Genomes of Subaerial Zygnematophyceae Provide Insights into Land Plant Evolution.

The transition to a terrestrial environment, termed terrestrialization, is generally regarded as a pivotal event in the evolution and diversification of the land plant flora that changed the surface of our planet. Through phylogenomic studies, a group of streptophyte algae, the Zygnematophyceae, have recently been recognized as the likely sister group to land plants (embryophytes). Here, we report genome sequences and analyses of two early diverging Zygnematophyceae (Spirogloea muscicola gen. nov. and Mesotaenium endlicherianum) that share the same subaerial/terrestrial habitat with the earliest-diverging embryophytes, the bryophytes. We provide evidence that genes (i.e., GRAS and PYR/PYL/RCAR) that increase resistance to biotic and abiotic stresses in land plants, in particular desiccation, originated or expanded in the common ancestor of Zygnematophyceae and embryophytes, and were gained by horizontal gene transfer (HGT) from soil bacteria. These two Zygnematophyceae genomes represent a cornerstone for future studies to understand the underlying molecular mechanism and process of plant terrestrialization.

Lower Statistical Support with Larger Data Sets: Insights from the Ochrophyta Radiation.

It is commonly assumed that increasing the number of characters has the potential to resolve evolutionary radiations. Here, we studied photosynthetic stramenopiles (Ochrophyta) using alignments of heterogeneous origin mitochondrion, plastid, and nucleus. Surprisingly while statistical support for the relationships between the six major Ochrophyta lineages increases when comparing the mitochondrion (6,762 sites) and plastid (21,692 sites) trees, it decreases in the nuclear (209,105 sites) tree. Statistical support is not simply related to the data set size but also to the quantity of phylogenetic signal available at each position and our ability to extract it. Here, we show that this ability for current phylogenetic methods is limited, because conflicting results were obtained when varying taxon sampling. Even though the use of a better fitting model improved signal extraction and reduced the observed conflicts, the plastid data set provided higher statistical support for the ochrophyte radiation than the larger nucleus data set. We propose that the higher support observed in the plastid tree is due to an acceleration of the evolutionary rate in one short deep internal branch, implying that more phylogenetic signal per position is available to resolve the Ochrophyta radiation in the plastid than in the nuclear data set. Our work therefore suggests that, in order to resolve radiations, beyond the obvious use of data sets with more positions, we need to continue developing models of sequence evolution that better extract the phylogenetic signal and design methods to search for genes/characters that contain more signal specifically for short internal branches.

Plastid Genome Evolution of Two Colony-Forming Benthic Ochrosphaera neapolitana Strains (Coccolithales, Haptophyta).

Coccolithophores are well-known haptophytes that produce small calcium carbonate coccoliths, which in turn contribute to carbon sequestration in the marine environment. Despite their important ecological role, only two of eleven haptophyte plastid genomes are from coccolithophores, and those two belong to the order Isochrysidales. Here, we report the plastid genomes of two strains of Ochrosphaera neapolitana (Coccolithales) from Spain (CCAC 3688 B) and the USA (A15,280). The newly constructed plastid genomes are the largest in size (116,906 bp and 113,686 bp, respectively) among all the available haptophyte plastid genomes, primarily due to the increased intergenic regions. These two plastid genomes possess a conventional quadripartite structure with a long single copy and short single copy separated by two inverted ribosomal repeats. These two plastid genomes share 110 core genes, six rRNAs, and 29 tRNAs, but CCAC 3688 B has an additional CDS (ycf55) and one tRNA (trnL-UAG). Two large insertions at the intergenic regions (2 kb insertion between ycf35 and ycf45; 0.5 kb insertion in the middle of trnM and trnY) were detected in the strain CCAC 3688 B. We found the genes of light-independent protochlorophyllide oxidoreductase (chlB, chlN, and chlL), which convert protochlorophyllide to chlorophyllide during chlorophyll biosynthesis, in the plastid genomes of O. neapolitana as well as in other benthic Isochrysidales and Coccolithales species, putatively suggesting an evolutionary adaptation to benthic habitats.

