The PROTON GRADIENT REGULATION5 (PGR5) protein is required for trans-thylakoid proton gradient formation and acclimation to fluctuating light (FL). PGR5 functionally interacts with two other thylakoid proteins, PGR5-like 1 (PGRL1) and 2 (PGRL2); however, the molecular details of these interactions are largely unknown. In the Arabidopsis (Arabidopsis thaliana) pgr5-1 mutant, the PGR5G130S protein accumulates in only small amounts. In this work, we generated a knockout allele of PGR5 (pgr5-Cas) using CRISPR-Cas9 technology. Like pgr5-1, pgr5-Cas is seedling-lethal under FL, but photosynthesis and particularly cyclic electron flow, as well as chlorophyll content, are less severely affected in both pgr5-Cas and pgrl1ab (which lacks PGRL1 and PGR5) than in pgr5-1. These differences are associated with changes in the levels of 260 proteins, including components of the Calvin–Benson cycle, photosystems II and I, and the NDH complex, in pgr5-1 relative to the wild type (WT), pgr5-Cas, and pgrl1ab. Some of the differences between pgr5-1 and the other mutant lines could be tentatively assigned to second-site mutations in the pgr5-1 line, identified by whole-genome sequencing. However, others, particularly the more pronounced photosynthetic defects and PGRL1 depletion (compared to pgr5-Cas), are clearly due to specific negative effects of the amino-acid substitution in PGR5G130S, as demonstrated by complementation analysis. Moreover, pgr5-1 and pgr5-Cas plants are less tolerant to long-term exposure to high light than pgrl1ab plants. These results imply that, in addition to the previously reported necessity of PGRL1 for optimal PGR5 function, PGR5 is required alongside PGRL1 to avoid harmful effects on plant performance.

中文翻译：

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拟南芥质子梯度调节5变体光合功能的共性和特异

质子梯度调节 5 (PGR5) 蛋白是跨类囊体质子梯度形成和适应波动光 (FL) 所必需的。PGR5 与其他两种类囊体蛋白 PGR5-like 1 (PGRL1) 和 2 (PGRL2) 在功能上相互作用；然而，这些相互作用的分子细节在很大程度上是未知的。在拟南芥 (Arabidopsis thaliana) pgr5-1 突变体中，PGR5G130S 蛋白仅少量积累。在这项工作中，我们使用 CRISPR-Cas9 技术生成了 PGR5 (pgr5-Cas) 的敲除等位基因。与 pgr5-1 一样，pgr5-Cas 在 FL 下对幼苗是致命的，但 pgr5-Cas 和 pgrl1ab（缺乏 PGRL1 和 PGR5）对光合作用和特别是循环电子流以及叶绿素含量的影响不如 pgr5 -1。这些差异与 260 种蛋白质水平的变化有关，包括相对于野生型 (WT)、pgr5-Cas 和 pgrl1ab 在 pgr5-1 中的 Calvin-Benson 循环、光系统 II 和 I 以及 NDH 复合物的成分。pgr5-1 和其他突变系之间的一些差异可以暂时归类为 pgr5-1 系中的第二位点突变，通过全基因组测序鉴定。然而，其他的，特别是更明显的光合缺陷和 PGRL1 消耗（与 pgr5-Cas 相比），显然是由于 PGR5G130S 中氨基酸取代的特定负面影响，如互补分析所示。此外，pgr5-1 和 pgr5-Cas 植物对长期暴露于强光的耐受性低于 pgrl1ab 植物。这些结果表明，除了先前报道的 PGRL1 对最佳 PGR5 功能的必要性外，