Plasmodesmata dynamics in bryophyte model organisms: secondary formation and developmental modifications of structure and function.

MAIN CONCLUSION: Developing bryophytes differentially modify their plasmodesmata structure and function. Secondary plasmodesmata formation via twinning appears to be an ancestral trait. Plasmodesmata networks in hornwort sporophyte meristems resemble those of angiosperms. All land-plant taxa use plasmodesmata (PD) cell connections for symplasmic communication. In angiosperm development, PD networks undergo an extensive remodeling by structural and functional PD modifications, and by postcytokinetic formation of additional secondary PD (secPD). Since comparable information on PD dynamics is scarce for the embryophyte sister groups, we investigated maturating tissues of Anthoceros agrestis (hornwort), Physcomitrium patens (moss), and Marchantia polymorpha (liverwort). As in angiosperms, quantitative electron microscopy revealed secPD formation via twinning in gametophytes of all model bryophytes, which gives rise to laterally adjacent PD pairs or to complex branched PD. This finding suggests that PD twinning is an ancient evolutionary mechanism to adjust PD numbers during wall expansion. Moreover, all bryophyte gametophytes modify their existing PD via taxon-specific strategies resembling those of angiosperms. Development of type II-like PD morphotypes with enlarged diameters or formation of pit pairs might be required to maintain PD transport rates during wall thickening. Similar to angiosperm leaves, fluorescence redistribution after photobleaching revealed a considerable reduction of the PD permeability in maturating P. patens phyllids. In contrast to previous reports on monoplex meristems of bryophyte gametophytes with single initials, we observed targeted secPD formation in the multi-initial basal meristems of A. agrestis sporophytes. Their PD networks share typical features of multi-initial angiosperm meristems, which may hint at a putative homologous origin. We also discuss that monoplex and multi-initial meristems may require distinct types of PD networks, with or without secPD formation, to control maintenance of initial identity and positional signaling.

Contrasting pectin polymers in guard cell walls of Arabidopsis and the hornwort Phaeoceros reflect physiological differences.

BACKGROUND AND AIMS: In seed plants, stomata regulate CO2 acquisition and water relations via transpiration, while minimizing water loss. Walls of guard cells are strong yet flexible because they open and close the pore by changing shape over the substomatal cavity. Pectins are necessary for wall flexibility and proper stomata functioning. This study investigates the differences in pectin composition in guard cells of two taxa that represent key lineages of plants with stomata: Arabidopsis, an angiosperm with diurnal stomatal activity, and Phaeoceros, a bryophyte that lacks active stomatal movement. METHODS: Using immunolocalization techniques in transmission electron microscopy, this study describes and compares the localization of pectin molecule epitopes essential to stomata function in guard cell walls of Arabidopsis and Phaeoceros. KEY RESULTS: In Arabidopsis, unesterified homogalacturonans very strongly localize throughout guard cell walls and are interspersed with arabinan pectins, while methyl-esterified homogalacturonans are restricted to the exterior of the wall, the ledges and the junction with adjacent epidermal cells. In contrast, arabinans are absent in Phaeoceros, and both unesterified and methyl-esterified homogalacturonans localize throughout guard cell walls. CONCLUSIONS: Arabinans and unesterified homogalacturonans are required for wall flexibility, which is consistent with active regulation of pore opening in Arabidopsis stomata. In contrast, the lack of arabinans and high levels of methyl-esterified homogalacturonans in guard cell walls of Phaeoceros are congruent with the inability of hornwort stomata to open and close with environmental change. Comparisons across groups demonstrate that variations in guard cell wall composition reflect different physiological activity of stomata in land plants.

Hornwort stomata do not respond actively to exogenous and environmental cues.

Backgrounds and Aims: Because stomata in bryophytes occur on sporangia, they are subject to different developmental and evolutionary constraints from those on leaves of tracheophytes. No conclusive experimental evidence exists on the responses of hornwort stomata to exogenous stimulation. Methods: Responses of hornwort stomata to abscisic acid (ABA), desiccation, darkness and plasmolysis were compared with those in tracheophyte leaves. Potassium ion concentrations in the guard cells and adjacent cells were analysed by X-ray microanalysis, and the ontogeny of the sporophytic intercellular spaces was compared with those of tracheophytes by cryo-scanning electron microscopy. Key Results: The apertures in hornwort stomata open early in development and thereafter remain open. In hornworts, the experimental treatments, based on measurements of >9000 stomata, produced only a slight reduction in aperture dimensions after desiccation and plasmolysis, and no changes following ABA treatments and darkness. In tracheophytes, all these treatments resulted in complete stomatal closure. Potassium concentrations are similar in hornwort guard cells and epidermal cells under all treatments at all times. The small changes in hornwort stomatal dimensions in response to desiccation and plasmolysis are probably mechanical and/or stress responses of all the epidermal and spongy chlorophyllose cells, affecting the guard cells. In contrast to their nascent gas-filled counterparts across tracheophytes, sporophytic intercellular spaces in hornworts are initially liquid filled. Conclusions: Our experiments demonstrate a lack of physiological regulation of opening and closing of stomata in hornworts compared with tracheophytes, and support accumulating developmental and structural evidence that stomata in hornworts are primarily involved in sporophyte desiccation and spore discharge rather than the regulation of photosynthesis-related gaseous exchange. Our results run counter to the notion of the early acquisition of active control of stomatal movements in bryophytes as proposed from previous experiments on mosses.

