Auxin signaling is essential for organogenesis but not for cell survival in the liverwort Marchantia polymorpha.

Auxin plays pleiotropic roles in plant development via gene regulation upon its perception by the receptors TRANSPORT INHIBITOR RESPONSE 1/AUXIN SIGNALING F-BOX (TIR1/AFBs). This auxin-regulated transcriptional control mechanism originated in the common ancestor of land plants. Although the complete loss of TIR1/AFBs causes embryonic lethality in Arabidopsis thaliana, it is unclear whether the requirement for TIR1-mediated auxin perception in cell viability can be generalized. The model liverwort Marchantia polymorpha has a minimal auxin signaling system with only a single TIR1/AFB, MpTIR1. Here we show by genetic, biochemical, and transcriptomic analyses that MpTIR1 functions as an evolutionarily conserved auxin receptor. Null mutants and conditionally knocked-out mutants of MpTIR1 were viable but incapable of forming any organs and grew as cell masses. Principal component analysis performed using transcriptomes at various developmental stages indicated that MpTIR1 is involved in the developmental transition from spores to organized thalli, during which apical notches containing stem cells are established. In Mptir1 cells, stem cell- and differentiation-related genes were up- and downregulated, respectively. Our findings suggest that, in M. polymorpha, auxin signaling is dispensable for cell division but is essential for three-dimensional patterning of the plant body by establishing pluripotent stem cells for organogenesis, a derived trait of land plants.

Diminished Auxin Signaling Triggers Cellular Reprogramming by Inducing a Regeneration Factor in the Liverwort Marchantia polymorpha.

Regeneration in land plants is accompanied by the establishment of new stem cells, which often involves reactivation of the cell division potential in differentiated cells. The phytohormone auxin plays pivotal roles in this process. In bryophytes, regeneration is enhanced by the removal of the apex and repressed by exogenously applied auxin, which has long been proposed as a form of apical dominance. However, the molecular basis behind these observations remains unexplored. Here, we demonstrate that in the liverwort Marchantia polymorpha, the level of endogenous auxin is transiently decreased in the cut surface of decapitated explants, and identify by transcriptome analysis a key transcription factor gene, LOW-AUXIN RESPONSIVE (MpLAXR), which is induced upon auxin reduction. Loss of MpLAXR function resulted in delayed cell cycle reactivation, and transient expression of MpLAXR was sufficient to overcome the inhibition of regeneration by exogenously applied auxin. Furthermore, ectopic expression of MpLAXR caused cell proliferation in normally quiescent tissues. Together, these data indicate that decapitation causes a reduction of auxin level at the cut surface, where, in response, MpLAXR is up-regulated to trigger cellular reprogramming. MpLAXR is an ortholog of Arabidopsis ENHANCER OF SHOOT REGENERATION 1/DORNRÖSCHEN, which has dual functions as a shoot regeneration factor and a regulator of axillary meristem initiation, the latter of which requires a low auxin level. Thus, our findings provide insights into stem cell regulation as well as apical dominance establishment in land plants.

Positional cues regulate dorsal organ formation in the liverwort Marchantia polymorpha.

Bryophytes and vascular plants represent the broadest evolutionary divergence in the land plant lineage, and comparative analyses of development spanning this divergence therefore offer opportunities to identify truisms of plant development in general. In vascular plants, organs are formed repetitively around meristems at the growing tips in response to positional cues. In contrast, leaf formation in mosses and leafy liverworts occurs from clonal groups of cells derived from a daughter cell of the apical stem cell known as merophytes, and cell lineage is a crucial factor in repetitive organ formation. However, it remains unclear whether merophyte lineages are a general feature of repetitive organ formation in bryophytes as patterns of organogenesis in thalloid liverworts are unclear. To address this question, we developed a clonal analysis method for use in the thalloid liverwort Marchantia polymorpha, involving random low-frequency induction of a constitutively expressed nuclear-targeted fluorescent protein by dual heat-shock and dexamethasone treatment. M. polymorpha thalli ultimately derive from stem cells in the apical notch, and the lobes predominantly develop from merophytes cleft to the left and right of the apical cell(s). Sector induction in gemmae and subsequent culture occasionally generated fluorescent sectors that bisected thalli along the midrib and were maintained through several bifurcation events, likely reflecting the border between lateral merophytes. Such thallus-bisecting sectors traversed dorsal air chambers and gemma cups, suggesting that these organs arise independently of merophyte cell lineages in response to local positional cues.

