Autophagy is a catabolic process for bulk and selective degradation of cytoplasmic components in the vacuole/lysosome. In Saccharomyces cerevisiae, ATG genes were identified as essential genes for autophagy, and most ATG genes are highly conserved among eukaryotes, including plants. Although reverse genetic analyses have revealed that autophagy is involved in responses to abiotic and biotic stresses in land plants, our knowledge of its molecular mechanism remains limited. This limitation is partly because of the multiplication of some ATG genes, including ATG8 , in widely used model plants such as Arabidopsis thaliana , which adds complexity to functional studies. Furthermore, due to limited information on the composition and functions of the ATG genes in basal land plants and charophytes, it remains unclear whether multiplication of ATG genes is associated with neofunctionalization of these genes. To gain insight into the diversification of ATG genes during plant evolution, we compared the composition of ATG genes in plants with a special focus on a liverwort and two charophytes, which have not previously been analyzed. Our results showed that the liverwort Marchantia polymorpha and the charophytes Klebsormidium nitens and Chara braunii harbor fundamental sets of ATG genes with low redundancy compared with those of A. thaliana and the moss Physcomitrella patens , suggesting that multiplication of ATG genes occurred during land plant evolution. We also attempted to establish an experimental system for analyzing autophagy in M. polymorpha . We generated transgenic plants expressing fluorescently tagged MpATG8 to observe its dynamics in M. polymorpha and produced autophagy-defective mutants by genome editing using the CRISPR/Cas9 system. These tools allowed us to demonstrate that MpATG8 is transported into the vacuole in an MpATG2-, MpATG5-, and MpATG7-dependent manner, suggesting that fluorescently tagged MpATG8 can be used as an autophagosome marker in M. polymorpha . M. polymorpha can provide a powerful system for studying the mechanisms and evolution of autophagy in plants.

Gametogenesis is an essential biological event for sexual reproduction in various organisms. Bryophytes and some other plants employ motile sperms (spermatozoids) as male gametes, which self-locomote to the egg cells to accomplish fertilization. Spermatozoids of bryophytes harbor distinctive morphological characteristics, including the cell body with a helical slender shape and two motile flagella at the anterior edge. During transformation from a spermatid to spermatozoid (spermiogenesis), the shape and cellular contents of spermatids are dynamically reorganized. However, how each organelle is reorganized during plant spermiogenesis remains obscure. In this study, we classified the developmental processes during spermiogenesis in the liverwort Marchantia polymorpha according to cellular and nuclear shapes and flagella development. We then examined the remodeling of microtubules and reorganization of endomembrane organelles during spermiogenesis. The results indicate that the state of post-translational modification of tubulin is dynamically changed during the formation of the flagella and spline, and the plasma membrane and endomembrane organelles are drastically reorganized in a precisely regulated manner during spermiogenesis. These findings are expected to provide useful indexes to classify developmental and subcellular processes of spermiogenesis in bryophytes.Summary statementWe classified developmental processes of spermatozoids into 1 + 5 stages and characterized remodeling of organelles and microtubules during spermiogenesis in the liverwort Marchantia polymorpha.

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Autophagy is a highly conserved system for degrading and recycling cytoplasmic components. The identification of autophagy-related (ATG) genes, required for autophagosome formation, has led to numerous studies using atg mutants. These studies have revealed the physiological significance of autophagy in various functions of diverse organisms. In land plants, autophagy is required for higher-order functions such as stress responses and development. Although defective autophagy does not result in any marked defect in the reproductive processes of Arabidopsis thaliana under laboratory conditions, several studies have shown that autophagy plays a pivotal role in male reproduction in several land plants. In this review, we aim to summarize information on the role of autophagy in male reproductive processes in land plants.