Silencing the Oryza sativa plasma membrane H+-ATPase isoform OsA2 affects grain yield and shoot growth and decreases nitrogen concentration

Abstract

The plasma membrane (PM) H⁺-ATPase (EC 3.6.1.3.) is a key component involved in nutrient uptake. There are 10 PM H⁺-ATPase isoforms in the rice genome (OsA1-OsA10), and OsA2 is highly responsive to nitrate (NO₃^-). We investigated the role that the OsA2 isoform plays in the total N and growth of rice (Oryza sativa). By the use of artificial microRNA, mutant osa2 rice lines presented ∼70 % downregulated levels of OsA2. Three osa2 lines and control plants (transformed with an empty IRS154 vector and named IRS) were cultivated in the greenhouse to evaluate grain and shoot production. For hydroponic experiments, the same lines were grown in Hoagland solution under two different NO₃^- levels for 30 days - 0.2 mM NO₃^--N (low N) or 2.0 mM NO₃^--N (sufficient N) - or were grown for three days without NO₃^- (starvation) after 27 days under 2.0 mM NO₃^--N. In the greenhouse experiments, compared with the IRS plants, the osa2 lines had lower shoot fresh weights, grain yields and SPAD values. Moreover, compared with the IRS plants, the three osa2 lines grown hydroponically under low NO₃^- levels had lower N concentration and net flux of NO₃^-. PM H⁺-ATPase activity was lower in the osa2 mutants than in the IRS plants. The relatively low N concentration in the osa2 lines was not due to lower expression of OsNRT2.1, OsNRT2.2, or OsNAR2.1. These results indicate that the specific PM H⁺-ATPase isoform OsA2 affects the net flux of NO₃^-, N concentration, and grain yield.

Keywords: Gene silencing; Nitrate transport; Proton pump.