Chromosome-level genome of Pedinomonas minor (Chlorophyta) unveils adaptations to abiotic stress in a rapidly fluctuating environment.

The Pedinophyceae (Viridiplantae) comprise a class of small uniflagellate algae with a pivotal position in the phylogeny of the Chlorophyta as the sister group of the 'core chlorophytes'. We present a chromosome-level genome assembly of the freshwater type species of the class, Pedinomonas minor. We sequenced the genome using Pacbio, Illumina and Hi-C technologies, performed comparative analyses of genome and gene family evolution, and analyzed the transcriptome under various abiotic stresses. Although the genome is relatively small (55 Mb), it shares many traits with core chlorophytes including number of introns and protein-coding genes, messenger RNA (mRNA) lengths, and abundance of transposable elements. Pedinomonas minor is only bounded by the plasma membrane, thriving in temporary habitats that frequently dry out. Gene family innovations and expansions and transcriptomic responses to abiotic stresses have shed light on adaptations of P. minor to its fluctuating environment. Horizontal gene transfers from bacteria and fungi have possibly contributed to the evolution of some of these traits. We identified a putative endogenization site of a nucleocytoplasmic large DNA virus and hypothesized that endogenous viral elements donated foreign genes to the host genome, their spread enhanced by transposable elements, located at gene boundaries in several of the expanded gene families.

Evolution of chloroplast retrograde signaling facilitates green plant adaptation to land.

Chloroplast retrograde signaling networks are vital for chloroplast biogenesis, operation, and signaling, including excess light and drought stress signaling. To date, retrograde signaling has been considered in the context of land plant adaptation, but not regarding the origin and evolution of signaling cascades linking chloroplast function to stomatal regulation. We show that key elements of the chloroplast retrograde signaling process, the nucleotide phosphatase (SAL1) and 3'-phosphoadenosine-5'-phosphate (PAP) metabolism, evolved in streptophyte algae-the algal ancestors of land plants. We discover an early evolution of SAL1-PAP chloroplast retrograde signaling in stomatal regulation based on conserved gene and protein structure, function, and enzyme activity and transit peptides of SAL1s in species including flowering plants, the fern Ceratopteris richardii, and the moss Physcomitrella patens Moreover, we demonstrate that PAP regulates stomatal closure via secondary messengers and ion transport in guard cells of these diverse lineages. The origin of stomata facilitated gas exchange in the earliest land plants. Our findings suggest that the conquest of land by plants was enabled by rapid response to drought stress through the deployment of an ancestral SAL1-PAP signaling pathway, intersecting with the core abscisic acid signaling in stomatal guard cells.

Rubisco and carbon-concentrating mechanism co-evolution across chlorophyte and streptophyte green algae.

Green algae expressing a carbon-concentrating mechanism (CCM) are usually associated with a Rubisco-containing micro-compartment, the pyrenoid. A link between the small subunit (SSU) of Rubisco and pyrenoid formation in Chlamydomonas reinhardtii has previously suggested that specific RbcS residues could explain pyrenoid occurrence in green algae. A phylogeny of RbcS was used to compare the protein sequence and CCM distribution across the green algae and positive selection in RbcS was estimated. For six streptophyte algae, Rubisco catalytic properties, affinity for CO2 uptake (K0.5 ), carbon isotope discrimination (δ13 C) and pyrenoid morphology were compared. The length of the ßA-ßB loop in RbcS provided a phylogenetic marker discriminating chlorophyte from streptophyte green algae. Rubisco kinetic properties in streptophyte algae have responded to the extent of inducible CCM activity, as indicated by changes in inorganic carbon uptake affinity, δ13 C and pyrenoid ultrastructure between high and low CO2 conditions for growth. We conclude that the Rubisco catalytic properties found in streptophyte algae have coevolved and reflect the strength of any CCM or degree of pyrenoid leakiness, and limitations to inorganic carbon in the aquatic habitat, whereas Rubisco in extant land plants reflects more recent selective pressures associated with improved diffusive supply of the terrestrial environment.

