3D‐reconstructions of zygospores in Zygnema vaginatum (Charophyta) reveal details of cell wall formation, suggesting adaptations to extreme habitats

Abstract

Abstract The streptophyte green algal class Zygnematophyceae is the immediate sister lineage to land plants. Their special form of sexual reproduction via conjugation might have played a key role during terrestrialization. Thus, studying Zygnematophyceae and conjugation is crucial for understanding the conquest of land. Moreover, sexual reproduction features are important for species determination. We present a phylogenetic analysis of a field‐sampled Zygnema strain and analyze its conjugation process and zygospore morphology, both at the micro‐ and nanoscale, including 3D‐reconstructions of the zygospore architecture. Vegetative filament size (26.18 ± 1.07 μm) and reproductive features allowed morphological determination of Zygnema vaginatum, which was combined with molecular analyses based on rbcL sequencing. Transmission electron microscopy (TEM) depicted a thin cell wall in young zygospores, while mature cells exhibited a tripartite wall, including a massive and sculptured mesospore. During development, cytological reorganizations were visualized by focused ion beam scanning electron microscopy (FIB‐SEM). Pyrenoids were reorganized, and the gyroid cubic central thylakoid membranes, as well as the surrounding starch granules, degraded (starch granule volume: 3.58 ± 2.35 μm3 in young cells; 0.68 ± 0.74 μm3 at an intermediate stage of zygospore maturation). Additionally, lipid droplets (LDs) changed drastically in shape and abundance during zygospore maturation (LD/cell volume: 11.77% in young cells; 8.79% in intermediate cells, 19.45% in old cells). In summary, we provide the first TEM images and 3D‐reconstructions of Zygnema zygospores, giving insights into the physiological processes involved in their maturation. These observations help to understand mechanisms that facilitated the transition from water to land in Zygnematophyceae.

Plant phenotyping relevance

FIB-SEM/TEMによる3D画像取得と再構成が研究の中心で、接合胞子の形態・細胞内構造・体積変化を定量的に評価しているため、画像ベースの植物表現型計測として含める。

Code and data availability

The paper's FIB-SEM image stacks of Zygnema vaginatum zygospores (the raw imaging data underlying the 3D reconstructions) are publicly deposited as supplementary Movies S2–S4 on figshare. No author analysis code or trained models are explicitly deposited; other supporting data are available only upon request.