution.
Since the terrestrialization of plants in the Ordovician period, land plants have evolved a complex and diversified morphology. Such morphogenesis requires coordinated cell growth and elongation (Yoshida et al. 2019), regulated by the plant hormone auxin. Auxin is transported with a specific vector between cells, causing a directional auxin flow within tissues called polar auxin transport. Thus, a gradient of auxin level is formed among cells, leading to cell and tissue differentiation, such as embryogenesis, leaf vasculature patterning, lateral root formation, and shoot branching, or causing differential cell elongation for tropic responses (Peer et al. 2011).
The auxin influx into the cell is run by both passive diffusion and proton symporters. On the other hand, due to the pH gradient between cytosol and the cell wall, auxin efflux relies on the efflux carriers, including ABCB transporters and PIN FORMED (PIN) transporters (Vanneste and Friml 2009). Among these, the auxin efflux carrier PIN is considered the key regulator of auxin transport and the resulting auxin gradient formation within tissues (Vanneste and Friml 2009). PIN proteins are separated into two subfamilies, long PINs or short PINs, depending on the length of the central hydrophilic loop. Among them, the long PINs, or canonical PINs, are localized to the plasma membrane in a polar manner, determining the direction of auxin efflux and causing polar auxin transport (Figure 1A, Křeček et al. 2009). The establishment and reorienting of the PIN localization in different manners, depending on the cell, constructs an organized canal of auxin transport. Thus, the tissues develop in a determined structure or exhibit tropic growth in response to environmental stimuli (Vanneste and Friml 2009, Hajný et al. 2022).
Various proteins are identified as the regulator of auxin transport in Arabidopsis thaliana (Naramoto 2017). For example, membrane trafficking regulators GNOM ARF-GEF and VAN3 ARF-GAP mediate the polar localization of PIN proteins by controlling the endocytic recycling of PIN proteins (Geldner et al. 2003, Scarpella et al. 2006, Naramoto et al. 2009, 2010, Naramoto and Kyozuka 2018). Members of AGC protein kinase, PINOID (PID)/WAG1, WAG2, and D6 PROTEIN KINASE (D6PK) directly phosphorylate PIN and positively regulate its activity (Zourelidou et al. 2014). MAB4, an NPH3 (NON-PHOTOTROPIC HYPOCOTYL 3)-like protein, forms a complex at the plasma membrane with PIN, PID and D6PK, where MAB4 stabilizes the polar localization of PIN by limiting the endocytosis and lateral diffusion of PIN proteins (Furutani et al. 2011, Glanc et al. 2021). 14-3-3 interacts with NPH3 in a blue light-dependent manner and, together with D6PK, is predicted to regulate PIN activity and contribute to phototropism (Keicher et al. 2017, Reuter et al. 2021). PRAF/RLD is known to interact with GNOM physically, controlling the PIN localization to establish body axis and cell polarity (Wang et al. 2022). LZY (LAZY-like) interacts with PRAF/RLD and, through the translocation of LZY from the amyloplasts to the plasma membrane by gravistimulation and the resulting amyloplast sedimentation, changes the localization of PIN and contributes to gravitropism (Yoshihara et al. 2013, Furutani et al. 2020, Nishimura et al. 2023). Studies of genetic analysis report that these genes are mostly conserved among all land plants, even in bryophytes moss Physcomitrium patens (synonym: Physcomitrella patens) and liverwort Marchantia polymorpha (Table 1, Suetsugu et al. 2016). Therefore, it is suggested that the molecular mechanisms to establish polar auxin transport were mainly acquired at the common ancestor of land plants (Naramoto et al. 2022). As charophytes such as Chara and Klebsormidium also possess PIN and some of the regulators (Table 1), and directional auxin transport is observed in Chara as well as in bryophytes (Figure 1), the acquisition may have been even earlier (Cooke et al. 2002, Boot et al. 2012). However, the morphology of these plants is highly diversified and different from angiosperms, suggesting that the function of PIN and its regulation may be varied. In fact, non-polar PIN localization is observed in bryophytes P. patens and M. polymorpha (Viaene et al. 2014, Fisher et al. 2023), implying auxin transportation without clear directionality. Therefore, to understand the evolutionary process of PIN and auxin transport and their contribution to the morphological diversification of plants, it would be essential to understand their function in different organisms and how it has changed among Streptophyta.
Figure 1 Polar auxin transport is detected in Charales, Moss, Liverwort, and Ferns.
(A) The polarly-localized PIN protein defines the direction of auxin transport, thereby inducing a unidirectional flow of auxin (polar auxin transport). (B-E) Polar auxin transport is detected directly by the observation of radioactive 14C-IAA or 3H-IAA in Charales, Moss, Liverwort, and Ferns. Schematic illustrations of the direction of auxin transport detected (blue arrow) in the internodal cells of charales Chara corallina (B), the sporophyte of moss Polytrichum ohioense (C), the thallus of liverwort Marchantia polymorpha (D), and the rachis of fern Regnellidium diphyllum are shown (Marovolo et al. 1976, Walters and Osborne 1979, Gaal et al. 1982, Fujita et al. 2008, Boot et al. 2012) .
To address this question, we will summarize the current findings on the function of PIN proteins and auxin transport in charophytes, bryophytes, and ferns. However, due to the lack of transformation techniques, genetic studies among these organisms regarding the localization of PIN proteins, the phenotypes of pin knockout mutants, and the relation to auxin biosynthesis and signaling genes, are mostly limited to moss P. patens and liverwort M. polymorpha. Therefore, we will also refer to pharmacological studies, such as the effects of exogenous auxin and auxin transport inhibitors reported in charophytes, bryophytes, and ferns. Based on these knowledges, we will discuss how the role of PIN proteins and auxin transport evolved in streptophytes and how it has affected plant morphology.
Table 1 The homologous proteins related to PIN or auxin in Klebsormidium nitens, Chara braunii, Physcomitrium patens, Marchantia polymorpha, Ceratopteris richardii, and Arabidopsis thaliana.
Protein homologs were searched on NCBI ( https://www.ncbi.nlm.nih.gov/) by BLAST, using the amino acid sequence of each protein in A. thaliana as the query. Among the proteins identified as homologous, proteins that have their function analyzed are written as “Functional” , and others are written as “Exist” . Both K. nitens and C. braunii possessed an NPH3 family protein homolog; however, these proteins clustered to neither land plant NPH3 nor MAB4 through phylogenetic analysis. Therefore, we referred to these proteins as “Undistinguishable” . Nishiyama et al. (2018) addressed the Aux/IAA in K. nitens and C. braunii as “non-canonical” , and we followed that description.