Biomass and phycobiliprotein production of Galdieria sulphuraria, immobilized on a twin-layer porous substrate photobioreactor.

The extremophile red alga Galdieria sulphuraria was successfully grown immobilized in a twin-layer porous substrate bioreactor (TL-PSBR). A maximal biomass growth rate of 10 g dry weight m-2 day-1 was measured at a photon fluence rate of 200 µmol photons m-2 s-1 with addition of 1% CO2 and a temperature of 34 °C. Under these conditions, a maximal biomass value of 232 g m-2 was attained after 33 days of growth. Phycobilin productivity, however, was highest at a lower photon fluence rate of 100 µmol photons m-2 s-1 and reached a phycobilin value of 14 g m-2, a phycobilin content in the biomass of 63 mg g-1 and a phycobilin growth rate of 0.28 g m-2 day-1 for phycocyanin and 0.23 g m-2 day-1 for allophycocyanin. Addition of CO2 was essential to enhance growth and phycobilin production in G. sulphuraria and further optimization of the cultivation process in the TL-PSBR appears possible using a multi-phase approach, higher growth temperatures and optimization of nutrient supply. It is concluded that autotrophic cultivation of G. sulphuraria in a TL-PSBR is an attractive alternative to suspension cultivation for phycobilin production and applications in bioremediation.

Genetic Analysis of Physcomitrella patens Identifies ABSCISIC ACID NON-RESPONSIVE, a Regulator of ABA Responses Unique to Basal Land Plants and Required for Desiccation Tolerance.

The anatomically simple plants that first colonized land must have acquired molecular and biochemical adaptations to drought stress. Abscisic acid (ABA) coordinates responses leading to desiccation tolerance in all land plants. We identified ABA nonresponsive mutants in the model bryophyte Physcomitrella patens and genotyped a segregating population to map and identify the ABA NON-RESPONSIVE (ANR) gene encoding a modular protein kinase comprising an N-terminal PAS domain, a central EDR domain, and a C-terminal MAPKKK-like domain. anr mutants fail to accumulate dehydration tolerance-associated gene products in response to drought, ABA, or osmotic stress and do not acquire ABA-dependent desiccation tolerance. The crystal structure of the PAS domain, determined to 1.7-Å resolution, shows a conserved PAS-fold that dimerizes through a weak dimerization interface. Targeted mutagenesis of a conserved tryptophan residue within the PAS domain generates plants with ABA nonresponsive growth and strongly attenuated ABA-responsive gene expression, whereas deleting this domain retains a fully ABA-responsive phenotype. ANR orthologs are found in early-diverging land plant lineages and aquatic algae but are absent from more recently diverged vascular plants. We propose that ANR genes represent an ancestral adaptation that enabled drought stress survival of the first terrestrial colonizers but were lost during land plant evolution.

Gene transfers from diverse bacteria compensate for reductive genome evolution in the chromatophore of Paulinella chromatophora.

Plastids, the photosynthetic organelles, originated >1 billion y ago via the endosymbiosis of a cyanobacterium. The resulting proliferation of primary producers fundamentally changed global ecology. Endosymbiotic gene transfer (EGT) from the intracellular cyanobacterium to the nucleus is widely recognized as a critical factor in the evolution of photosynthetic eukaryotes. The contribution of horizontal gene transfers (HGTs) from other bacteria to plastid establishment remains more controversial. A novel perspective on this issue is provided by the amoeba Paulinella chromatophora, which contains photosynthetic organelles (chromatophores) that are only 60-200 million years old. Chromatophore genome reduction entailed the loss of many biosynthetic pathways including those for numerous amino acids and cofactors. How the host cell compensates for these losses remains unknown, because the presence of bacteria in all available P. chromatophora cultures excluded elucidation of the full metabolic capacity and occurrence of HGT in this species. Here we generated a high-quality transcriptome and draft genome assembly from the first bacteria-free P. chromatophora culture to deduce rules that govern organelle integration into cellular metabolism. Our analyses revealed that nuclear and chromatophore gene inventories provide highly complementary functions. At least 229 nuclear genes were acquired via HGT from various bacteria, of which only 25% putatively arose through EGT from the chromatophore genome. Many HGT-derived bacterial genes encode proteins that fill gaps in critical chromatophore pathways/processes. Our results demonstrate a dominant role for HGT in compensating for organelle genome reduction and suggest that phagotrophy may be a major driver of HGT.