The Roles of the Sole Activator-Type Auxin Response Factor in Pattern Formation of Marchantia polymorpha.

Cell division patterning is important to determine body shape in plants. Nuclear auxin signaling mediated by AUXIN RESPONSE FACTOR (ARF) transcription factors affects plant growth and development through regulation of cell division, elongation and differentiation. The evolutionary origin of the ARF-mediated pathway dates back to at least the common ancestor of bryophytes and other land plants. The liverwort Marchantia polymorpha has three phylogenetically distinct ARFs: MpARF1, the sole 'activator' ARF; and MpARF2 and MpARF3, two 'repressor' ARFs. Genetic screens for auxin-resistant mutants revealed that loss of MpARF1 function conferred auxin insensitivity. Mparf1 mutants showed reduced auxin-inducible gene expression and various developmental defects, including thallus twisting and gemma malformation. We further investigated the role of MpARF1 in gemma development, which is traceable at the cellular level. In wild-type plants, a gemma initial first undergoes several transverse divisions to generate a single-celled stalk and a gemma proper, followed by rather synchronous longitudinal divisions in the latter. Mparf1 mutants often contained multicelled stalks and showed defects in the execution and timing of the longitudinal divisions. While wild-type gemmae finally generate two meristem notches, Mparf1 gemmae displayed various numbers of ectopic meristems. These results suggest that MpARF1 regulates formative cell divisions and axis formation through auxin responses. The mechanism for activator ARF regulation of pattern formation may be shared in land plants and therefore important for the general acquisition of three-dimensional body plans.

An ancestral function of strigolactones as symbiotic rhizosphere signals.

In flowering plants, strigolactones (SLs) have dual functions as hormones that regulate growth and development, and as rhizosphere signaling molecules that induce symbiosis with arbuscular mycorrhizal (AM) fungi. Here, we report the identification of bryosymbiol (BSB), an SL from the bryophyte Marchantia paleacea. BSB is also found in vascular plants, indicating its origin in the common ancestor of land plants. BSB synthesis is enhanced at AM symbiosis permissive conditions and BSB deficient mutants are impaired in AM symbiosis. In contrast, the absence of BSB synthesis has little effect on the growth and gene expression. We show that the introduction of the SL receptor of Arabidopsis renders M. paleacea cells BSB-responsive. These results suggest that BSB is not perceived by M. paleacea cells due to the lack of cognate SL receptors. We propose that SLs originated as AM symbiosis-inducing rhizosphere signaling molecules and were later recruited as plant hormone.

Auxin Biology in Bryophyta: A Simple Platform with Versatile Functions.

Bryophytes, including liverworts, mosses, and hornworts, are gametophyte-dominant land plants that are derived from a common ancestor and underwent independent evolution from the sporophyte-dominant vascular plants since their divergence. The plant hormone auxin has been shown to play pleiotropic roles in the haploid bodies of bryophytes. Pharmacological and chemical studies identified conserved auxin molecules, their inactivated forms, and auxin transport in bryophyte tissues. Recent genomic and molecular biological studies show deep conservation of components and their functions in auxin biosynthesis, inactivation, transport, and signaling in land plants. Low genetic redundancy in model bryophytes enable unique assays, which are elucidating the design principles of the auxin signaling pathway. In this article, the physiological roles of auxin and regulatory mechanisms of gene expression and development by auxin in Bryophyta are reviewed.

Design principles of a minimal auxin response system.

Auxin controls numerous growth processes in land plants through a gene expression system that modulates ARF transcription factor activity1-3. Gene duplications in families encoding auxin response components have generated tremendous complexity in most land plants, and neofunctionalization enabled various unique response outputs during development1,3,4. However, it is unclear what fundamental biochemical principles underlie this complex response system. By studying the minimal system in Marchantia polymorpha, we derive an intuitive and simple model where a single auxin-dependent A-ARF activates gene expression. It is antagonized by an auxin-independent B-ARF that represses common target genes. The expression patterns of both ARF proteins define developmental zones where auxin response is permitted, quantitatively tuned or prevented. This fundamental design probably represents the ancestral system and formed the basis for inflated, complex systems.