Charophyte algae is the group of algae most closely related to land plants (embryophytes). As a paraphyletic group, the morphology shows a wide variety; while species in the genus Chara form complex bodies with differentiated cells, there are single cellular organisms such as the Closterium (Nishiyama et al. 2018, Tsuchikane and Sekimoto 2019). Among charophytes, there are several studies relevant to auxin transport in Klebsormidiophyceae and Charophyceae. As the organisms in these phyla currently lack transformation procedures, these studies have focused on the pharmacological analysis, such as the exogenous application of auxin or auxin transport inhibitors, including NPA (N-1-naphthylphthalamic acid) and TIBA (2,3,5-triiodobenzoic acid) (Dibb-Fuller and Morris 1992, Klämbt et al. 1992, Ohtaka et al. 2017). Moreover, they conducted biochemical measurements of auxin transport activity using H3 or C14 labeled IAA (Indole-3-acetic acid) or examined the effect of ectopically expressing PIN in other organisms (Boot et al. 2012, Ohtaka et al. 2017, Skokan et al. 2019).
Klebsormidium nitens is a multicellular alga with a non-branching filamentous body without any specialized or differentiated cells, which possesses a single PIN protein (KfPIN) (Hori et al. 2014). Ectopic expression in P. patens, A. thaliana, Nicotiana tabacum BY-2 cells, and Xenopus laevis oocytes showed that KfPIN is localized to the plasma membrane and is functional as auxin efflux carriers in each organism. The difference from land plant PINs was that KfPIN is not localized to the region connected to neighboring cells (cross wall) but to the regions facing the outer environment. When ectopically expressed in other plant organisms, KfPIN localization showed no clear polarity within the plasma membrane (Skokan et al. 2019). Treatment with highly concentrated exogenous auxin IAA, IBA (Indole-3-butyric acid) or NAA (1-Naphthaleneacetic acid), or the auxin transport inhibitor TIBA, but not NPA delays K. nitens growth and cell division (Ohtaka et al. 2017). These observations suggest that the role of KfPIN is to transport auxin out to the environment, not between cells, to maintain the intracellular level of auxin (Ohtaka et al. 2017, Skokan et al. 2019). Analysis of the genome sequence has identified that K. nitens possess auxin biosynthetic genes such as TAA (TRIPTOPHANE AMINO TRANSFERASE OF ARABIDOPSIS) and YUC (YUCCA) but lacks the TIR1/AFB - Aux/IAA - ARF auxin signaling pathway commonly used in land plants (Table 1, Hori et al. 2014, Ohtaka et al. 2017). K. nitens growth is suppressed by the application of auxin biosynthesis inhibitors BBo (4-biphenylboronic acid) and PPBo (4-phenoxyphenylboronic acid), indicating that auxin biosynthesis and transportation are both essential for K. nitens development (Ohtaka et al. 2017). As an auxin receptor, ABP1 (AUXIN BINDING PROTEIN 1) and ABL (ABP1-like) is present in the K. nitens genome, which can bind to auxin at the apoplast and regulate a non-transcriptional auxin signaling pathway through protein phosphorylation in angiosperms (Table 1, Hori et al. 2014, Friml et al. 2022, Yu et al. 2023). Although whether the ABP1 and ABL of K. nitens works as an auxin receptor is unclarified, IAA induces the phosphorylation of RAF-like kinase in K. nitens as well as in land plants (Kuhn et al. 2024). Therefore, the non-transcriptional auxin signaling pathway seems to be conserved among streptophytes, and the results of pharmacological analysis on K. nitens may be related to this pathway. Other than the non-transcriptional auxin signaling pathway, Ohtaka et al. (2017) reported that IAA treatment induces the expression of KnLBD (LATERAL ORGAN BOUNDARIES-DOMAIN). This implies that auxin can regulate gene expression in K. nitens, but as the TIR1/AFB-Aux/IAA-ARF auxin signaling is absent in K. nitens, how such regulation is caused remains unclear (Ohtaka et al. 2017).
In contrast to Klebsormidium, Chara develops a complex morphology with several differentiated tissues: the stem-like axis and the whorl of branchlets formed from the node comprises the thallus, and rhizoids are formed at the base of the thallus (Nishiyama et al. 2018). Among these tissues, basipetal polar auxin transport is observed in the internodal cells of C. corallina (Figure 1B, Boot et al. 2012). As a pharmacological analysis, Klämbt et al. (1992) reported that auxin transport inhibitor NPA suppressed the rhizoid growth in C. globularis. However, the effect of NPA on IAA accumulation in the cell varies among reports. Dibb-Fuller and Morris (1992) reported that the IAA efflux of C. vulgaris thallus cells was unaffected by either NPA or TIBA treatment. In contrast, Klämbt et al. (1992) reported that NPA treatment led to auxin accumulation in rhizoids of C. globularis. The growth inhibition of rhizoids was recovered by applying IAA (Klämbt et al. 1992), suggesting that NPA had additional effects on Chara rhizoids other than intracellular IAA accumulation (Cooke et al. 2002). Genome sequence analysis has suggested the existence of an intracellular auxin receptor and ARF; however, only a non-canonical Aux/IAA is found in the C. braunii genome (Table 1, Nishiyama et al. 2018). Thus, how Chara operates auxin signaling and the role of auxin transport remains unclear. Still, the land plant-like basipetal auxin transport and its complex morphology may have some correlation (Figure 1B, Vosolsobě et al. 2020), and further investigations are awaited.
Among charophytes, Zygnematophyceae, the phyla of unicellular or filamentous organisms without branching, are identified as the sister clade of land plants rather than the morphologically more complex Charales (Wickett et al. 2014). PIN proteins are present in these organisms, and stable transformation procedures by using particle bombardments are established in the unicellular Closterium despite the lower transformation efficiency (Abe et al. 2011). Recently, a new transformation technique employing electroporation has been established, which shows higher transformation efficiency that is amenable to experimental investigation (Kawai et al. 2022). Future genetic studies on Closterium PIN may give new insights into the evolution of auxin transport and its role in the development of charophytes (Bennett 2015).