Functional and topological diversity of LOV domain photoreceptors.

Light-oxygen-voltage sensitive (LOV) flavoproteins are ubiquitous photoreceptors that mediate responses to environmental cues. Photosensory inputs are transduced into signaling outputs via structural rearrangements in sensor domains that consequently modulate the activity of an effector domain or multidomain clusters. Establishing the diversity in effector function and sensor-effector topology will inform what signaling mechanisms govern light-responsive behaviors across multiple kingdoms of life and how these signals are transduced. Here, we report the bioinformatics identification of over 6,700 candidate LOV domains (including over 4,000 previously unidentified sequences from plants and protists), and insights from their annotations for ontological function and structural arrangements. Motif analysis identified the sensors from ∼42 million ORFs, with strong statistical separation from other flavoproteins and non-LOV members of the structurally related Per-aryl hydrocarbon receptor nuclear translocator (ARNT)-Sim family. Conserved-domain analysis determined putative light-regulated function and multidomain topologies. We found that for certain effectors, sensor-effector linker length is discretized based on both phylogeny and the preservation of α-helical heptad repeats within an extended coiled-coil linker structure. This finding suggests that preserving sensor-effector orientation is a key determinant of linker length, in addition to ancestry, in LOV signaling structure-function. We found a surprisingly high prevalence of effectors with functions previously thought to be rare among LOV proteins, such as regulators of G protein signaling, and discovered several previously unidentified effectors, such as lipases. This work highlights the value of applying genomic and transcriptomic technologies to diverse organisms to capture the structural and functional variation in photosensory proteins that are vastly important in adaptation, photobiology, and optogenetics.

Phylogenomics Provides New Insights into Gains and Losses of Selenoproteins among Archaeplastida.

Selenoproteins that contain selenocysteine (Sec) are found in all kingdoms of life. Although they constitute a small proportion of the proteome, selenoproteins play essential roles in many organisms. In photosynthetic eukaryotes, selenoproteins have been found in algae but are missing in land plants (embryophytes). In this study, we explored the evolutionary dynamics of Sec incorporation by conveying a genomic search for the Sec machinery and selenoproteins across Archaeplastida. We identified a complete Sec machinery and variable sizes of selenoproteomes in the main algal lineages. However, the entire Sec machinery was missing in the Bangiophyceae-Florideophyceae clade (BV) of Rhodoplantae (red algae) and only partial machinery was found in three species of Archaeplastida, indicating parallel loss of Sec incorporation in different groups of algae. Further analysis of genome and transcriptome data suggests that all major lineages of streptophyte algae display a complete Sec machinery, although the number of selenoproteins is low in this group, especially in subaerial taxa. We conclude that selenoproteins tend to be lost in Archaeplastida upon adaptation to a subaerial or acidic environment. The high number of redox-active selenoproteins found in some bloom-forming marine microalgae may be related to defense against viral infections. Some of the selenoproteins in these organisms may have been gained by horizontal gene transfer from bacteria.

Identification of an algal xylan synthase indicates that there is functional orthology between algal and plant cell wall biosynthesis.

