Flourishing in water: the early evolution and diversification of plant receptor-like kinases

Receptor-like kinases (RLKs) play significant roles in mediating innate immunity and development of plants. The evolution of plant RLKs has been characterized by extensive variation in copy numbers and domain configurations. However, much remains unknown about the origin, evolution, and early diversification of plant RLKs. Here, we perform phylogenomic analyses of RLKs across plants (Archaeplastida), including embryophytes, charophytes, chlorophytes, prasinodermophytes, glaucophytes, and rhodophytes. We identify the presence of RLKs in all the streptophytes (land plants and charophytes), nine out of 18 chlorophytes, one prasinodermophyte, and one glaucophyte, but not in rhodophytes. Interestingly, the copy number of RLKs increased drastically in streptophytes after the split of the clade of Mesostigmatophyceae and Chlorokybophyceae and other streptophytes. Moreover, phylogenetic analyses suggest RLKs from charophytes form diverse distinct clusters, and are dispersed along the diversity of land plant RLKs, indicating that RLKs have extensively diversified in charophytes and charophyte RLKs seeded the major diversity of land plant RLKs. We identify at least 81 and 76 different kinase-associated domains for charophyte and land plant RLKs, 23 of which are shared, suggesting that RLKs might have evolved in a modular fashion through frequent domain gains or losses. We also detect signatures of positive selection for many charophyte RLK groups, indicating potential functions in host–microbe interaction. Taken together, our findings provide significant insights into the early evolution and diversification of plant RLKs and the ancient evolution of plant–microbe symbiosis.     

Occurrence and evolutionary significance of RESISTANT CELL WALLS IN CHAROPHYTES AND BRYOPHYTES

A survey of charophycean green algal and bryophyte taxa revealed the frequent occurrence of vegetative cell walls that were characterized by a specific form of autofluorescence and resistance to high temperature acid treatment (acetolysis). The time of production and the location of resistant, autofluorescent cell walls varied among charophyte and bryophyte taxa in patterns that suggest that bryophytes inherited the capacity to produce such walls from charophyte ancestors. A number of charophytes produced resistant walls in response to desiccation stress, suggesting an evolutionarily early adaptive response. Coleochaete was unique among charophytes, but similar to all bryophytes tested in that sexual reproduction induced autofluorescence in cell walls of well-hydrated tissues at the placental junction. Maternal tissues in apical portions of the pseudoseta bearing Sphagnum sporophytes were characterized by autofluorescent, acetolysis-resistant cell walls similar to those observed in maternal cells adjacent to Coleochaete zygotes. These observations suggest that cell–cell stimulus–response interactions regulate deposition of autofluorescent compounds in placental cell walls, and that this characteristic may have been shared by the earliest embryophytes and their charophyte ancestors. Various bryophytes deposit autofluorescent, acid-resistant compounds at other adaptively significant sites including sporangial epidermis, spiral thickenings of elaters, rhizoids, and leaves in the special case of Sphagnum moss. Sphagnum and liverwort sporangial epidermis, which had been subjected to acetolysis or strong acid procedures commonly used to release microfossils from rock matrices, resembled published photographs of Ordovician–Devonian microfossils consisting of cellular scraps that have been attributed to earliest land plants. Our work suggests that at least some of these fossils, previously thought to represent “dispersed cuticles,” could be reinterpreted as earliest known remains of plant sporophytic tissues, and that they may be homologous with resistant sporangial epidermis of modern bryophytes. In general, the patterns of occurrence of resistant, autofluorescent cell walls in charophytes and bryophytes suggest repeated exaptation. Regulation of deposition appears to have been modified through time, so that resistant wall compounds have had a sequence of functions: desiccation resistance and/or microbial resistance in lower charophytes, a role in embryogenesis in Coleochaete and embryophytes, and finally, decay resistance in innovative structures that characterize bryophytes, such as rhizoids, sporangial epidermis, and elaters.     

