Laboratory of Tropical Crop Science, Graduate School of Agricultural Science, Kobe University, Kobe, 657-8501, Japan.
Laboratory of Crop Science, Graduate School of Agricultural Science, Kobe University, Kobe, 657-8501, Japan.
Photosynth Res. 2018 Sep;137(3):465-474. doi: 10.1007/s11120-018-0525-9. Epub 2018 May 30.
The relationship between ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) and Rubisco activase (Rca) levels was studied using transgenic rice overexpressing maize Rca (OX-mRca) and knockdown transgenic rice expressing antisense Rca (KD-Rca). The ratio of Rubisco to total soluble protein was lower in OX-mRca, whereas it was higher in KD-Rca than in WT, indicating that Rca expression was negatively correlated with Rubisco content. The expressions of other Calvin-Benson-Bassham cycle enzymes such as sedoheptulose-1,7-bisphosphatase and phosphoribulokinase analyzed by immunoblotting did not show such a negative correlation with Rca, suggesting that the effect of Rca on protein expression may be specific for Rubisco. Although Rubisco content was decreased in OX-mRca, the transcript levels of the Rubisco large subunit (OsRbcL) and the Rubisco small subunit mostly increased in OX-mRca as well as in KD-Rca. Additionally, polysome loading of OsRbcL was slightly higher in OX-mRca than it was in WT, suggesting that the OsRbcL translation activity was likely stimulated by overexpression of Rca. S-methionine labeling experiments demonstrated that there was no significant difference in the stability of newly synthesized Rubisco among genotypes. However, S-methionine-labeled Rubisco was marginally decreased in OX-mRca and increased in KD-Rca compared to the WT. These results suggest that Rca negatively affects the Rubisco content, possibly in the synthesis step.
我们利用过量表达玉米 Rca（OX-mRca）和反义抑制表达 Rca（KD-Rca）的转基因水稻研究了核酮糖-1,5-二磷酸羧化酶/加氧酶（Rubisco）和 Rubisco 激活酶（Rca）之间的关系。与 WT 相比，OX-mRca 中 Rubisco 与总可溶性蛋白的比值较低，而 KD-Rca 中则较高，表明 Rca 的表达与 Rubisco 含量呈负相关。通过免疫印迹分析，其他 Calvin-Benson-Bassham 循环酶如 sedoheptulose-1,7-双磷酸酶和磷酸核糖激酶的表达并没有与 Rca 表现出这种负相关，这表明 Rca 对蛋白质表达的影响可能是针对 Rubisco 的。尽管 OX-mRca 中的 Rubisco 含量降低，但 Rubisco 大亚基（OsRbcL）和 Rubisco 小亚基的转录水平在 OX-mRca 和 KD-Rca 中均增加。此外，与 WT 相比，OX-mRca 中 OsRbcL 的多核糖体加载量略高，这表明 Rca 的过表达可能刺激了 OsRbcL 的翻译活性。S-甲硫氨酸标记实验表明，不同基因型之间新合成的 Rubisco 的稳定性没有显著差异。然而，与 WT 相比，OX-mRca 中 S-甲硫氨酸标记的 Rubisco 略有减少，KD-Rca 中则略有增加。这些结果表明，Rca 可能在合成步骤中负向影响 Rubisco 含量。