Insights into the evolution of plant cell walls have important implications for comprehending these diverse and abundant biological structures. In order to understand the evolving structure-function relationships of the plant cell wall, it is imperative to trace the origin of its different components. The present study is focused on plant 1,4-ß-xylan, tracing its evolutionary origin by genome and transcriptome mining followed by phylogenetic analysis, utilizing a large selection of plants and algae. It substantiates the findings by heterologous expression and biochemical characterization of a charophyte alga xylan synthase. Of the 12 known gene classes involved in 1,4-ß-xylan formation, XYS1/IRX10 in plants, IRX7, IRX8, IRX9, IRX14 and GUX occurred for the first time in charophyte algae. An XYS1/IRX10 ortholog from Klebsormidium flaccidum, designated K. flaccidumXYLAN SYNTHASE-1 (KfXYS1), possesses 1,4-ß-xylan synthase activity, and 1,4-ß-xylan occurs in the K. flaccidum cell wall. These data suggest that plant 1,4-ß-xylan originated in charophytes and shed light on the origin of one of the key cell wall innovations to occur in charophyte algae, facilitating terrestrialization and emergence of polysaccharide-based plant cell walls.

Insights into the Evolution of Hydroxyproline-Rich Glycoproteins from 1000 Plant Transcriptomes.

The carbohydrate-rich cell walls of land plants and algae have been the focus of much interest given the value of cell wall-based products to our current and future economies. Hydroxyproline-rich glycoproteins (HRGPs), a major group of wall glycoproteins, play important roles in plant growth and development, yet little is known about how they have evolved in parallel with the polysaccharide components of walls. We investigate the origins and evolution of the HRGP superfamily, which is commonly divided into three major multigene families: the arabinogalactan proteins (AGPs), extensins (EXTs), and proline-rich proteins. Using motif and amino acid bias, a newly developed bioinformatics pipeline, we identified HRGPs in sequences from the 1000 Plants transcriptome project (www.onekp.com). Our analyses provide new insights into the evolution of HRGPs across major evolutionary milestones, including the transition to land and the early radiation of angiosperms. Significantly, data mining reveals the origin of glycosylphosphatidylinositol (GPI)-anchored AGPs in green algae and a 3- to 4-fold increase in GPI-AGPs in liverworts and mosses. The first detection of cross-linking (CL)-EXTs is observed in bryophytes, which suggests that CL-EXTs arose though the juxtaposition of preexisting SPn EXT glycomotifs with refined Y-based motifs. We also detected the loss of CL-EXT in a few lineages, including the grass family (Poaceae), that have a cell wall composition distinct from other monocots and eudicots. A key challenge in HRGP research is tracking individual HRGPs throughout evolution. Using the 1000 Plants output, we were able to find putative orthologs of Arabidopsis pollen-specific GPI-AGPs in basal eudicots.

Molecular phylogeny, systematics, and revision of the type species of Lobomonas, L. francei (Volvocales, Chlorophyta) and closely related taxa.

In the present study, three new strains of the rare volvocalean green alga Lobomonas were isolated from field-collected samples, one from Sardinia (Italy) and two from Argentina, and comparatively studied. The Sardinian and one of the Argentinian strains were identified as Lobomonas francei, the type species of the genus, whereas the second Argentinian strain corresponded to L. panduriformis. Two additional nominal species of Lobomonas from culture collections (L. rostrata and L. sphaerica) were included in the analysis and shown to be morphologically and molecularly identical to the L. francei strains. The presence, number, and shapes of cell wall lobes, the diagnostic criterion of Lobomonas, were shown to be highly variable depending on the chemical composition of the culture medium used. The analyses by SEM gave evidence that the cell wall lobes in Lobomonas originate at the junctions of adjacent cell wall plates by extrusion of gelatinous material. The four L. francei strains had identical nrRNA gene sequences and differed by only one or two substitutions in the ITS1 + ITS2 sequences. In the phylogenetic analyses, L. francei and L. panduriformis were sister taxa; however, another nominal Lobomonas species (L. monstruosa) did not belong to this genus. Lobomonas, together with taxa designated as Vitreochlamys, Tetraspora, and Paulschulzia, formed a monophyletic group that in the combined analyses was sister to the "Chlamydomonas/Volvox-clade." Based on these results, Lobomonas was revised, the diagnosis of the type species emended, a lectotype and an epitype designated, and several taxa synonymized with the type species.