Bryophytes are comprised of three groups, moss, liverwort, and hornwort, and are considered monophyletic, a sister clade to vascular plants (Donoghue et al. 2021, Harris et al. 2022). Among the three, liverwort and moss are also called the Setaphyta clade, named from their morphology of sporophytes, and hornworts are believed to be the first to diverge from the other two (Donoghue et al. 2021). Importantly, although bryophytes are often referred to as “ancestral” land plants, they may possess systems or phenomena gained after the divergence from vascular plants regarding the phylogenetic relation. Therefore, the function of PIN and auxin transport shared among bryophytes may not always be “ancestral” . To reveal the evolutional process of such functions and mechanisms, a comparison among bryophytes, vascular plants, and charophytes would be necessary.
As a study covering all three groups of bryophytes, Poli et al. (2003) investigated the existence of polar auxin transport in sporophytes of moss Polytrichum ohioense, liverwort Pellia epiphylla, and hornwort Phaeoceros pearsonii, by observing the movement of radioactive 3H-IAA. Among them, only the young sporophyte of moss P. ohioense showed evidence of polar auxin transport, which was in a basipetal direction (Figure 1C). There are currently no other reports on auxin transport in hornworts. However, in the gametophyte stage of liverwort M. polymorpha, polar auxin transport from the apical to the basal side of the thallus is detected (Figure 1D, Maravolo 1976, Gaal et al. 1982). The difference in the developmental stage where polar auxin transport is detected (that is, sporophyte generation or gametophyte generation) implies that, through the evolution of moss and liverwort, they may have newly gained or lost the ability to transport auxin in a polar manner within their tissues. Thus, liverworts and mosses are suitable subjects to study how the role of auxin transport has evolved in plants, as the different features may reflect the divergence in the role of auxin transport. To address this question, we will examine the currently elucidated function of PINs and auxin in each developmental stage of moss P. patens and liverwort M. polymorpha, as representative model organisms of each phylum. As canonical PINs, P. patens possesses three, called PpPIN A, B, and C (Viaene et al. 2014). M. polymorpha possesses a single canonical PIN, named MpPIN1 or MpPINZ (Fisher et al. 2023, Tang et al. 2023).
The life cycle of P. patens can be divided into three stages: the filamentous gametophyte protonema, the three-dimensional leafy shoot gametophore, and the sporophyte comprised of a short seta and a single spore capsule (Prigge and Bezanilla 2010). Among these tissues, basipetal auxin transport has been detected in the sporophyte of several species of mosses (Figure 1C, Poli et al. 2003, Fujita et al. 2008). In contrast, the gametophore of P. patens and several other mosses transport roughly the same amount of auxin in both basipetal and acropetal directions (Fujita et al. 2008). Still, the GmGH3:GUS assay reported an auxin gradient within the shoot, where the expression of GUS is limited to the stem of shoots (Bennett et al. 2014). In the leaves, observation of the R2D2 reporter line has identified that auxin distribution moves towards the basal side as leaves develop (Thelander et al. 2019). The protonema cells also exhibit an auxin gradient (Bierfreund et al. 2003, Thelander et al. 2019); therefore, although it may not be through polar auxin transport, the regulation of auxin gradient exists in all stages of the P. patens life cycle.
Upon spore germination or regeneration from injured tissues, P. patens produces protonema, a two-dimensional network-like structure formed from filamentous tissue with side branches. Protonemata first emerge as chloronema cells, containing fully developed chloroplasts in short cells with a perpendicular cell division plane. As the chloronemata elongates by cell division, the transition from chloronema to caulonema cells occurs at the apical tip, which contains fewer and smaller chloroplasts in long cells with an oblique cell division plane (Menand et al. 2007). The pina pinb double knockout mutant shows an earlier transition from chloronema to caulonema, whereas the overexpression of PINA prolongs the chloronema cell stage (Viaene et al. 2014). Treatment of exogenous auxin induces similar phenotypes as the pin knockout mutants (Jenkins and Cove 1983, Decker et al. 2006), suggesting that the phenotype of knockout mutant was caused by the accumulation of auxin, whereas that of overexpression mutants was caused by auxin starvation within the cells (Viaene et al. 2014). Thus, PIN proteins can be proposed as a regulator of the cell fate transition in protonema, where they maintain chloronema by auxin efflux (Viaene et al. 2014).
A concern following such phenomena would be the direction of PIN-mediated auxin transport and the resulting auxin gradient. In protonema, PpPINA is polarly distributed to the apical side of plasma membrane, and the expression is strongest at the tip of the apical cell (Viaene et al. 2014, Tang et al. 2023). Such localization implies an acropetal auxin transport from the basal cells towards the apex and out to the environment from the apical tip. An auxin gradient is observed by several reporter lines in P. patens protonema, perhaps relevant to the PIN localization (Viaene et al. 2014). However, the auxin distribution pattern varies depending on the reports or the kind of auxin reporters used. The R2D2 reporter line indicates a higher auxin sensing level at the base of filaments and gradually lower towards the tip (Thelander et al. 2019). In contrast, expression of DR5rev2:GUS was detected not only in the basal cells but also in the caulonema tip cells (Thelander et al. 2019). The expression pattern of GmGH3:GUS, whether it is strong at the basal cells or the tip cells, completely conflicts among reports (Bierfreund et al. 2003, Thelander et al. 2019). This variation may reflect the possibility that the Auxin Responsive Elements (AuxREs) in seed plants are potentially non-functional in mosses. Therefore, it can be assumed that the R2D2 reporter, which does not use the AuxREs (Jedličková et al. 2022), indicates the intracellular auxin level, or at least the sensing level, more precisely. As such, the auxin sensing level is presumably lower at the tip cells (Thelander et al. 2019). Considering that the expression of PpPINA in protonema is strongest at the apical tip (Viaene et al. 2014), it is conceivable that PIN is responsible for removing auxin from the apical cell. The site of auxin biosynthesis or the activity of the auxin receptor TIR1/AFB, both currently undescribed in P. patens protonema, may provide a better understanding of the cause of such auxin sensing gradient and the involvement of PIN.
In angiosperms, the localization of PIN is regulated through various processes, and the acquisition of similar mechanisms in bryophytes are suggested from the genetic analysis of M. polymorpha and P. patens (Table 1, Suetsugu et al. 2016, Naramoto 2017). However, ectopic expression of AtPIN1 in P. patens, or PpPINA, MpPIN1 in A. thaliana, all localized to the plasma membrane but did not recapitulate the polar localization of native PINs (Tang et al. 2023). In P. patens, the PIN localization in protonema is indicated as cytoskeleton-dependent; the treatment of microtubule depolymerizer Oryzalin abolishes the plasma membrane localization of PIN proteins, whereas actin depolymerizer LatB narrows down the region of PIN localization to a more limited area at the apical tip (Tang et al. 2023). This regulatory mechanism is not shared with A. thaliana and only partly with M. polymorpha, as AtPIN2 distribution in the root hair of A. thaliana was unaffected by both Oryzalin and LatB, and only Oryzalin disturbed the MpPIN1 distribution in M. polymorpha rhizoids (Tang et al. 2023). As such, the mechanism of PIN trafficking and polarization is diverged among land plants (Tang et al. 2023).