Primary cell wall composition of bryophytes and charophytes

Major differences in primary cell wall (PCW) components between non-vascular plant taxa are reported. (1) Xyloglucan: driselase digestion yielded isoprimeverose (the diagnostic repeat unit of xyloglucan) from PCW-rich material of Anthoceros (a hornwort), mosses and both leafy and thalloid liverworts, as well as numerous vascular plants, showing xyloglucan to be a PCW component in all land plants tested. In contrast, charophycean green algae (Klebsormidium flaccidium, Coleochaete scutata and Chara corallina), thought to be closely related to land plants, did not contain xyloglucan. They did not yield isoprimeverose; additionally, charophyte material was not digestible with xyloglucan-specific endoglucanase or cellulase to give xyloglucan-derived oligosaccharides. (2) Uronic acids: acid hydrolysis of PCW-rich material from the charophytes, the hornwort, thalloid and leafy liverworts and a basal moss yielded higher concentrations of glucuronic acid than that from the remaining land plants including the less basal mosses and all vascular plants tested. Polysaccharides of the hornwort Anthoceros contained an unusual repeat-unit, glucuronic acid-alpha(1-->3)-galactose, not found in appreciable amounts in any other plants tested. Galacturonic acid was consistently the most abundant PCW uronic acid, but was present in higher concentrations in acid hydrolysates of bryophytes and charophytes than in those of any of the vascular plants. Mannuronic acid was not detected in any of the species surveyed. (3) Mannose: acid hydrolysis of charophyte and bryophyte PCW-rich material also yielded appreciably higher concentrations of mannose than are found in vascular plant PCWs. (4) Mixed-linkage glucan (MLG) was absent from all algae and bryophytes tested; however, upon digestion with licheninase, PCW-rich material from the alga Ulva lactuca and the leafy liverwort Lophocolea bidentata yielded penta- to decasaccharides, indicating the presence of MLG-related polysaccharides. Our results show that major evolutionary events are often associated with changes in PCW composition. In particular, the acquisition of xyloglucan may have been a pre-adaptive advantage that allowed colonization of land.     

The GapA/B gene duplication marks the origin of Streptophyta (charophytes and land plants)

Independent evidence from morphological, ultrastructural, biochemical, and molecular data have shown that land plants originated from charophycean green algae. However, the branching order within charophytes is still unresolved, and contradictory phylogenies about, for example,the position of the unicellular green alga Mesostigma viride are difficult to reconcile. A comparison of nuclear-encoded Calvin cycle glyceraldehyde-3-phosphate dehydrogenases (GAPDH) indicates that a crucial duplication of the GapA gene occurred early in land plant evolution. The duplicate called GapB acquired a characteristic carboxy-terminal extension (CTE) from the general regulator of the Calvin cycle CP12. This CTE is responsible for thioredoxin-dependent light/dark regulation. In this work, we established GapA, GapB, and CP12 sequences from bryophytes, all orders of charophyte as well as chlorophyte green algae, and the glaucophyte Cyanophora paradoxa. Comprehensive phylogenetic analyses of all available plastid GAPDH sequences suggest that glaucophytes and green plants are sister lineages and support a positioning of Mesostigma basal to all charophycean algae. The exclusive presence of GapB in terrestrial plants, charophytes, and Mesostigma dates the GapA/B gene duplication to the common ancestor of Streptophyta. The conspicuously high degree of GapB sequence conservation suggests an important metabolic role of the newly gained regulatory function. Because the GapB-mediated protein aggregation most likely ensures the complete blockage of the Calvin cycle at night, we propose that this mechanism is also crucial for efficient starch mobilization. This innovation may be one prerequisite for the development of storage tissues in land plants.     

Evolutionary patterns in auxin action

This review represents the first effort ever to survey the entire literature on auxin (indole-3-acetic acid, IAA) action in all plants, with special emphasis on the green plant lineage, including charophytes (the green alga group closest to the land plants), bryophytes (the most basal land plants), pteridophytes (vascular non-seed plants), and seed plants. What emerges from this survey is the surprising perspective that the physiological mechanisms for regulating IAA levels and many IAA-mediated responses found in seed plants are also present in charophytes and bryophytes, at least in nascent forms. For example, the available evidence suggests that the apical regions of both charophytes and liverworts synthesize IAA via a tryptophan-independent pathway, with IAA levels being regulated via the balance between the rates of IAA biosynthesis and IAA degradation. The apical regions of all the other land plants utilize the same class of biosynthetic pathway, but they have the potential to utilize IAA conjugation and conjugate hydrolysis reactions to achieve more precise spatial and temporal control of IAA levels. The thallus tips of charophytes exhibit saturable IAA influx and efflux carriers, which are apparently not sensitive to polar IAA transport inhibitors. By contrast, two divisions of bryophyte gametophytes and moss sporophytes are reported to carry out polar IAA transport, but these groups exhibit differing sensitivities to those inhibitors. Although the IAA regulation of charophyte development has received almost no research attention, the bryophytes manifest a wide range of developmental responses, including tropisms, apical dominance, and rhizoid initiation, which are subject to IAA regulation that resembles the hormonal control over corresponding responses in seed plants. In pteridophytes, IAA regulates root initiation and vascular tissue differentiation in a manner also very similar to its effects on those processes in seed plants. Thus, it is concluded that the seed plants did not evolve de novo mechanisms for mediating IAA responses, but have rather modified pre-existing mechanisms already operating in the early land plants. Finally, this paper discusses the encouraging prospects for investigating the molecular evolution of auxin action.     

