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[...]pioneers of the P. patens plant model system envisaged using classical forward genetics for the dissection of developmental processes.Since protonemal tissue, the first tissue to be formed from a germinating moss spore, can be propagated vegetatively, the culture of mutants that were abnormal even at an early stage of development presented no problems—although when such mutants were sterile, orthodox genetic analysis was not possible.Forward genetic dissection of a developmental process using somatic hybridisation is compromised when the causal mutations are dominant [10].[...]it was another three decades before a candidate-gene strategy showed that 8 of 17 chemically-induced nar (1-naphthalene acetic acid and indole-3-acetic acid resistant) strains harbour mutations distributed among the three P. patens homologues of Arabidopsis Aux/IAA (auxin/indole-3-acetic acid) genes [11].[...]forward genetic analysis in P. patens was unattractive to most researchers, and their disaffection was exacerbated by the paucity of markers required for effective mapping of new mutations that existed prior to the generation of a genetic linkage map based on numerous molecular markers and anchored to the genome sequence [12].The corresponding gene, designated ARK (ABA and abiotic stress-responsive Raf-like kinase), encodes a B3 Raf-like mitogen-activated protein (MAP) kinase kinase kinase involved in integrating ABA and osmotic signals upstream from sucrose nonfermenting 1 (SNF1)-related protein kinase 2, known to be a central regulator of stress signaling in plants [15].

Sporopollenin is the main constituent of the exine layer of spore and pollen walls. The anther-specific chalcone synthase-like (ASCL) enzyme of Physcomitrella patens, PpASCL, has previously been implicated in the biosynthesis of sporopollenin, the main constituent of exine and perine, the two outermost layers of the moss spore cell wall. We made targeted knockouts of the corresponding gene, PpASCL, and phenotypically characterized ascl sporophytes and spores at different developmental stages. Ascl plants developed normally until late in sporophytic development, when the spores produced were structurally aberrant and inviable. The development of the ascl spore cell wall appeared to be arrested early in microspore development, resulting in small, collapsed spores with altered surface morphology. The typical stratification of the spore cell wall was absent with only an abnormal perine recognisable above an amorphous layer possibly representing remnants of compromised intine and/or exine. Equivalent resistance of the spore walls of ascl mutants and the control strain to acetolysis suggests the presence of chemically inert, defective sporopollenin in the mutants. Anatomical abnormalities of late-stage ascl sporophytes include a persistent large columella and an air space incompletely filled with spores. Our results indicate that the evolutionarily conserved PpASCL gene is needed for proper construction of the spore wall and for normal maturation and viability of moss spores.

Vascular plants appeared ~410 million years ago, then diverged into several lineages of which only two survive: the euphyllophytes (ferns and seed plants) and the lycophytes. We report here the genome sequence of the lycophyte Selaginella moellendorffii (Selaginella), the first nonseed vascular plant genome reported. By comparing gene content in evolutionarily diverse taxa, we found that the transition from a gametophyte- to a sporophyte-dominated life cycle required far fewer new genes than the transition from a nonseed vascular to a flowering plant, whereas secondary metabolic genes expanded extensively and in parallel in the lycophyte and angiosperm lineages. Selaginella differs in posttranscriptional gene regulation, including small RNA regulation of repetitive elements, an absence of the trans-acting small interfering RNA pathway, and extensive RNA editing of organellar genes.

Type III polyketide synthases (PKSs) have co-evolved with terrestrial plants such that each taxon can generate a characteristic collection of polyketides, fine-tuned to its needs. 2'-Oxoalkylresorcinol synthase from Physcomitrella patens (PpORS) is basal to all plant type III PKSs in phylogenetic trees and may closely resemble their most recent common ancestor. To gain insight into the roles that ancestral plant type III PKSs might have played during early land plant evolution, we constructed and phenotypically characterized targeted knockouts of PpORS. Ors gametophores, unless submerged in water while they were developing, displayed various leaf malformations that included grossly misshapen leaves, missing or abnormal midribs, multicellular protuberances and localized necrosis. Ors leaves, particularly abnormal ones, showed increased permeability to the hydrophilic dye, toluidine blue. Ors gametophores lost water faster and were more susceptible to dehydration than those the control strain. Our findings are consistent with ors leaves possessing a partially defective cuticle and implicate PpORS in synthesis of the intact cuticle. PpORS orthologs are present in a few moss species but have not been found in other plants. However, conceivably an ancestral ORS in early land plants may have contributed to their protection from dehydration.