As the protonema of P. patens mature, some side branches start transitioning from filamentous growth to three-dimensional growth in a cytokinin-dependent manner. A single stem cell at the apex of the cell mass continues to divide, gradually forming leaf-like structures comprised of a single cell layer. New leaves are produced at a certain angle to the previously formed leaves, thus generating the gametophores (shoot-like structures) as mature gametophytic tissue. (Harrison et al. 2009, Kamamoto et al. 2021). Within this gametophore tissue, PIN proteins are expressed in the developing leaves (Bennett et al. 2014, Viaene et al. 2014, Lüth et al. 2023). All three PpPINs show a similar expression pattern, where it is initially strong at the tip of young leaves, and as the leaves develop, the expressed region gradually moves toward the basal area (Viaene et al. 2014, Lüth et al. 2023). Interestingly, this wave of PIN expression in the leaves correlates with the region where the transition from cell division to cell elongation occurs, suggesting the involvement of PIN and auxin transport in this process (Viaene et al. 2014).
The phenotypes observed in PIN overexpression and knockout mutants have supported this hypothesis (Bennett et al. 2014, Viaene et al. 2014). PINA overexpression leads to the formation of short and narrow leaves comprised of smaller cells than WT (Viaene et al. 2014). pina pinb double knockout mutants produce long, narrow leaves, phenocopied by gametophores treated with NPA and/or NAA (Bennett et al. 2014, Viaene et al. 2014). The cells of pina pinb mutant were long and wide, therefore implies that the accumulation of auxin promotes cell elongation and inhibits cell division (Viaene et al. 2014). This implication is consistent with the auxin gradient observed by the R2D2 reporter line. High auxin level was detected in the basal area of developing leaves, where cell elongation occurs (Thelander et al. 2019). A possible founder of this auxin gradient is the PIN proteins. However, the subcellular localization of PIN is complex in the leaves, in which a specific direction of auxin flow cannot be identified. Precisely, cells closer to the tip exhibit bipolar PIN distribution at the apical and basal side of the plasma membrane, whereas the basal cells lose PIN polar localization and PIN proteins are localized to all sides (Viaene et al. 2014, Tang et al. 2023). Thus, how PIN contributes to the auxin gradient within gametophore leaves remains unclear. Nevertheless, based on the phenotypes observed in knockout mutants and NAA/NPA-treated gametophores, the involvement of PIN and auxin in the development of gametophores seems unquestionable (Bennett et al. 2014, Viaene et al. 2014).
Within the gametophore tissue, the apex is identified as the place of auxin biosynthesis by the expression analysis of PpTAR (TAA-related enzyme), an auxin biosynthesis enzyme homologous to TAA (Landberg et al. 2021). However, the R2D2 reporter indicated an auxin sensing minimum in the shoot apex despite the biosynthesis of auxin (Thelander et al. 2019). An immunolocalization assay using the anti-maize PIN1 antibody detected PpPIN localization at the plasma membrane of the shoot apical stem cell. However, the specific PIN localization and the direction of auxin transport are unclear (Bennett et al. 2014). Treatment of highly concentrated NAA or 2,4-D, or co-treatment of NPA and NAA, inhibits the formation of gametophore leaves, suggesting that auxin accumulation leads to the disturbance of stem cell activity (Bennett et al. 2014). Therefore, it is conceivable that PIN maintains stem cell activity by controlling the intracellular auxin level (Bennett et al. 2014). However, as pina pinb double knockout mutants (Bennett et al. 2014), and even the pina pinb pinc triple knockout mutants can still produce gametophore leaves (Suemitsu and Naramoto unpublished), PIN-mediated auxin efflux seems not to be essential in this process.
In P. patens, both male (antheridia) and female (archegonia) reproductive organs are formed at the apex of a single gametophore (Hohe et al. 2002, Landberg et al. 2021). Despite the defects in gametophore morphology, pina pinb double knockout mutants form normal gametangia (Bennett et al. 2014). The detailed localization and expression of PINA and PINB in gametangia is not investigated, but for PINC, its expression at the mature antheridium and archegonium is reported (Lüth et al. 2023). Whereas pinc single knockout mutant shows no phenotype in the morphology of the plant body, several phenotypes in the reproductive process are observed, such as the increase of sperm motility, fertilization rate, and the abortion (falling out of sporophytes before maturation) rate. The direction of auxin transport in gametangia, as well as the involvement of PINA and PINB, remain unclear. Thus, further research is needed to identify how auxin transport contributes to these phenomena (Lüth et al. 2023).
After fertilization, the zygotes develop into the diploid tissue, sporophyte, with a short seta and a single spore capsule (sporangium) (Hohe et al. 2002). The existence of polar auxin transport is not investigated in P. patens sporophytes. However, basipetal auxin transport is detected in the sporophyte of several other mosses (Poli et al. 2003, Fujita et al. 2008); therefore, it can be assumed that the sporophytes of P. patens are the same. Consistently, the treatment of the auxin transport inhibitor NPA induces morphological defects in sporophyte development, such as malformed or multiple sporangia formation (Fujita et al. 2008). Similarly, pinb single knockout mutant produces two sporangia from a single seta, whereas pina single knockout mutant and pinc single knockout mutant form normal sporophyte tissues (Bennett et al. 2014, Lüth et al. 2023). More severe phenotypes in sporophyte development are observed in pina pinb double knockout mutant, suggesting the redundancy of PIN proteins in this process and PINB having the primary contribution (Bennett et al. 2014). The duplicated sporangia in pin mutants can be considered branching, one of the prominent phenomena polarly transported auxin regulates in angiosperms. Thus, the results of NPA treatment and the phenotypes of pin mutants in P. patens imply that growth regulation through polar auxin transport is conserved among the sporophyte of bryophytes and angiosperms (Bennett et al. 2014).