Charophyten und Otolithen aus den Cyrenen-Schichten des nördlichen Alpenvorlandes

Charophytes and otoliths from theCyrena Beds of the northern Alpine foreland (Hausham and Miesbach synclines, Upper Bavaria) have been studied. A very abundant charophyte flora comprising 12 species, includingStephanochara martinii n. sp., is described and figured. The moderately diverse otolith fauna is documented by tables and SEM-pictures. The studiedCyrena Beds can be dated biostratigraphically as early Chattian and correlated with theChara microcera Zone. The charophytes show strong paleobiogeographic relations to floras in the Mainz Basin/Upper Rhine Graben as well as to western Switzerland and southern France. The otolith fauna is strikingly different from late Chattian faunas of the western Paratethys. The paleoecology indicated by charophytes, otoliths and the accompanying fauna suggests short term variation including freshwater, brackish water and marine influenced environments.     

Multigene phylogeny of the green lineage reveals the origin and diversification of land plants

The Viridiplantae (green plants) include land plants as well as the two distinct lineages of green algae, chlorophytes and charophytes. Despite their critical importance for identifying the closest living relatives of land plants, phylogenetic studies of charophytes have provided equivocal results [1-5]. In addition, many relationships remain unresolved among the land plants, such as the position of mosses, liverworts, and the enigmatic Gnetales. Phylogenomics has proven to be an insightful approach for resolving challenging phylogenetic issues, particularly concerning deep nodes [6-8]. Here we extend this approach to the green lineage by assembling a multilocus data set of 77 nuclear genes (12,149 unambiguously aligned amino acid positions) from 77 taxa of plants. We therefore provide the first multigene phylogenetic evidence that Coleochaetales represent the closest living relatives of land plants. Moreover, our data reinforce the early divergence of liverworts and the close relationship between Gnetales and Pinaceae. These results provide a new phylogenetic framework and represent a key step in the evolutionary interpretation of developmental and genomic characters in green plants.     

The complete mitochondrial DNA sequence of Mesostigma viride identifies this green alga as the earliest green plant divergence and predicts a highly compact mitochondrial genome in the ancestor of all green plants.

To gain insights into the nature of the mitochondrial genome in the common ancestor of all green plants, we have completely sequenced the mitochondrial DNA (mtDNA) of Mesostigma viride. This green alga belongs to a morphologically heterogeneous class (Prasinophyceae) that includes descendants of the earliest diverging green plants. Recent phylogenetic analyses of ribosomal RNAs (rRNAs) and concatenated proteins encoded by the chloroplast genome identified Mesostigma as a basal branch relative to the Streptophyta and the Chlorophyta, the two phyla that were previously thought to contain all extant green plants. The circular mitochondrial genome of Mesostigma resembles the mtDNAs of green algae occupying a basal position within the Chlorophyta in displaying a small size (42,424 bp) and a high gene density (86.6% coding sequences). It contains 65 genes that are conserved in other mtDNAs. Although none of these genes represents a novel coding sequence among green plant mtDNAs, four of them (rps1, sdh3, sdh4, and trnL[caa]) have not been reported previously in chlorophyte mtDNAs, and two others (rpl14 and trnI[gau]) have not been identified in the streptophyte mtDNAs examined so far (land-plant mtDNAs). Phylogenetic analyses of 19 concatenated mtDNA-encoded proteins favor the hypothesis that Mesostigma represents the earliest branch of green plant evolution. Four group I introns (two in rnl and two in cox1) and three group II introns (two in nad3 and one in cox2), two of which are trans-spliced at the RNA level, reside in Mesostigma mtDNA. The insertion sites of the three group II introns are unique to this mtDNA, suggesting that trans-splicing arose independently in the Mesostigma lineage and in the Streptophyta. The few structural features that can be regarded as ancestral in Mesostigma mtDNA predict that the common ancestor of all green plants had a compact mtDNA containing a minimum of 75 genes and perhaps two group I introns. Considering that the mitochondrial genome is much larger in size in land plants than in Mesostigma, we infer that mtDNA size began to increase dramatically in the Streptophyta either during the evolution of charophyte green algae or during the transition from charophytes to land plants.     