Algal ancestor of land plants was preadapted for symbiosis.

Colonization of land by plants was a major transition on Earth, but the developmental and genetic innovations required for this transition remain unknown. Physiological studies and the fossil record strongly suggest that the ability of the first land plants to form symbiotic associations with beneficial fungi was one of these critical innovations. In angiosperms, genes required for the perception and transduction of diffusible fungal signals for root colonization and for nutrient exchange have been characterized. However, the origin of these genes and their potential correlation with land colonization remain elusive. A comprehensive phylogenetic analysis of 259 transcriptomes and 10 green algal and basal land plant genomes, coupled with the characterization of the evolutionary path leading to the appearance of a key regulator, a calcium- and calmodulin-dependent protein kinase, showed that the symbiotic signaling pathway predated the first land plants. In contrast, downstream genes required for root colonization and their specific expression pattern probably appeared subsequent to the colonization of land. We conclude that the most recent common ancestor of extant land plants and green algae was preadapted for symbiotic associations. Subsequent improvement of this precursor stage in early land plants through rounds of gene duplication led to the acquisition of additional pathways and the ability to form a fully functional arbuscular mycorrhizal symbiosis.

Independent optical excitation of distinct neural populations.

Optogenetic tools enable examination of how specific cell types contribute to brain circuit functions. A long-standing question is whether it is possible to independently activate two distinct neural populations in mammalian brain tissue. Such a capability would enable the study of how different synapses or pathways interact to encode information in the brain. Here we describe two channelrhodopsins, Chronos and Chrimson, discovered through sequencing and physiological characterization of opsins from over 100 species of alga. Chrimson's excitation spectrum is red shifted by 45 nm relative to previous channelrhodopsins and can enable experiments in which red light is preferred. We show minimal visual system-mediated behavioral interference when using Chrimson in neurobehavioral studies in Drosophila melanogaster. Chronos has faster kinetics than previous channelrhodopsins yet is effectively more light sensitive. Together these two reagents enable two-color activation of neural spiking and downstream synaptic transmission in independent neural populations without detectable cross-talk in mouse brain slice.

All-optical electrophysiology in mammalian neurons using engineered microbial rhodopsins.

All-optical electrophysiology-spatially resolved simultaneous optical perturbation and measurement of membrane voltage-would open new vistas in neuroscience research. We evolved two archaerhodopsin-based voltage indicators, QuasAr1 and QuasAr2, which show improved brightness and voltage sensitivity, have microsecond response times and produce no photocurrent. We engineered a channelrhodopsin actuator, CheRiff, which shows high light sensitivity and rapid kinetics and is spectrally orthogonal to the QuasArs. A coexpression vector, Optopatch, enabled cross-talk-free genetically targeted all-optical electrophysiology. In cultured rat neurons, we combined Optopatch with patterned optical excitation to probe back-propagating action potentials (APs) in dendritic spines, synaptic transmission, subcellular microsecond-timescale details of AP propagation, and simultaneous firing of many neurons in a network. Optopatch measurements revealed homeostatic tuning of intrinsic excitability in human stem cell-derived neurons. In rat brain slices, Optopatch induced and reported APs and subthreshold events with high signal-to-noise ratios. The Optopatch platform enables high-throughput, spatially resolved electrophysiology without the use of conventional electrodes.

Growth and biomass productivity of Scenedesmus vacuolatus on a twin layer system and a comparison with other types of cultivations.