Tropic bending of angiosperms is one of the major roles of PIN and polar auxin transport. The localization or expression of PINs is changed in response to environmental cues, such as light or gravity, causing a differential auxin distribution within tissues. This auxin gradient causes differential growth, resulting in the bending of the tissue (Figure 2A). Mutants with defects in constructing this auxin gradient, such as the root of Atpin2 knockout mutant, lose the ability to bend in response to light, gravity, and other environmental cues (Han et al. 2021).
In P. patens, the gametophore shoots and protonema exhibit tropism based on the vectorial information of light and gravity. In response to light, protonema grows towards the direction of red light, a process controlled by the photoreceptor phytochrome (Jenkins and Cove 1983, Mittmann et al. 2004). Shoots exhibit positive phototropism towards the unilateral far-red, red, and blue light (Bao et al. 2015). The response to gravity can only be seen in dark conditions, where caulonema and shoots elongate in a negative-gravitropic direction (Jenkins et al. 1986). As a filamentous tissue, the tropic response of protonema is caused by the change in the direction of tip growth (Jenkins and Cove 1983). In contrast, the tropic bending of multicellular gametophores is likely caused by differential growth between the sides facing or opposite the environmental cue (Bao et al. 2015), similar to the root and shoot tropism of angiosperms. Mutants with defected caulonema gravitropism have been reported to exhibit normal gravitropism in shoots (Jenkins et al. 1986), implying that the mechanism underlying the tropism of protonema and gametophore may be different.
The phototropism and gravitropism of shoots were lost in pina pinb mutants, consistent with the pin knockout mutants in angiosperm. This was phenocopied by the application of exogenous auxin 2,4-D (2,4-dichlorophenoxyacetic acid), which is known to flatten the auxin gradient within tissues in seed plants, suggesting that the PIN-dependent auxin gradient within the shoot tissues regulates shoot tropism of P. patens (Bennett et al. 2014). In contrast, the negative-gravitropic growth of caulonema was not disturbed in pina pinb double mutant (Bennett et al. 2014). Since PINC remains functional in the double knockout mutant, there is a possibility that the defect was not evident due to the presence of the remaining canonical PIN. However, as the expression level of PINC is the lowest among PpPINs and scarcely expressed in protonema (Lüth et al. 2023), a more probable interpretation would be that the gravitropism of caulonema is PIN-independent (Bennett et al. 2014). Future study awaits to clarify this issue. In summary, the tropic response of P. patens shoots is assumed to be dependent on the auxin gradient caused by PIN (Bennett et al. 2014). The molecular mechanism of this regulation process remains unclear, such as whether PIN localization or expression changes in response to environmental stimuli, as observed in angiosperms. Future detailed study of the subcellular localization of PINs, together with the identification of other factors involved in PIN-mediated tropism of P. patens, may lead us to understand how the tropic responses have evolved in plants.
In contrast to moss, which had polar auxin transport detected in sporophytes, polar auxin transport is observed at the gametophytic stage of liverwort M. polymorpha (Figure 1C and 1D, Maravolo 1976, Gaal et al. 1982, Poli et al. 2003, Fujita et al. 2008). Consistently, treatment of exogenous auxin or auxin transport inhibitors to M. polymorpha, and the recently reported Mppin1 knockout mutant exhibit defects in various aspects of gametophyte development (Ishizaki et al. 2012, Fisher et al. 2023, Suzuki et al. 2023). In the sporophyte, GmGH3:GUS auxin reporter has identified that the auxin distribution moves towards the basal region as the sporophyte develops, and MpPIN1 expression is observed in the seta of sporophytes (Ishizaki et al. 2012, Fisher et al. 2023). However, the pin1 knockout mutant displayed no apparent phenotype in sporophyte development, suggesting the limited role of auxin transport at this stage (Fisher et al. 2023). As PIN-dependent polar auxin transport is deeply involved in the sporophyte development in moss P. patens (Fujita et al. 2008, Bennett et al. 2014), further analysis may help clarify the evolutionary process of auxin transport in diploid tissues. Nevertheless, the amount of information currently accessible is not sufficient to discuss this issue. Therefore, for now, we will review the role of auxin and its transportation during the gametophytic stage of M. polymorpha.
M. polymorpha germinates from either spores or gemmae, produced via sexual or vegetative reproduction, respectively. Upon spore germination, M. polymorpha produces filamentous protonema but subsequently shifts to two-dimensional growth, forming a cell mass. After a while, this cell mass generates an apical stem cell and develops an apical notch, forming a heart-shaped prothallus. As this prothallus grows, the division of the apical stem cell shifts to three-dimensional, thereby forming a mature thallus, with air pores and gemma cups formed at the dorsal side and rhizoids and ventral scales formed at the ventral side (Shimamura 2016, Naramoto et al. 2022). As a vegetative reproduction, M. polymorpha produces a gemma cup at the dorsal side of thallus, which produces gemmae inside. These gemmae possess two notches, each containing two potential apical stem cells aligned perpendicularly to the gemma (Shimamura 2016). After being dispersed from gemma cups, the dorsoventral axis is determined depending on the direction of light or gravity (Fitting 1936). Halbsguth (1937) suggested that this process is caused by the deactivation of one of the potential stem cells within each notch, with the cell at the side of light irradiation losing stem cell activity. The deactivated side differentiates into dorsal tissues, while the opposite side develops ventral tissues. Thus, the gemma generates a mature thallus with a dorsoventral axis (Bowman 2016). M. polymorpha can also regenerate from injured thalli by producing a callus-like cell mass from the apical side of the excised thallus, which gradually differentiates into a mature thallus (Ishida et al. 2022). Through the application of exogenous auxin or auxin inhibitors and phenotype analysis of Mppin1 mutant, auxin and PIN are identified to regulate all these processes (Eklund et al. 2015, Flores-Sandoval et al. 2015, 2018, Fisher et al. 2023, Suzuki et al. 2023).