Activity of Redox Enzymes in the Thallus of Anthoceros natalensis.

Anthocerotophyta (hornworts) belong to a group of ancient nonvascular plants and originate from a common ancestor with contemporary vascular plants. Hornworts represent a unique model for investigating mechanisms of formation of stress resistance in higher plants due to their high tolerance to the action of adverse environmental factors. In this work, we demonstrate that the thallus of Anthoceros natalensis exhibits high redox activity changing under stress. Dehydration of the thallus is accompanied by the decrease in activities of intracellular peroxidases, DOPA-peroxidases, and tyrosinases, while catalase activity increases. Subsequent rehydration results in the increase in peroxidase and catalase activities. Kinetic features of peroxidases and tyrosinases were characterized as well as the peroxidase isoenzyme composition of different fractions of the hornwort cell wall proteins. It was shown that the hornwort peroxidases are functionally similar to peroxidases of higher vascular plants including their ability to form superoxide anion-radical. The biochemical mechanism was elucidated, supporting the possible participation of peroxidases in the formation of reactive oxygen species (ROS) via substrate-substrate interactions in the hornwort thallus. It has been suggested that the ROS formation by peroxidases is an evolutionarily ancient process that emerged as a protective mechanism for enhancing adaptive responses of higher land plants and their adaptation to changing environmental conditions and successful colonization of various ecological niches.

Correlates of monoicy and dioicy in hornworts, the apparent sister group to vascular plants.

BACKGROUND: Whether male and female gametes are produced by single or separate individuals shapes plant mating and hence patterns of genetic diversity among and within populations. Haploid-dominant plants ("bryophytes": liverworts, mosses and hornworts) can have unisexual (dioicous) or bisexual (monoicous) gametophytes, and today, 68% of liverwort species, 57% of moss species, and 40% of hornwort species are dioicous. The transitions between the two sexual systems and possible correlations with other traits have been studied in liverworts and mosses, but not hornworts. Here we use a phylogeny for 98 of the 200 species of hornworts, the sister group to vascular plants, representing roughly equal proportions of all monoicous and all dioicous species, to test whether transitions in sexual systems are predominantly from monoicy to dioicy as might be expected based on studies of mosses. We further investigate possible correlations between sexual system and spore size, antheridium number, ploidy level, and diversification rate, with character selection partly based on findings in mosses and liverworts. RESULTS: Hornworts underwent numerous transitions between monoicy and dioicy. The transition rate from dioicy to monoicy was 2× higher than in the opposite direction, but monoicous groups have higher extinction rates; diversification rates do not correlate with sexual system. A correlation important in mosses, that between monoicy and polyploidy, apparently plays a small role: of 20 species with chromosome counts, only one is polyploid, the monoicous Anthoceros punctatus. A contingency test revealed that transitions to dioicy were more likely in species with small spores, supporting the hypothesis that small but numerous spores may be advantageous for dioicous species that depend on dense carpets of gametophytes for reproductive assurance. However, we found no evidence for increased antheridium-per-chamber numbers in dioicous species. CONCLUSIONS: Sexual systems in hornworts are labile, and the higher number of extant monoicous species (60%) may be largely due to frequent transitions to monoicy.

Fungal symbioses in hornworts: a chequered history.

Hornworts are considered the sister group to vascular plants, but their fungal associations remain largely unexplored. The ancestral symbiotic condition for all plants is, nonetheless, widely assumed to be arbuscular mycorrhizal with Glomeromycota fungi. Owing to a recent report of other fungi in some non-vascular plants, here we investigate the fungi associated with diverse hornworts worldwide, using electron microscopy and molecular phylogenetics. We found that both Glomeromycota and Mucoromycotina fungi can form symbioses with most hornworts, often simultaneously. This discovery indicates that ancient terrestrial plants relied on a wider and more versatile symbiotic repertoire than previously thought, and it highlights the so far unappreciated ecological and evolutionary role of Mucoromycotina fungi.

Ceratophyllum demersum L. and Potamogeton alpinus Balb. from Iset' river, Ural region, Russia differ in adaptive strategies to heavy metals exposure--a comparative study.

We examined the uptake of five heavy metals (Cu, Fe, Ni, Zn, and Mn) in Ceratophyllum demersum L. (hornwort) and Potamogeton alpinus Balb. (pondweed) from Iset' river, Ural region, Russia. This study was conducted in a territory that is highly urbanized where the surface waters are contaminated by a wide spectrum of pollutants. The environmental situation in this territory drastically deteriorated due to anthropogenic activity. The water quality in most of the water bodies in the Ural region is rather poor. In a comparative study of C. demersum and P. alpinus, differential accumulation pattern was noted for heavy metals (HMs). Higher amounts of HMs accumulated in C. demersum compared to P. alpinus. Also it was shown that in leaves of C. demersum there were high amount of total phosphorus, nitrogen, organics acids and ash; high activity of guaiacol peroxidase; high content of nonenzymatic antioxidants viz., flavonoids, ascorbate, glutathione and proline; high amount of thiols (soluble and membrane bound) compared to P. alpinus.