# Vacuolar compartments preserved among loosely packed amyloplasts account for heat-induced rice chalky formation under low nitrogen conditions

## TL;DR

This study shows that applying nitrogen reduces chalky rice kernels caused by high temperatures by regulating vacuolar compartments and protein synthesis.

## Contribution

The study reveals how nitrogen application mitigates heat-induced rice chalkiness through cellular and organelle-level changes.

## Key findings

- Nitrogen application reduced chalky kernels from 25.0% to 10.7% under high temperatures.
- Chalky cells showed smaller amyloplasts and protein bodies compared to translucent cells.
- Protein storage vacuole enlargement and arrested amyloplast development were linked to chalky kernel formation.

## Abstract

The regulation of vacuolar compartmentation and protein synthesis during the early ripening stage might be responsible for rice appearance at high temperature.

High temperature at the early ripening stage disrupts protein synthesis to arrest starch and storage protein accumulation in the rice endosperms, leading to the occurrence of chalky kernels (CK), such as white-back kernels (WBK) and basal-white kernels (BWK). In contrast, adequate nitrogen (N) application might sustain protein synthesis and reduce chalky kernels. These processes might be associated with the regulation of vacuolar compartmentation and protein synthesis during heat adaptation, yet the exact cellular dynamics behind the reduction of endosperm air space when applying N have not been examined in the fields. In this study, plants at different N levels were treated under the same high temperatures in the fields and morphological analysis were carried out to examine the time course of changes in organelles compartmentation during the N-enhanced mitigation process. Light and transmission electron microscopic observations were conducted at dorsal and basal endosperm cells, at which chalky formation was presumed to occur under low N conditions during kernel development at high temperature. Results show that CK reached 25.0% in no N-applied plants under heat, whereas N treatment contrastingly decreased CK formation down to 10.7%. In the mature kernels, the areas of chalky cells, amyloplasts, and protein bodies (PBs) were smaller in chalky cells, compared with translucent cells. At the middle ripening stage, volumetric enlargement of protein storage vacuole concomitant with the arrested amyloplast development were both observed in the putative growing chalky cells, resulting in the formation of CK at the late ripening stage. In contrast, N application ameliorated the effect on rice appearance by diminishing the vacuolar size and enhancing protein synthesis to ensure cell size and amyloplast and PB development, increasing the transparency. Therefore, it is proposed that regulation of vacuolar compartmentation and protein synthesis at the ripening stage might be responsible for rice appearance under field conditions.

The online version contains supplementary material available at 10.1007/s00425-025-04793-z.