During sporeling development, the Mppin1 knockout mutant exhibits delayed phase transition and cell differentiation (Fisher et al. 2023). Similar phenotypes are observed during the regeneration from injured thallus, where the growth of cell mass and differentiation of dorsal air pores are delayed in Mppin1 mutants (Fisher et al. 2023). This partly corresponds to the phenotypes of Mptaa, Mptir1 knockout mutants, or ectopic expression of MpARF3, which lack auxin biosynthesis, the intracellular auxin receptor, or repress auxin-induced gene expression throughout the thallus, respectively. These mutants produce a cell mass due to defects in cell differentiation (Eklund et al. 2015, Flores-Sandoval et al. 2018, Suzuki et al. 2023). In contrast, auxin-treated gemmae or auxin-hypersensitive mutants exhibit a severe growth defect and ectopic rhizoid differentiation (Flores-Sandoval et al. 2015, Suzuki et al. 2023). The GUS reporter analysis identified that the expression of auxin biosynthesis genes, as well as the repressor ARF, is high at the notches of the thallus (Eklund et al. 2015, Flores-Sandoval et al. 2018), thereby implying that auxin is synthesized but the sensitivity is low at the notch. Auxin sensing minima is indicated as an essential factor for establishing apical stem cells in M. polymorpha, as auxin inhibits the regeneration from an excised thallus (Ishida et al. 2022). Consistently, MpPIN1 expression first appears at the notch of the prothallus during sporeling development and at the notch of the regenerated thallus (Fisher et al. 2023). Therefore, the role of PIN in sporeling development or thallus regeneration is suggested to be the transportation of auxin away from the meristem to maintain the stem cell activity (Fisher et al. 2023).
Upon germination from gemma, Mppin1 mutants produce two shoots from each of the notches, resulting in the development of a mirrored thallus; one displaying a typical, and the other displaying an opposite dorsoventral orientation that has dorsal air pores facing downwards (Fisher et al. 2023). This phenotype resembles the gemma germinated under microgravity and without unidirectional light, suggesting that PIN regulates the light/gravity-mediated stem cell deactivation (Fitting 1936, Fisher et al. 2023). A similar phenotype is observed under the treatment of auxin transport inhibitor TIBA (Allsopp et al. 1968). In contrast, the application of exogenous auxin induces ectopic rhizoid formation from both sides without forming a mirrored thallus (Flores-Sandoval et al. 2015). The expression of MpPIN1 is detected from the notch of gemma, initially at both upper and lower sides, but loses its expression on the side illuminated by unidirectional light (Fisher et al. 2023). The side with remaining PIN expression develops as the apical meristem, whereas the opposite side differentiates into dorsal tissues, implying that stem cell activity was maintained by PIN auxin efflux, as in spore germination (Fisher et al. 2023). However, Mppin1 mutant exhibits defective dorsoventral pattern formation only in gemmaling development and not in sporeling development. Therefore, it is conceivable that MpPIN1 is not involved in apical stem cell initialization (Fisher et al. 2023).
As explained above, thallus is the mature gametophytic tissue of M. polymorpha, a flat three-dimensional tissue with a dorsoventral axis (Shimamura 2016). As they grow, each of the apical stem cells divides horizontally in two, thus causing branching of the thallus (Hirakawa et al. 2020). Within this thallus, polar auxin transport is detected from the midrib in a basipetal direction (Figure 1D, Maravolo 1976, Gaal et al. 1982). From the phenomenon observed in the regeneration from excised thallus, the repression of ectopic stem cell initiation is suggested as the role of this auxin transport (Ishida et al. 2022). Callus formation from an injured thallus occurs only when the notch is absent and mainly from the apical side of the excised thallus. This suggests that an inhibitory signal, presumably auxin, is generated from the apical notch and is transported basipetally. MpLAXR (LOW-AUXIN RESPONSIVE), a transcription factor expressed when the auxin level is low, is identified as a regulator of regeneration, by causing differentiated cells to re-enter the cell cycle (Ishida et al. 2022). The overexpression of MpLAXR produces cell mass without cell differentiation, reminiscent of the auxin-insensitive mutants such as Mptir1 (Ishida et al. 2022, Suzuki et al. 2023). As such, the basipetally transported auxin is assumed to repress the expression of MpLAXR, thus inhibiting the cells at the basal region entering the cell cycle (Ishida et al. 2022).
Contrary to the reported basipetal auxin transport, MpPIN1 localization is reported to exhibit no clear polarity on the plasma membrane throughout the thallus cells (Fisher et al. 2023). The Mppin1 knockout mutant thallus exhibits hyponastic growth, with the lateral side of the thallus curled inward to the midrib (Fisher et al. 2023). This phenotype is induced by the application of the auxin transport inhibitor TIBA but not by the application of exogenous auxin, and is not observed in the other auxin-related mutants (Eklund et al. 2015, Flores-Sandoval et al. 2015, 2018, Ishida et al. 2022, Fisher et al. 2023, Suzuki et al. 2023). Thus, auxin transport seems important for flat thallus development, but the molecular mechanism underlying this process remains to be determined. At a more local level, the Mppin1 mutant exhibits defects in the morphology of air pores and gemma cups (Fisher et al. 2023). These phenotypes can be induced by the application of exogenous auxin 2,4-D, the auxin transport inhibitor TIBA, or by the overexpression of auxin biosynthesis enzyme MpYUC, implying that the accumulation of auxin in the cells is responsible for such phenotypes (Eklund et al. 2015, Flores-Sandoval et al. 2015, Fisher et al. 2023). The phenotypes of the Mppin1 mutant, including the hyponastic growth, are complemented by expressing MpPIN1 using the constitutive promoter proMpEF1 or pro35S, as well as the native promoter (Fisher et al. 2023). Therefore, PIN and auxin transport are suggested to be functional throughout the thallus, regulating the cell morphology by controlling intracellular auxin levels (Fisher et al. 2023).