Contributions of plant molecular systematics to studies of molecular evolution

Dobzhansky stated that nothing in biology makes sense except in the light of evolution. A close corollary, and the central theme of this paper, is that everything makes a lot more sense in the light of phylogeny. Systematics is in the midst of a renaissance, heralded by the widespread application of new analytical approaches and the introduction of molecular techniques. Molecular phylogenetic analyses are now commonplace, and they have provided unparalleled insights into relationships at all levels of plant phylogeny. At deep levels, molecular studies have revealed that charophyte green algae are the closest relatives of the land plants and suggested that liverworts are sister to all other extant land plants. Other studies have suggested that lycopods are sister to all other vascular plants and clarified relationships among the ferns. The impact of molecular phylogenetics on the angiosperms has been particularly dramatic – some of the largest phylogenetic analyses yet conducted have involved the angiosperms. Inferences from three genes (rbcL, atpB, 18S rDNA) agree in the major features of angiosperm phylogeny and have resulted in a reclassification of the angiosperms. This ordinal-level reclassification is perhaps the most dramatic and important change in higher-level angiosperm taxonomy in the past 200 years. At lower taxonomic levels, phylogenetic analyses have revealed the closest relatives of many crops and model organisms for studies of molecular genetics, concomitantly pointing to possible relatives for use in comparative studies and plant breeding. Furthermore, phylogenetic information has contributed to new perspectives on the evolution of polyploid genomes. The phylogenetic trees now available at all levels of the taxonomic hierarchy for angiosperms and other green plants should play a pivotal role in comparative studies in diverse fields from ecology to molecular evolution and comparative genetics.     

Charophyte algae and land plant origins

The charophyte algae are six distinct groups of mostly freshwater green algae that are related to modern land plants. Charophyte algae exhibit diverse morphologies and reproductive strategies, from unicells to branching erect forms, and from swimming asexual spores to sex involving eggs and sperm, respectively. The green algae known as stoneworts (Charales) are suggested to be the extant sister group to all land plants, although the phylogeny is not conclusive. Here we review recent molecular phylogenetic work on the charophyte algae and its implications for our understanding of the origins of land plants and of characters in their aquatic ancestors that might have played a role in the explosive diversification of plants on land.     

Green algae to land plants: An evolutionary transition

Studies focused upon the evolutionary transition from ancestral green algae to the earliest land plants are important from a range of ecological, molecular and evolutionary perspectives. A substantial suite of ultrastructural, biochemical and molecular data supports the concept that land plants (embryophytes) are monophyletically derived from an ancestral charophycean alga. However, the details of phylogenetic branching patterns linking extant charophytes and seedless embryophytes are currently unclear. Moreover, the fossil record has so far been mute regarding the algae-land plant transition. Nevertheless, an accurate reflection of major evolutionary events in the history of the earliest land plants can be obtained by comparative paleontological-neontological studies, and comparative molecular, cellular and developmental investigations of extant charophytes and bryophytes. This review focuses upon research progress toward understanding three clade-specific adaptations that were important in the successful colonization of land by plants: the histogenetic apical meristem, the matrotrophic embryo, and decay-resistant cell wall polymers.     

The colonisation of disturbed freshwater habitats byCharaceae

Various species of charophytes are able rapidly to colonise disturbed or cleaned shallow water bodies. This fact may be explained by the long-lasting viability of charophyte oospores, their easy dispersal and, probably, enhanced germination after passage through digestive tracts of waterfowl. Vegetative parts of Charophytes are sometimes resistant to herbicide treatment.     

