Development of a novel aluminum tolerance phenotyping platform used for comparisons of cereal aluminum tolerance and investigations into rice aluminum tolerance mechanisms

Abstract

The genetic and physiological mechanisms of aluminum (Al) tolerance have been well studied in certain cereal crops, and Al tolerance genes have been identified in sorghum (Sorghum bicolor) and wheat (Triticum aestivum). Rice (Oryza sativa) has been reported to be highly Al tolerant; however, a direct comparison of rice and other cereals has not been reported, and the mechanisms of rice Al tolerance are poorly understood. To facilitate Al tolerance phenotyping in rice, a high-throughput imaging system and root quantification computer program was developed, permitting quantification of the entire root system, rather than just the longest root. Additionally, a novel hydroponic solution was developed and optimized for Al tolerance screening in rice and compared with the Yoshida's rice solution commonly used for rice Al tolerance studies. To gain a better understanding of Al tolerance in cereals, comparisons of Al tolerance across cereal species were conducted at four Al concentrations using seven to nine genetically diverse genotypes of wheat, maize (Zea mays), sorghum, and rice. Rice was significantly more tolerant than maize, wheat, and sorghum at all Al concentrations, with the mean Al tolerance level for rice found to be 2- to 6-fold greater than that in maize, wheat, and sorghum. Physiological experiments were conducted on a genetically diverse panel of more than 20 rice genotypes spanning the range of rice Al tolerance and compared with two maize genotypes to determine if rice utilizes the well-described Al tolerance mechanism of root tip Al exclusion mediated by organic acid exudation. These results clearly demonstrate that the extremely high levels of rice Al tolerance are mediated by a novel mechanism, which is independent of root tip Al exclusion.

Figure 1.

Mean root growth (±sd) of seven rice genotypes in Yoshida's (gray diamonds) and modified Magnavaca's (black diamonds) control and Al solutions. The Al concentrations represent previously reported concentrations for rice Al tolerance screening in Yoshida's (1,297 μm) and concentrations for modified Magnavaca's (540 μm) determined in this study. In control solutions (0 μm), root growth is identical. A, Total rootgrowth in response to the concentration of soluble Al in Yoshida's and modified Magnavaca nutrient solutions (r² = 0.92). B, Total root growth in response to concentration of total Al in Yoshida's and modified Magnavaca's nutrient solutions (r² = 0.76).

Figure 2.

Example where growth of the longest root in an Al-grown (right) and a control-grown (left) rice seedling is similar but total root growth is significantly different. Images are of plants representative of the mean growth of the longest root in control (−Al) and treatment (+Al) solutions for genotype NSF4. The mean longest root growth was 1.8 ± 0.14 cm in control solution and 2.0 ± 0.18 cm in treatment solution. However, the mean total root growth was 50.29 ± 7.3 cm in control and 27.10 ± 2.47 cm in treatment. The mean longest root RRG was 1.11, although the mean total root RRG was only 0.54.

Figure 3.

Average Al tolerance (RRG) of rice (n = 8), maize (n = 9), wheat (n = 8), and sorghum (n = 7) at three Al³⁺ activities ± sd.

Figure 4.

Phenotypic distribution of Al tolerance in rice, maize, sorghum, and wheat at three Al³⁺ activities: 160 μm (A), 27 μm (B), and 8.75 μm (C). [See online article for color version of this figure.]

Figure 5.

Correlation of root tip (1 cm) Al accumulation and Al tolerance across 23 genetically diverse rice genotypes. Genotypes were selected to represent the genetic and Al tolerance variation across the Indica (crosses) and Japonica (triangles) varietal groups. Note that there is no correlation between root tip Al accumulation and Al tolerance (r² = 0.002).

Figure 6.

Correlation of root exudates of citrate (A and D), malate (B and E), and phosphate (C and F) with Al tolerance (RRG) in the left column and root tip Al content in the right column for 11 genetically diverse Indica varieties. A significant negative correlation is observed between root citrate exudation and root tip Al content (D). However, there is no correlation between root citrate exudation and Al tolerance. There is a slight correlation between malate exudation and Al tolerance, although there is no relation between malate exudation and root tip Al exclusion (A). For the rest of the parameters, there is either no or very weak correlation.

Figure 7.

Relative gene expression determined using quantitative RT-PCR for the rice homolog of the sorghum Al tolerance gene, SbMATE, in roots of four rice genotypes that represent a wide range of Al tolerance. The Al tolerance (RRG) for each genotype is indicated below the name of each genotype.