As explained earlier, tropic responses of angiosperms are regulated via PIN-mediated differential auxin distribution. In M. polymorpha, thalli, rhizoids, and gametangia stalks exhibit tropism in response to light or gravity. Thalli exhibit positive phototropism under the radiation of weak blue light by forming a narrow thallus that grows towards the light source (Komatsu et al. 2019). As a plagiotropic response, thallus placed with the dorsal side facing downwards gradually reorient its dorsoventrality as they grow (Fisher et al. 2023). Rhizoids, formed from the ventral side of the thallus, exhibit negative phototropism in response to blue light (Fisher et al. 2023). However, these tropic responses are not defective in the Mppin1 mutant, indicating that they are regulated by a PIN-independent mechanism (Fisher et al. 2023). The only tropic growth disturbed in the Mppin1 mutant was the phototropism/gravitropism of the gametangia stalk (Fisher et al. 2023). In WT, gametangia stalks are produced from the edge of the thallus and grow upright in a positive-phototropic and negative-gravitropic manner (Figure 2B, Inoue et al. 2019). In contrast, the Mppin1 mutant stalk grows without clear directionality, and although it can bend, it is apolar (Fisher et al. 2023). Treatment of exogenous auxin 2,4-D, or auxin transport inhibitors TIBA or NPA to the whole gametangia induces similar phenotypes, giving evidence that defective auxin transport abolished the tropism of the stalks of Mppin1 mutant (Fisher et al. 2023). Moreover, the application of 2,4-D on one side of the stalk induces bending towards the direction in which auxin was supplied (Fisher et al. 2023). Therefore, it is conceivable that PIN-mediated auxin transport generated an auxin gradient within the stalk tissues and caused differential growth reminiscent of the mechanism in angiosperms. In A. thaliana root and shoot, the change of PIN localization or expression is responsible for such differential auxin distribution (Han et al. 2021). In straightly developing M. polymorpha stalks, MpPIN1 is polarly localized to the basal sides of the plasma membrane, implying a basipetal auxin transport. However, upon light stimulation and the resulting tropic bending, the PIN polar localization of the cells at the bending region is lost, exhibiting distribution to all sides of the plasma membrane (Fisher et al. 2023). As auxin application to one side of the stalk induces bending towards that direction, it is assumed that this non-polar PIN distribution somehow causes auxin accumulation at the side facing the light source (Figure 2B). More precise observation of PIN localization may explain how M. polymorpha stalk establishes this auxin gradient. In summary, the positive phototropic growth of the M. polymorpha gametangia stalk is regulated by the shift of PIN localization, assumingly forming an auxin gradient between the sides of the stalk that results in differential growth (Fisher et al. 2023).
In the aspect of auxin distribution and the resulting bending direction, the stalk bending of M. polymorpha resembles the roots of angiosperms rather than the shoots. In the shoots of angiosperms, auxin accumulation promotes cell elongation, whereas in roots, auxin accumulation inhibits cell elongation (Han et al. 2021). The latter appears to be more similar to the case of the M. polymorpha stalk since growth was inhibited on the auxin-applied side (Figure 2A and 2B, Fisher et al. 2023). There are currently no reports on the cell length between each side of the bent stalk. Therefore, it is unclear whether the bending mechanisms of the gametangia stalk and the roots of angiosperms are genuinely the same. Nevertheless, the tropic responses caused by differential growth through the regulation of auxin distribution by PIN proteins are shared among liverworts and angiosperms (Fisher et al. 2023).
Figure 2 Tropic bending is caused by the generation of auxin gradient in angiosperms and liverworts.
(A, B) Auxin gradient induces bending of A. thaliana hypocotyl and root (A) and M. polymorpha gametangia stalk (B). The white arrowhead indicates the direction of light, and the black arrowhead indicates the direction of gravity. The blue color in the enlarged box indicates the intracellular auxin level. The blue arrow indicates the direction and the intensity of auxin transport.
Ferns are unique from the other land plant phylum in the aspect of ploidy, as the haploid and diploid tissues are viable independently, not reliant on one or the other. Moreover, while the morphology of sporophytic tissues is reminiscent of that of seed plants, with vasculatures, true leaves, and roots, the gametophytic tissue is similar to some of the bryophyte tissues. For example, the hermaphrodite gametophyte of Ceratopteris richardii and other ferns resembles the shape of M. polymorpha prothallus (Plackett et al. 2014), and Lygodium japonicum produces filamentous protonema as a male gametophyte (Ohishi et al. 2021). These traits in ferns provide a fundamental subject to study for elucidating the evolutionary processes of plant morphology. Among ferns, the technique for stable transformation by particle bombardment is established in C. richardii (Plackett et al. 2014). However, despite the academic importance, reports on the function of proteins in C. richardii involved in auxin transport and signaling scarcely exist. Nevertheless, pharmacological analysis focusing on the function of auxin and its transport has been reported in both the gametophytic and sporophytic stages of several fern species, giving us insights into how auxin is involved in the development of these plants.
Consistent with the morphological similarity, several reports have suggested a shared role of auxin among sporophytes of ferns and angiosperms. A basipetal auxin transport is detected in the rachis of Osmunda cinnamomea and Regnellidium diphyllum (Figure 1E, Steeves and Briggs 1960, Walters and Osborne 1979). The pinnae of O. cinnamomea and the leaflets of R. diphyllum are assumed to be the site of auxin biosynthesis, whose removal diminishes cell elongation at the rachis (Steeves and Briggs 1960, Walters and Osborne 1979). The pinnae removal of O. cinnamomea also inhibits the maturation of xylem cells in the rachis, where the formation of secondary cell walls is delayed (Steeves and Briggs, 1960). As auxin application recovers such phenomena, it is conceivable that auxin transport from the biosynthesis site is essential for rachis development (Steeves and Briggs 1960, Walters and Osborne 1979). In Pteris longifolia, excision of the primary leaf apex is reported to induce the lateral growth of a new leaf. Auxin application on the excised site inhibits such leaf formation, implying that auxin plays a role in the apical dominance of P. longifolia (Albaum 1938). These suggested roles of auxin transport in fern sporophytes: cell elongation, vasculature development, and apical dominance are common in angiosperms (Scarpella et al. 2006, Peer et al. 2011). Therefore, while auxin transport in bryophytes seems to work differently from angiosperms (as explained above), fern sporophytes may be using a system more similar to angiosperms.
In contrast to sporophytes, there are no reports on the measurement of polar auxin transport in fern gametophytes. However, previous studies have suggested auxin transport as a critical factor in their morphogenesis. The developmental process upon spore germination is well described in C. richardii, where they first produce a tongue-like thallus, which in hermaphrodite gametophytes, a lateral meristem is formed and thus developing a heart-like-shaped prothallus with a single notch (Conway and Di Stilio 2020). The meristem of C. richardii is identified as the site of auxin biosynthesis by the expression analysis of CrTAA. (Withers et al. 2023). Application of exogenous auxin 2,4-D, NAA, and IAA at a high concentration diminishes the formation of a lateral meristem, while the auxin transport inhibitor NPA induces the formation of multiple meristems (Gregorich and Fisher 2006, Withers et al. 2023). The formation of multiple meristems is also observed in L. japonicum by applying the auxin transport inhibitor TIBA (Ohishi et al. 2021). Other than the treatment of auxin transport inhibitors, the formation of a new meristem in fern prothallus is observed by the removal of the original meristem by laser ablation on C. richardii (Withers et al. 2023) and by the excision of P. longifolia (Albaum 1938). In P. longifolia, auxin application on the excised prothallus inhibits regeneration and the formation of new meristems (Albaum 1938). In summary of the reports of several fern species, it is conceivable that auxin synthesized at the meristem is transported to other cells of the prothallus, thus inhibiting the formation of an ectopic meristem and defining the morphology. Such presumed role of auxin in fern gametophytes is strongly reminiscent of that observed in M. polymorpha, in contrast to the angiosperm-like role of auxin in fern sporophytes. The molecular mechanisms underlying these phenomena, such as the function of PIN proteins, would be the subsequent interest. Future genetic studies are awaited to understand the precise function of auxin transport in ferns, which may help elucidate the evolution from bryophytes to seed plants.