Stable transformation and reverse genetic analysis of Penium margaritaceum: a platform for studies of charophyte green algae,the immediate ancestors of land plants

The charophyte green algae (CGA, Streptophyta, Viridiplantae) occupy a key phylogenetic position as the immediate ancestors of land plants but, paradoxically, are less well‐studied than the other major plant lineages. This is particularly true in the context of functional genomic studies, where the lack of an efficient protocol for their stable genetic transformation has been a major obstacle. Observations of extant CGA species suggest the existence of some of the evolutionary adaptations that had to occur for land colonization; however, to date, there has been no robust experimental platform to address this genetically. We present a protocol for high‐throughput Agrobacterium tumefaciens‐mediated transformation of Penium margaritaceum, a unicellular CGA species. The versatility of Penium as a model for studying various aspects of plant cell biology and development was illustrated through non‐invasive visualization of protein localization and dynamics in living cells. In addition, the utility of RNA interference (RNAi) for reverse genetic studies was demonstrated by targeting genes associated with cell wall modification (pectin methylesterase) and biosynthesis (cellulose synthase). This provided evidence supporting current models of cell wall assembly and inter‐polymer interactions that were based on studies of land plants, but in this case using direct observation in vivo. This new functional genomics platform has broad potential applications, including studies of plant organismal biology and the evolutionary innovations required for transition from aquatic to terrestrial habitats.     

Seasonal variation in invertebrate grazing on Chara connivens and C. tomentosa in Kõiguste Bay, NE Baltic Sea

Charophytes are a highly endangered group of algae. In the Baltic Sea, the number of species, distribution area and biomass of charophytes have significantly decreased in recent decades. Although eutrophication triggers their initial decline, the mechanism of the final extinction of charophyte populations is not fully understood. An in situ experiment was performed to study the role of the mesoherbivores Idotea baltica, Gammarus oceanicus and Palaemon adspersus in the decline of charophytes in the north-eastern Baltic Sea. Invertebrate grazing showed a clear seasonality: grazing pressure was low in April, moderate in July, and high in October. Grazing on charophytes by P. adspersus was negligible, whereas I. baltica and G. oceanicus significantly reduced the biomass of charophytes in the field. Low photosynthetic activity (high decomposition rate) of the charophytes favoured grazing. The invertebrates studied preferred Chara tomentosa to C. connivens. Low consumption of C. connivens may reflect its non-native origin. The experiment suggests that, under moderately eutrophic conditions, grazers are not likely to control charophyte populations. However, grazers have the potential to eliminate charophytes in severely eutrophic systems under the stress of filamentous algae.Communicated by H.D. Franke     

PHYLOGENETIC RELATIONSHIPS AMONG STREPTOPHYTES AS INFERRED FROM CHLOROPLAST SMALL AND LARGE SUBUNIT rRNA GENE SEQUENCES1

To gain insights into the phylogeny of charophytes and into their relationships with other green algae and bryophytes, we analyzed the chloroplast small and large subunit rRNA sequences of charophytes belonging to five orders (Charales, Coleochaetales, Desmidiales, Klebsormidiales, and Zygnematales), of chlorophytes from the four remaining classes of green algae, and of bryophytes representing the three classes reported in this group of land plants. We also probed the gene organization and intron content of the chloroplast rDNA operon in charophytes and bryophytes. The organization of this operon proved to be highly conserved, except in members of the Desmidiales and Zygnematales. Homologous group II introns were identified in the trnA(UGC) gene of all charophyte groups examined and in the trnI(GAU) gene of charophytes from all orders except the Desmidiales and Zygnematales. Phylogenetic analyses of concatenated rDNA sequences consistently placed the prasinophyte Mesostigma viride Lauterborn at the base of the Streptophyta and Chlorophyta, although alternative topologies positioning Mesostigma within the Streptophyta could not be rejected. A sister group relationship was unambiguously established between Chaetosphaeridium globosum (Nordstedt) Klebahn and members of the Coleochaetales. The Charales, Coleochaetales, Desmidiales, and Zygnematales were found to be monophyletic, and a sister group relationship was observed for the Desmidiales and Zygnematales. Although our analyses failed to resolve the branching order of the Coleochaetales, Charales, Desmidiales/Zygnematales, and bryophytes, they revealed that the problematic charophyte taxon Entransia fimbriata Hughes strongly clusters with Klebsormidium flaccidum (Kützing) Silva, Mattox et Blackwell to form a basal lineage relative to the other charophyte orders examined.     

