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CRISPR/Cas knockout of the NADPH oxidase gene OsRbohB reduces ROS overaccumulation and enhances heat stress tolerance in rice
Thursday, 2024/11/21 | 08:24:29

Xiaolong LiuPing JiJingpeng LiaoXimiao DuanZhiyang LuoXin YuChang-Jie JiangChen XuHongtao YangBo PengKai Jiang

Plant Biotechnology Journal; 01 November 2024; https://doi.org/10.1111/pbi.14500

Summary

Heat stress (HS) has become a major factor limiting crop yields worldwide. HS inhibits plant growth by ROS accumulation, and NADPH oxidases (Rbohs) are major ROS producers in plants. Here, we show that CRISPR/Cas knockout of the OsRbohB (OsRbohB-KO) significantly increased rice tolerance to HS imposed at various different growth stages. We produced OsRbohB-KO and OsRbohB-overexpression (OsRbohB-OE) lines in a japonica cultivar, Nipponbare. Compared with nontransgenic wild-type (WT) plants, the OsRbohB-KO lines showed a significant increase in chlorophyll contents (5.2%–58.0%), plant growth (48.2%–65.6%) and grain yield (8.9%–20.5%), while reducing HS-induced ROS accumulation in seeds (21.3%–33.0%), seedlings (13.0%–30.4%), anthers (13.1%–20.3%) and grains (9.7%–22.1%), under HS conditions. Analysis of yield components revealed that the increased yield of OsRbohB-KO plants was due to increased starch synthetase activity, spikelets per panicle (2.0%–9.3%), filled spikelets (4.8%–15.5%), percentage of filled spikelets (2.4%–6.8%) and 1000-grain weight (2.9%–7.4%) under HS conditions during the reproductive stage. Grain milling and appearance quality, and starch content were also significantly increased in OsRbohB-KO plants under HS conditions during the mature stage. Furthermore, OsRbohB-KO significantly upregulated the expression levels of heat shock-related genes, OsHSP23.7OsHSP17.7OsHSF7 and OsHsfA2a, in rice seedlings and grains under long-term HS conditions. Conversely, OsRbohB-OE resulted in phenotypes that were opposite to OsRbohB-KO in most cases. Our results suggest that suppression of OsRbohB provides an effective approach for alleviating heat damage and improving grain yield and quality of rice under long-term HS conditions.

 

See https://onlinelibrary.wiley.com/doi/10.1111/pbi.14500

 

Figure 7: OsRbohB-suppression enhances heat stress tolerance at multiple rice growth stages. Heat stress induces OsRbohB expression, leading to harmful ROS accumulation (blue arrows) that damages cells, inhibits plant growth and leads to yield loss (black arrow). Blocking OsRbohB through genome editing, targeted mutations or chemical inhibitors (red lines) reduces ROS and mitigates oxidative damage (green arrows). This promotes heat stress tolerance by maintaining cellular integrity, triggering heat shock proteins and boosting starch production (green arrows). Benefits vary across seed, vegetative (seedling), reproductive (anther) and grain filling stages. Arrows and lines with a bar at the end indicate positive and negative regulation, respectively.

 

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