As a fundamental phytohormone, auxin displays various effects on plant morphogenesis, from charophytes to angiosperms. Among them, the most conserved feature of auxin function may be the regulation of stem cell identity. The inhibition of cell division by auxin application is observed in all phyla described in this review, indicating a shared role of auxin as an inhibitor of cell proliferation. However, as inhibiting auxin biosynthesis in K. nitens suppresses its growth (Ohtaka et al. 2017), the role of auxin as a growth promoter seems to be conserved as well. Therefore, maintaining an optimal intracellular auxin level would be essential. The localization of PIN in K. nitens was at the side of filaments, not at the cell boundaries, suggesting KfPIN functions in actively exporting auxin to the outer environment (Skokan et al. 2019). Such PIN localization, which is not present at the cell boundaries, can be observed in land plants as well, in P. patens protonema, M. polymorpha rhizoids, and in A. thaliana root hair (Viaene et al. 2014, Tang et al. 2023). This shared feature among streptophytes suggests that this type of PIN localization pattern may be ancestral, with a role in maintaining intracellular auxin levels by actively removing auxin from the cell (Figure 3A). Cell-to-cell auxin transport is detected in the complex algae Chara (Boot et al. 2012). Therefore, PIN localization on the cell boundaries is presumably established at the common ancestor of charales and land plants. Auxin transport between cells determines the asymmetric distribution of auxin, which in turn gives characteristics to the cell in response to auxin levels. Charales can be specialized from other charophytes in the aspect of morphological complexity (Nishiyama et al. 2018). Taken together, the evolution of PIN auxin transport from “cell-to-environment” to “cell-to-cell” may have promoted cell differentiation and the resulting complex morphology by constructing a differential auxin distribution between tissues.
In angiosperms, this differential auxin distribution is determined through a complex PIN localization, constructing a patterned and canalized auxin flow, commonly called polar auxin transport (Figure 3C). Such auxin patterning regulates the morphology of tissues, such as the leaf vasculature patterning (Scarpella et al. 2006). However, from the observation of PIN localization in P. patens and M. polymorpha, the canalized auxin transport appears to be absent in the gametophyte of bryophytes (Viaene et al. 2014, Fisher et al. 2023). Considering the bryophyte-like simple tissues and the phenomena observed in several species, gametophytes of ferns can be assumed to be the same. Rather than forming a patterned canal of auxin transport, PIN proteins in gametophytes appear to function as a hose to remove auxin away from the apical stem cell (Figure 3B, Bennett et al. 2014, Fisher et al. 2023). Auxin is synthesized at the meristem of bryophytes and ferns (Eklund et al. 2015, Landberg et al. 2021 Withers et al. 2023). However, as this auxin inhibits cell division, auxin removal is essential for stem cell maintenance (Ishida et al. 2022, Thelander et al. 2019). PIN proteins in the gametophytes of non-seed plants may mainly be responsible of efficiently removing auxin from the stem cell region (Figure 3B). Given the fact that auxin is essential for cell differentiation and organogenesis in M. polymorpha (Suzuki et al. 2023), such auxin transportation may also serve as a provider of auxin to basal cells. In summary, the role of PIN in gametophytes of bryophytes and ferns seems to be relatively simple: transporting auxin away from the stem cells and maintaining the cell division ability, as well as providing auxin to basal cells to induce cell differentiation. Maravolo (1976) and Gaal et al. (1982) may have detected such transport of auxin as basipetal in the thallus of M. polymorpha (Figure 1D). Thus, the role of PIN auxin efflux can be classified into three types: 1) the “cell-to-environment auxin transport” to maintain the auxin level of the cell itself (Figure 3A), 2) the “non-canalized intercellular auxin transport” from the apical stem cell region to the surrounding cells to maintain stem cell activity and induce cell differentiation (Figure 3B), 3) the establishment of a “canalized auxin transport” to produce patterned cell differentiation and differential growth (Figure 3C). Considering the polar auxin transport previously observed, the one in the seta of moss sporophytes and the rachis of fern sporophytes (Figure 1C, 1E, Steeves and Briggs 1960, Walters and Osborne 1979, Poli et al. 2003, Fujita et al. 2008) can be considered as canalized, as auxin is transported in a long distance with clear directionality. However, the complexity may be lower than the canalized auxin transport in angiosperms. Still, a complexly canalized auxin flow can be assumed to be established in fern sporophytes, as they possess complex tissues such as the organized leaf vasculature, and auxin is involved in the vasculature formation of ferns (Steeves and Briggs 1960). The polar auxin transport observed in Chara is at a single-cell level (Figure 1B, Boot et al. 2012); therefore, it is difficult to determine whether it is canalized or non-canalized. For clarification, future studies on the auxin transport throughout the plant body, the subcellular PIN localization in the internodal cells, and identifying the auxin biosynthesis site would be necessary. Further research on the role of auxin transport and PIN in various organisms are awaited, which would give us new insights into the evolution of the function of auxin transport and when and how the transition of such roles occurred.
Figure 3 The three different models of PIN-mediated auxin transport in streptophytes.
(A-C) The hypothetical model of different types of PIN-mediated auxin transport in streptophytes and the constructed auxin gradient. (A) Auxin transport from the cell to the environment, caused by PIN localization at the outer side of the plasma membrane in the filamentous organism. (B) Non-canalized intercellular auxin transport away from the meristem caused by PIN localization at the cell boundaries facing away from the apical stem cell (marked with an asterisk). (C) Canalized auxin transport caused by cell-type-dependent differential PIN localization. The blue line indicates the localization of PIN proteins, and the blue arrow indicates the direction of auxin flow. The color of the cells indicates the assumed intracellular auxin level. (D) Phylogenetic tree of streptophytes and the establishment of each type of auxin transport.
This work was supported by a Grants-in-Aid for Scientific Research from the Japan Society for the Promotion of Science (KAKENHI grant number 22H04708 for S. N.) and the Japan Science and Technology Agency (PRESTO sakigake grant number JPMJPR22D6 for S. N.).