Green land: Multiple perspectives on green algal evolution and the earliest land plants

Green plants, broadly defined as green algae and the land plants (together, Viridiplantae), constitute the primary eukaryotic lineage that successfully colonized Earth's emergent landscape. Members of various clades of green plants have independently made the transition from fully aquatic to subaerial habitats many times throughout Earth's history. The transition, from unicells or simple filaments to complex multicellular plant bodies with functionally differentiated tissues and organs, was accompanied by innovations built upon a genetic and phenotypic toolkit that have served aquatic green phototrophs successfully for at least a billion years. These innovations opened an enormous array of new, drier places to live on the planet and resulted in a huge diversity of land plants that have dominated terrestrial ecosystems over the past 500 million years. This review examines the greening of the land from several perspectives, from paleontology to phylogenomics, to water stress responses and the genetic toolkit shared by green algae and plants, to the genomic evolution of the sporophyte generation. We summarize advances on disparate fronts in elucidating this important event in the evolution of the biosphere and the lacunae in our understanding of it. We present the process not as a step-by-step advancement from primitive green cells to an inevitable success of embryophytes, but rather as a process of adaptations and exaptations that allowed multiple clades of green plants, with various combinations of morphological and physiological terrestrialized traits, to become diverse and successful inhabitants of the land habitats of Earth.     

Sugar composition of the pectic polysaccharides of charophytes,the closest algal relatives of land-plants: presence of 3-O-methyl-d-galactose residues

Background and Aims During evolution, plants have acquired and/or lost diverse sugar residues as cell-wall constituents. Of particular interest are primordial cell-wall features that existed, and in some cases abruptly changed, during the momentous step whereby land-plants arose from charophytic algal ancestors.Methods Polysaccharides were extracted from four charophyte orders [Chlorokybales (Chlorokybus atmophyticus), Klebsormidiales (Klebsormidium fluitans, K. subtile), Charales (Chara vulgaris, Nitella flexilis), Coleochaetales (Coleochaete scutata)] and an early-diverging land-plant (Anthoceros agrestis). ‘Pectins’ and ‘hemicelluloses’, operationally defined as extractable in oxalate (100 °C) and 6 m NaOH (37 °C), respectively, were acid- or Driselase-hydrolysed, and the monosaccharides analysed chromatographically. One unusual monosaccharide, ‘U’, was characterized by ¹H/¹³C-nuclear magnetic resonance spectroscopy and also enzymically.Key Results ‘U’ was identified as 3-O-methyl-d-galactose (3-MeGal). All pectins, except in Klebsormidium, contained acid- and Driselase-releasable galacturonate, suggesting homogalacturonan. All pectins, without exception, released rhamnose and galactose on acid hydrolysis; however, only in ‘higher’ charophytes (Charales, Coleochaetales) and Anthoceros were these sugars also efficiently released by Driselase, suggesting rhamnogalacturonan-I. Pectins of ‘higher’ charophytes, especially Chara, contained little arabinose, instead possessing 3-MeGal. Anthoceros hemicelluloses were rich in glucose, xylose, galactose and arabinose (suggesting xyloglucan and arabinoxylan), none of which was consistently present in charophyte hemicelluloses.Conclusions Homogalacturonan is an ancient streptophyte feature, albeit secondarily lost in Klebsormidium. When conquering the land, the first embryophytes already possessed rhamnogalacturonan-I. In contrast, charophyte and land-plant hemicelluloses differ substantially, indicating major changes during terrestrialization. The presence of 3-MeGal in charophytes and lycophytes but not in the ‘intervening’ bryophytes confirms that cell-wall chemistry changed drastically between major phylogenetic grades.     

TRANSITION TO A LAND FLORA: A MOLECULAR PHYLOGENETIC PERSPECTIVE

Phylogenetic analyses of morphoultrastructural characters and DNA sequences of chloroplast, mitochondrial, and nuclear genes and investigations of genomic structural features in both organellar genomes have clarified several difficult issues in charophyte and bryophyte phylogenies. Mesostigma seems to represent the first lineage of streptophytes, and Charales are a derived member of charophytes. Liverworts and hornworts both are candidates for the basal-most lineage in land plants according to analyses of morphoultrastructural and sequence data, but mitochondrial intron distribution favors the former. Takakia is likely a basal moss. More studies using both multigene sequence analyses and genomic structural characters will significantly improve our understanding of the origin and early evolution of land plants.