4.3 Key regulatory factors contributed to rice quality formation under elevated temperature
Photosynthesis is the process by which light energy is converted into chemical energy and stored, and it is also the source of accumulation of rice grain assimilation. Chlorophyll content and metabolic enzyme activity are closely related to the strength of photosynthesis. In our case, chlorophyll a-b binding protein 1B-21, chlorophyll a-b binding protein P4, and chlorophyll a-b binding protein 7 were significantly up-regulated, which induced the acceleration of the synthesis and binding of chlorophyll (Ballottari et al., 2012). Meanwhile, the expression levels of photosystem I reaction center subunit VI and oxygen-evolving enhancer protein 3 from the photosystem I were also increased significantly under warming conditions, and that may explain the accelerated grain filling rate during the early grain-filling stage induced by elevated temperature, and the significant increase in the accumulation rate of grain materials compared with the normal temperature treatment. However, the expression of PSB28, which is responsible for water splitting, had a downward trend throughout the period, obtaining a significant low level at 12d after flowering under elevated temperature (0.4 folds). That may inhibit electron transfer and weakens signal transmission, thereby weakening photochemical reactions and resulting in decreased cell chlorophyll and photosynthesis (Wada et al., 2019). Previous research shows that the optical system II (PS II) is the most sensitive element to temperature in the electron transmission chain (Zhang et al., 2011). It would be interesting to further investigate whether PSB28 could be the most critical component affected by high temperature during the photosynthesis process.
To our knowledge, the contents and ratio of starch and storage protein in rice grains are the decisive factors in determining the final rice quality. Rice starch synthesis is regulated by various enzymes, including SSS, SBE, DBE and GBSS. GBSS is the main enzyme responsible for amylose synthesis. Wx protein encoded by the Waxy gene GBSS-I can tightly bind to the starch granules and promote the synthesis of amylose. High temperatures can lead to downregulation of gene expression that regulates GBSS synthesis, resulting in decreased amylose content and increased amylopectin content (Dian et al. 2005; Fujita et al. 2006). Research by Denver (1996) shows that GBSS is not only related to the synthesis of amylose, but also to the extension of amylopectin in starch granules. However, the exact effect of GBSS on the extension of normal starch granules is still unclear. Our results showed that the GBSS enzyme was down-regulated at 6d after flowering under elevated temperature. However, enzymes related to amylopectin synthesis did not change significantly. From 6d to 12d after flowering, the expression level of granule-bound starch synthase was significantly lower than that of the control. Under high temperature, the amylose content of mature rice grains was significantly lower than that of CK treatment, while the amylopectin content was significantly increased (Ahmed et al., 2015). Expression levels of the soluble starch synthase SS4 and SSS2-3, responsible for the synthesis of amylopectin, were also decreased under high temperatures (Yamakawa, 2012). This change may reduce the activities of granular starch synthase and soluble starch synthase, and lead to change in the ratio of amylose and amylopectin, which eventually affected the physical and chemical properties of starches in rice grain (Tang et al., 2019).
Rice storage proteins include albumin, globulin, glutelin and prolamin. Prolamin is directly deposited in the endoplasmic reticulum cavity in the form of intracellular protein particles, and finally buds from the endoplasmic reticulum in the form of spherical protein bodies (PBIs). While glutelin is efficiently converted into mature form by vacuolar processing enzymes, and forms irregular protein bodies II (PBII) together with α-globulin (Krishnan et al., 1992; Kumamaru et al., 2010). The results of this study showed that warming had significant up- or down-regulation effects on the expression of storage protein family-related regulatory factors at different periods. For example, the expression of glutelin type-A and type-B proteins were either significantly up-regulated or down-regulated at 3d and 6d after flowering, and there is no obvious rule for the regulation mode of these regulatory factors under warming conditions. Based on our understanding of glutelin, its synthesis pathway is still unclear, and the presence of many unknown glutelin genes increases the difficulty in understanding the expression pattern under warming condition. Therefore, this study has not been able to essentially find the direct reasons for the changes in the final grain storage protein content.
However, several types of protein species (ribosomal protein species, superfamily II DNA and RNA helicase, and molecular chaperone IbpA) related to biosynthesis and processing of proteins was found to be affected by high temperature. Ribosomes are the primary sites for protein synthesis, and different species of ribosomal proteins play an essential role in translation, ribosome structure, and biogenesis in protein anabolism (Moin et al., 2016). In our study, the ribosomal protein species (25S, 30S, 40S, 50S and 60S) exhibited significant decreases during the middle stage of grain-filling, which may cause the reduction in the protein biosynthesis and maintain the balance between synthesis and degradation of proteins (Moin et al., 2016). The reduction in protein content related to translation, such as RNA recognition motif (RRM) domains, eukaryotic initiation factors (eIFs) and elongation factors (EFs), indicates the adverse effects of high temperature on rice protein synthesis. Furthermore, a series of molecular chaperone heat shock proteins (Hsps) were identified to be significantly up-regulated when exposed to high temperature. Heat shock proteins Heat shock protein is a highly conserved peptide in structure and could be activated and produced in large quantities when plants are subjected to abiotic stress (Timperio et al., 2008). In this study, the two most sensitive heat shock proteins are HSP70 and 26.7 kDa heat shock protein of the sHSPs (s