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The photosynthetic and anatomical responses of bryophytes to changes in gravity will provide crucial information for estimating how these plant traits evolved to adapt to changes in gravity in land plant history. We performed long-term hypergravity experiments at 10 g for 4 and 8 weeks using the moss Physcomitrella patens with two centrifuges equipped with lighting systems that enable long-term plant growth under hypergravity with irradiance. The aims of this study are (1) to quantify changes in the anatomy and morphology of P. patens, and (2) to analyze the post-effects of hypergravity on photosynthesis by P. patens in relation to these changes. We measured photosynthesis by P. patens for a population of gametophores (e.g., canopy) in Petri dishes and plant culture boxes. Gametophore numbers increased by 9% for a canopy of P. patens, with 24-27% increases in chloroplast sizes (diameter and thickness) in leaf cells. In a canopy of P. patens, the area-based photosynthesis rate ( A ) was increased by 57% at 10 g. The increase observed in A was associated with greater plant numbers and chloroplast sizes, both of which involved enhanced CO diffusion from the atmosphere to chloroplasts in the canopies of P. patens. These results suggest that changes in gravity are important environmental stimuli to induce changes in plant growth and photosynthesis by P. patens, in which an alteration in chloroplast size is one of the key traits. We are now planning an ISS experiment to investigate the responses of P. patens to microgravity. [ABSTRACT FROM AUTHOR]

In bryophytes (i.e., mosses, liverworts, and hornworts), extant representatives of early land plants, plasmodesmata have been described in a wide range of tissues. Although their contribution to bryophyte morphogenesis remains largely unexplored, several recent studies have suggested that the deposition of callose around plasmodesmata might regulate developmental and physiological responses in mosses. In this chapter, we provide a protocol to image and quantify callose levels in the filamentous body of the model moss Physcomitrium (Physcomitrella) patens and discuss possible alternatives and pitfalls. More generally, this protocol establishes a framework to explore the distribution of callose in other bryophytes., (© 2022. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

An important approach to investigate intercellular connectivity via plasmodesmata is to visualize and track the movement of fluorescent proteins between cells. The intercellular connectivity is largely controlled by the size exclusion limit of the pores. Over the past few decades, the technique to observe and analyze intercellular movement of a fluorescent protein has been developed mainly in angiosperms such as Arabidopsis thaliana. We recently applied the corresponding system to track the intercellular movement of the fluorescent protein Dendra2 in the moss Physcomitrium (Physcomitrella) patens. The protonemal tissues are particularly suited for observation of the intercellular movement due to the simple organization. Here, we describe a protocol suitable for the analysis of Dendra2 movement between cells in P. patens., (© 2022. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Shoot phototropism benefits growth and metabolism in land plants by enabling them to position their photosynthetic organs in favorable light conditions. Nonvascular land plants, like the ancestors of modern mosses, are believed to have been among the first plants to occupy the land. To understand the evolutional history of shoot phototropism in land plants, we have established a system for experimentally studying phototropism in gametophores of the moss Physcomitrella patens. Here we will describe the key points in our system, including obtaining etiolated gametophores, the light sources used for inducing bending, and the methods for evaluation of phototropic responses.