Although the evolution of spores was critical to the diversification of plants on land, sporogenesis is incompletely characterized for model plants such as Physcomitrium patens . In this study, the complete process of P. patens sporogenesis is detailed from capsule expansion to mature spore formation, with emphasis on the construction of the complex spore wall and proximal aperture. Both diploid (sporophytic) and haploid (spores) cells contribute to the development and maturation of spores. During capsule expansion, the diploid cells of the capsule, including spore mother cells (SMCs), inner capsule wall layer (spore sac), and columella, contribute a locular fibrillar matrix that contains the machinery and nutrients for spore ontogeny. Nascent spores are enclosed in a second matrix that is surrounded by a thin SMC wall and suspended in the locular material. As they expand and separate, a band of exine is produced external to a thin foundation layer of tripartite lamellae. Dense globules assemble evenly throughout the locule, and these are incorporated progressively onto the spore surface to form the perine external to the exine. On the distal spore surface, the intine forms internally, while the spiny perine ornamentation is assembled. The exine is at least partially extrasporal in origin, while the perine is derived exclusively from outside the spore. Across the proximal surface of the polar spores, an aperture begins formation at the onset of spore development and consists of an expanded intine, an annulus, and a central pad with radiating fibers. This complex aperture is elastic and enables the proximal spore surface to cycle between being compressed (concave) and expanded (rounded). In addition to providing a site for water intake and germination, the elastic aperture is likely involved in desiccation tolerance. Based on the current phylogenies, the ancestral plant spore contained an aperture, exine, intine, and perine. The reductive evolution of liverwort and hornwort spores entailed the loss of perine in both groups and the aperture in liverworts. This research serves as the foundation for comparisons with other plant groups and for future studies of the developmental genetics and evolution of spores across plants.

Enzymes of the chalcone synthase (CHS) superfamily catalyze the production of a variety of secondary metabolites in bacteria, fungi and plants. Some of these metabolites have played important roles during the early evolution of land plants by providing protection from various environmental assaults including UV irradiation. The genome of the moss, Physcomitrella patens, contains at least 17 putative CHS superfamily genes. Three of these genes (PpCHS2b, PpCHS3 and PpCHS5) exist in multiple copies and all have corresponding ESTs. PpCHS11 and probably also PpCHS9 encode non-CHS enzymes, while PpCHS10 appears to be an ortholog of plant genes encoding anther-specific CHS-like enzymes. It was inferred from the genomic locations of genes comprising it that the moss CHS superfamily expanded through tandem and segmental duplication events. Inferred exon-intron architectures and results from phylogenetic analysis of representative CHS superfamily genes of P. patens and other plants showed that intron gain and loss occurred several times during evolution of this gene superfamily. A high proportion of P. patens CHS genes (7 of 14 genes for which the full sequence is known and probably 3 additional genes) are intronless, prompting speculation that CHS gene duplication via retrotransposition has occurred at least twice in the moss lineage. Analyses of sequence similarities, catalytic motifs and EST data indicated that a surprisingly large number (as many as 13) of the moss CHS superfamily genes probably encode active CHS. EST distribution data and different light responsiveness observed with selected genes provide evidence for their differential regulation. Observed diversity within the moss CHS superfamily and amenability to gene manipulation make Physcomitrella a highly suitable model system for studying expansion and functional diversification of the plant CHS superfamily of genes.

KEY MESSAGE : The Physcomitrella pseudochromosomal genome assembly revealed previously invisible synteny enabling realisation of the full potential of shared synteny as a tool for probing evolution of this plant’s MADS-box gene family. Assembly of the sequenced genome of Physcomitrella patens into 27 mega-scaffolds (pseudochromosomes) has confirmed the major predictions of our earlier model of expansion of the MADS-box gene family in the Physcomitrella lineage. Additionally, microsynteny has been conserved in the immediate vicinity of some recent duplicates of MADS-box genes. However, comparison of non-syntenic MIKC MADS-box genes and neighbouring genes indicates that chromosomal rearrangements and/or sequence degeneration have destroyed shared synteny over longer distances (macrosynteny) around MADS-box genes despite subsets comprising two or three MIKC genes having remained syntenic. In contrast, half of the type I MADS-box genes have been transposed creating new syntenic relations with MIKC genes. This implies that conservation of ancient ancestral synteny of MIKC genes and of more recently acquired synteny of type I and MIKC genes may be selectively advantageous. Our revised model predicts the birth rate of MIKC genes in Physcomitrella is higher than that of type I genes. However, this difference is attributable to an early tandem duplication and an early segmental duplication of MIKC genes prior to the two polyploidisations that account for most of the expansion of the MADS-box gene family in Physcomitrella. Furthermore, this early segmental duplication spawned two chromosomal lineages: one with a MIKC C gene, belonging to the PPM2 clade, in close proximity to one or a pair of MIKC* genes and another with a MIKC C gene, belonging to the PpMADS-S clade, characterised by greater separation from syntenic MIKC* genes. Our model has evolutionary implications for the Physcomitrella karyotype.