Scenedesmus is a genus of microalgae employed for several industrial uses. Industrial cultivations are performed in open ponds or in closed photobioreactors (PBRs). In the last years, a novel type of PBR based on immobilized microalgae has been developed termed porous substrate photobioreactors (PSBR) to achieve significant higher biomass density during cultivation in comparison to classical PBRs. This work presents a study of the growth of Scenedesmus vacuolatus in a Twin Layer System PSBR at different light intensities (600 µmol photons m-2 s-1 or 1000 µmol photons m-2 s-1), different types and concentrations of the nitrogen sources (nitrate or urea), and at two CO2 levels in the gas phase (2% or 0.04% v/v). The microalgal growth was followed by monitoring the attached biomass density as dry weight, the specific growth rate and pigment accumulation. The highest productivity (29 g m-2 d-1) was observed at a light intensity of 600 µmol photons m-2 s-1 and 2% CO2. The types and concentrations of nitrogen sources did not influence the biomass productivity. Instead, the higher light intensity of 1000 µmol photons m-2 s-1 and an ambient CO2 concentration (0.04%) resulted in a significant decrease of productivity to 18 and 10-12 g m-2 d-1, respectively. When compared to the performance of similar cultivation systems (15-30 g m-2 d-1), these results indicate that the Twin Layer cultivation System is a competitive technique for intensified microalgal cultivation in terms of productivity and, at the same time, biomass density.

Horizontal transfer of an adaptive chimeric photoreceptor from bryophytes to ferns.

Ferns are well known for their shade-dwelling habits. Their ability to thrive under low-light conditions has been linked to the evolution of a novel chimeric photoreceptor--neochrome--that fuses red-sensing phytochrome and blue-sensing phototropin modules into a single gene, thereby optimizing phototropic responses. Despite being implicated in facilitating the diversification of modern ferns, the origin of neochrome has remained a mystery. We present evidence for neochrome in hornworts (a bryophyte lineage) and demonstrate that ferns acquired neochrome from hornworts via horizontal gene transfer (HGT). Fern neochromes are nested within hornwort neochromes in our large-scale phylogenetic reconstructions of phototropin and phytochrome gene families. Divergence date estimates further support the HGT hypothesis, with fern and hornwort neochromes diverging 179 Mya, long after the split between the two plant lineages (at least 400 Mya). By analyzing the draft genome of the hornwort Anthoceros punctatus, we also discovered a previously unidentified phototropin gene that likely represents the ancestral lineage of the neochrome phototropin module. Thus, a neochrome originating in hornworts was transferred horizontally to ferns, where it may have played a significant role in the diversification of modern ferns.

Phylotranscriptomic analysis of the origin and early diversification of land plants.

Reconstructing the origin and evolution of land plants and their algal relatives is a fundamental problem in plant phylogenetics, and is essential for understanding how critical adaptations arose, including the embryo, vascular tissue, seeds, and flowers. Despite advances in molecular systematics, some hypotheses of relationships remain weakly resolved. Inferring deep phylogenies with bouts of rapid diversification can be problematic; however, genome-scale data should significantly increase the number of informative characters for analyses. Recent phylogenomic reconstructions focused on the major divergences of plants have resulted in promising but inconsistent results. One limitation is sparse taxon sampling, likely resulting from the difficulty and cost of data generation. To address this limitation, transcriptome data for 92 streptophyte taxa were generated and analyzed along with 11 published plant genome sequences. Phylogenetic reconstructions were conducted using up to 852 nuclear genes and 1,701,170 aligned sites. Sixty-nine analyses were performed to test the robustness of phylogenetic inferences to permutations of the data matrix or to phylogenetic method, including supermatrix, supertree, and coalescent-based approaches, maximum-likelihood and Bayesian methods, partitioned and unpartitioned analyses, and amino acid versus DNA alignments. Among other results, we find robust support for a sister-group relationship between land plants and one group of streptophyte green algae, the Zygnematophyceae. Strong and robust support for a clade comprising liverworts and mosses is inconsistent with a widely accepted view of early land plant evolution, and suggests that phylogenetic hypotheses used to understand the evolution of fundamental plant traits should be reevaluated.