OsGLP8-7 interacts with OsPRX111 to detoxify excess copper in rice.

Lignin is a phenolic biopolymer generated from phenylpropanoid pathway in the secondary cell wall and is required for defense of plants against various stress. Although the fact of stress-induced lignin deposition has been clearly demonstrated, it remains largely elusive how the formation of lignin is promoted under Cu stress. The present study showed that OsGLP8-7, an extracellular glycoprotein of rice (Oryza sativa L.), plays an important function against Cu stress. The loss function of OsGLP8-7 results in Cu sensitivity whereas overexpression of OsGLP8-7 scavenges Cu-induced superoxide anion (O2•-). OsGLP8-7 interacts with apoplastic peroxidase111 (OsPRX111) and elevates OsPRX111 stability when exposed to excess Cu. In OsGLP8-7 overexpressing (OE) lines, the retention of Cu within cell wall limiting Cu uptake into cytoplasm is attributed to the enhanced lignification required for Cu tolerance. Exogenous application of a lignin inhibitor can impair the Cu tolerance of transgenic Arabidopsis lines overexpressing OsGLP8-7. In addition, co-expression of OsGLP8-7 and OsPRX111 genes in tobacco leaves leads to an improved lignin deposition compared to leaves expressing each gene individually or the empty vector. Taken together, our findings provided the convincing evidences that the interaction between OsGLP8-7 and OsPRX111 facilitates effectively lignin polymerization, thereby contributing to Cu tolerance in rice.

OsGLP participates in the regulation of lignin synthesis and deposition in rice against copper and cadmium toxicity.

Copper (Cu) and cadmium (Cd) are common heavy metal pollutants. When Cd and excessive Cu accumulate in plants, plant growth is reduced. Our previous study showed that Germin-like proteins (GLPs), which exist in tandem on chromosomes, are a class of soluble glycoproteins that respond to Cu stress. In this study, hydroponic cultures were carried out to investigate the effect of GLP on Cd and Cu tolerance and accumulation in rice. The results showed that knockout of a single OsGLP8-2 gene or ten OsGLP genes (OsGLP8-2 to OsGLP8-11) resulted in a similar sensitivity to Cd and Cu toxicity. When subjected to Cu and Cd stress, the glp8-2 and glp8-(2-11) mutants displayed a more sensitive phenotype based on the plant height, root length, and dry biomass of the rice seedlings. Correspondingly, Cu and Cd concentrations in the glp8-2 and glp8-(2-11) mutants were significantly higher than those in the wild-type (WT) and OsGLP8-2-overexpressing line. However, Cu and Cd accumulation in the cell wall was the opposite. Furthermore, we determined lignin accumulation. The overexpressing-OsGLP8-2 line had a higher lignin accumulation in the shoot and root cell walls than those of the WT, glp8-2, and glp8-(2-11). The expression of lignin synthesis genes in the OsGLP8-2-overexpressing line was significantly higher than that in the WT, glp8-2, and glp8-(2-11). The SOD activity of OsGLP8-2, Diaminobe-nzidine (DAB), propidium iodide (PI) staining, and Malondialdehyde (MDA) content determination suggested that OsGLP8-2 is involved in heavy metal-induced antioxidant defense in rice. Our findings clearly suggest that OsGLPs participate in responses to heavy metal stress by lignin deposition and antioxidant defense capacity in rice, and OsGLP8-2 may play a major role in the tandem repeat gene clusters of chromosome 8 under heavy metal stress conditions.

Excess copper inhibits the growth of rice seedlings by decreasing uptake of nitrate.

Copper (Cu) toxicity has a deleterious effect on plant growth, and different nitrogen (N) forms have significantly different impacts on the uptake and accumulation of heavy metals by plants. However, it remains unclear how excess Cu inhibits the growth of rice seedlings under different N forms. Here, we examined the mechanism of Cu toxicity inhibiting the growth of rice supplied with different N forms. Rice seedlings were grown in a nutrient solution with 0.81 mmol L-1 N, as ammonium (NH4+), nitrate (NO3-) and NH4+ + NO3-, or without N (0 N) in the presence of 0.2 µmol L-1 CuSO4 or 10 µmol L-1 CuSO4. The inhibition of shoot growth under excess Cu was more pronounced in plants that were supplied with NO3- than NH4+; such inhibition was not induced by higher Cu concentration in shoots. Applied with NO3- alone increased solution pH value up to 6.2, but supplied with NH4+ alone and NH4+ + NO3- decreased solution pH value to 4.0 and 4.2, respectively. The increment of solution pH reduced Cu concentration in shoots of rice supplied with NO3- alone. Copper toxicity decreased NO3- concentrations in rice seedlings that were supplied with NO3- alone but increased the NH4+ concentrations in plants that were supplied with NH4+ or NH4+ + NO3-. High Cu levels reduced the uptake of NO3- in roots by the analysis of net NO3- flux and NO3- assimilation enzymes activity. Under excess Cu, the transcript levels of OsNPF6.5, OsNPF2.2 and OsNPF2.4 genes were suppressed, while OsNRT2.1, OsNRT2.2 and OsNAR2.1 were raised in roots. In conclusion, Cu toxicity inhibits NO3- uptake and upward translocation by modulating the expression level of NO3- transporter genes. The reduction in the concentrations of NO3- and total N decreased shoot growth of rice seedlings when N was supplied as NO3-. Hence, rice seedlings supplied with NO3- had lower shoot biomass than those with NH4+ under Cu stress.