The First Automotive CFD Prediction Workshop was held in December 2019 at St Anne’s College at the University of Oxford with the aim to assess the ability of a broad range of computational fluid dynamics (CFD) methods to predict the flow over realistic automotive geometries. Here, results from 53 simulation data sets from 9 separate groups are analyzed for the open-source automotive DrivAer model (in the fastback and estate variants). The represented CFD approaches include Reynolds-averaged Navier-Stokes (RANS) approaches with a broad range of turbulence models, as well as scale-resolving approaches such as wall-modelled large-eddy simulation (WMLES) and hybrid RANS-LES methods (HRLM). A range of CFD codes was used, including commercial, academic, and open source. Compared to the two experimental data points, there was a large spread of CFD results. The difference between drag predictions among HRLM and RANS methods is significant, with an even larger mismatch for lift. The differences are found to be more significant for the estate geometry than for the fastback, with the former having larger areas of flow separation. In general it is found that the spread of HRLM is smaller than those for RANS approaches, with HRLM grouping closer to the range of experimental values. However, for HRLM, there is a systematic underprediction of the front lift coefficient that is irrespective of the mesh, turbulence model, and CFD code. Given that the majority of participants used the same mesh and boundary conditions, and in some cases the same CFD code, it suggests that also user choices around numerical schemes, convergence, and turbulence model coefficients may have a sizable impact, which was not possible to fully control in this first workshop. It is worth noting as well that the CFD simulations were conducted in a free-air environment and did not model the wind-tunnel geometry itself, which may also be an area requiring further study. The DrivAer model exhibits numerous complex flow physics, i.e., laminar/turbulent separation, diffusion of momentum in turbulent shear layers, and interactions of turbulent wakes with boundary layers. However, the nature of a community-driven workshop and the lack of extensive experimental data means that this article can only report the current state of the art and serve as a reference for future workshops and a springboard for more focused future studies where topics can be explored in greater detail.

We report the draft genome sequence of the model moss Physcomitrella patens and compare its features with those of flowering plants, from which it is separated by more than 400 million years, and unicellular aquatic algae. This comparison reveals genomic changes concomitant with the evolutionary movement to land, including a general increase in gene family complexity; loss of genes associated with aquatic environments (e.g., flagellar arms); acquisition of genes for tolerating terrestrial stresses (e.g., variation in temperature and water availability); and the development of the auxin and abscisic acid signaling pathways for coordinating multicellular growth and dehydration response. The Physcomitrella genome provides a resource for phylogenetic inferences about gene function and for experimental analysis of plant processes through this plant's unique facility for reverse genetics.

An imperiled population of endemic North American Burbot (Lota lota maculosa) inhabiting the lower Kootenai River in a transboundary region of Idaho and British Columbia struggles with recruitment failures linked to incompatibility between early life history patterns and upstream hydropower operations at Libby Dam in Montana. The loss of both river ice in winter and a flood pulse in spring has led to an unnatural post-dam thermal regime in spawning and early rearing habitats of Burbot. A series of laboratory experiments and river temperature modeling efforts were carried out to determine if simulated changes to the river thermal regime impact spawning, embryo development, and larval rearing. Contemporary warming in the post-dam regime between December−January does not appear to preclude functional gametogenesis, as river-adapted hatchery fish spawned viable gametes in February. However, elevated water temperatures of 4°C and 6°C shorten the spawning period to potentially coincide with suboptimal conditions for normal embryo development. Cold incubation temperatures are crucial during early embryogenesis, but the post-dam regime is often >3°C—and can be as warm as 5°C—which causes severe deformity and mortality of embryos. Later stages of embryonic development show reasonable tolerance to gradual warming and moderate thermal flux common in late winter and early spring. Larvae were observed inflating their swim bladders, foraging on rotifers, and exhibiting high survival through a critical period in waters <6°C, which is the typical post-dam condition in putative nursery habitats in April. However, such cold regimes also prolong resting behavior of yolk-sac larvae, delay swim bladder inflation, reduce feeding, slow growth, and diminish larvae performance. These important aspects of larval ecophysiology during a critical period of Burbot ontogeny could increase the risk of cumulative mortality and negatively impact recruitment. It was difficult to resolve a modeling solution of post-dam operations that better mimics pre-dam river temperatures in winter because daily changes in regional climate now explain most of the variance in river temperature between December−April. The unpopular option of restricting discharge from Libby Dam in December and January might return ice cover to the downstream meander reach, but a more realistic mitigation tool for Burbot conservation is restoring tributary, side-channel, and off-channel habitats potentially supportive of early life stages. Conservation aquaculture is rebuilding the population and improving opportunities for discovering the recruitment bottleneck(s). The focus of restoration, monitoring, and evaluation work should now be on improving embryo survival in tributaries and larvae survival through the critical period in nursery habitats of the meander reach. An unwavering commitment to hatcheries will also be a key management tool for combating the deleterious impacts of global warming on altered and natural spawning habitats of Burbot.