Research

Plant "innovation" - Reproductive mechanisms that support plant prosperity

Flowering plants are thought to have emerged in the early Cretaceous period and prospered explosively, reaching over 350,000 species, accounting for approximately 80% of all living terrestrial plants today. One of the angiosperm's "innovation" that supported this prosperity was the birth of sexual reproduction through pollen. To ensure the reproduction of offspring on dry land, female gametes evolved to be enclosed deep in the tissues of the pistil, and male gametes evolved to be enclosed in cells with a hard cell wall. These male cells are called pollen.

Pollen is an organ common to all angiosperms. Elucidating the process from its development to fertilization is important not only for understanding the fundamental principles of life and its evolutionary significance, but also for solving problems in the fields of agriculture, food, and the environment. However, because reproduction in angiosperms occurs deep within the flower, observation and analysis are difficult, and many mysteries regarding its mechanism remain.

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Reproductive process of angiosperms

Pollen is the male gametophyte of angiosperms, and each pollen grain contains two sperm cells, which are male gametes. Because the sperm cells of angiosperms lost their flagella during evolution, they are unable to move independently to reach the female. Therefore, pollen has developed a mechanism to extend a tubular cell called a pollen tube, which delivers the sperm cells inside to the ovule. The pollen tube extends through the pistil and reaches the ovule, delivering the sperm cells to the egg cell and central cell, resulting in the double fertilization.

These processes are unique reproductive methods that plants have evolved to ensure they can produce offspring and thrive on dry land.

Pollen development and differentiation

After meiosis, a single cell microspore undergoes asymmetric division and differentiates into a large vegetative cell and a small generative cell. However, if some stresses cause symmetric division, both cells will become vegetative cells, resulting in pollen that cannot be fertilized.

We have established the live imaging system for in vitro pollen development of Nicotiana benthamiana pollen (Mizuta 2023). We have also established a method for transiently introducing genes into microspores (Nagahara et al. 2021) to rapidly analyze intracellular structures and gene functions.

Analysis of the cell cycle and cytoskeleton dynamics revealed that actin filaments control the polarized movement of the nucleus before pollen mitosis 1, and the differentiation of vegetative cells after pollen mitosis 1 (Mizuta et al. 2025). Furthermore, by combining genetic and omics analyses, we are studying how a single cell produces two different cells and the molecular mechanisms by which cells become fertile.

Pollen tube guidance and polyspermy block

In Arabidopsis pistil, each ovule attract a single pollen tube and rejects others, a phenomenon known as one-to-one pollen tube attraction, i.e., polyspermy block. This allows more seeds to be produced with less pollen. One-to-one pollen tube attraction requires male-female communication. In recent years, several factors, including peptides and receptors, have been identified (Mizuta and Higashiyama 2018). However, because pollen tube attraction occurs deep within the pistil, live-analysis is difficult, hindering progress in research.

Using two-photon excitation microscopy, we have established a method for making the deep pistil transparent (Kurihara et al. 2015 ; Mizuta and Tsuda 2018 ; Niimi et al. 2022) (Figure A below) and a method for live imaging (Mizuta et al. 2015 ; Mizuta 2021) (Figure B below). Using mutants, we have revealed that the signal derived from the maternal somatic cell, and proteins on the plasma membranes of the female gametophyte, as well as the number and position of pollen tubes, are important for one-to-one pollen tube attraction. Our study revealed multi-steps and spatiotemporal mechanism for controlling pollen tube attraction and polyspermy block (Mizuta et al. 2024) (Figure C below). We aim to elucidate the mechanisms of plant sexual reproduction acquired by angiosperms during evolution.

“Design” of plant reproductive engineering

Pollen is easier to manipulate than female gametophytes. It also serves as a "vector" cell, carrying the male genome, which is passed on to the next generation. We are developing methods to introduce molecules into pollen to modify the genome and confer desired traits to the next generation. To date, we have developed a technology to rapidly edit the male genome by transiently introducing the genome editing tool CRISPR-Cas9 into pollen (Nagahara et al. 2021) (patent). We also developed a highly efficient genome editing enzyme (patent), an efficient introduction method (patent), and a method to collect a gene-introduced pollen using a near-infrared laser system (Kaneshiro et al. 2022). By combining these technologies, we aim to establish plant reproductive engineering technology that controls pollen production and designs plants.

Learn more (magazines, TV, etc.)

Science View Biological Resources, The Scientist "The Ingenious Mechanism of Plant Fertilization"　Jikkyo Publishing February 2026
AXIS Sci-Tech File " Peeking into the depths of flowers - What determines the fate of pollen?! " April 2025 (vol. 232)
The Scientist " The Hidden Dance of Plant Fertilization " April 1, 2025
The Economist " A flower's female sex organs can speed up fertilization " October 1, 2024
NHK Special: Super Evolution Special Edition (1) Message from Plants - The Amazing World that Colors the Earth May 22, 2024
Fluorescence observation of deep inside rice sprouts and stems using plant transparency technology Yusako Co., Ltd. July 2022