OsXTH19 Overexpression Improves Aluminum Tolerance via Xyloglucan Reduction in Rice Root Cell Wall

Abstract

Aluminum (Al) dissolves from soil at low pH and is absorbed by plants, inhibiting their growth. Since most of the Al absorbed by plants is present in the cell wall, it is thought that the binding of Al to cell wall polysaccharides alters the properties of the cell wall and inhibits cell elongation. However, it remains unclear in which component of the cell wall Al accumulates. In this study, we determined the distribution of Al in rice root cell wall fractions under Al stress conditions. The results show that Al accumulates predominantly in the hemicellulose fraction, with star1 mutants accumulating significantly more Al than WT plants. An analysis of cell wall sugars revealed an increase in xyloglucan content under Al stress, which influenced the inhibition of root elongation. OsXTH19, a member of the xyloglucan endotransglucosylase/hydrolase (XTH) family, exhibits only xyloglucan endohydrolase (XEH) activity and lacks endotransglucosylase (XET) activity. OsXTH19 overexpressor rice (OsXTH19-OX) enhances the degradation of xyloglucan. Furthermore, OsXTH19-OX rice with reduced xyloglucan levels exhibited reduced Al accumulation and enhanced root growth under Al stress.

Keywords: Aluminum; Oryza sativa; OsXTH19; xyloglucan.

Figure 1

Al content in different cell wall fractions of WT and aluminum-sensitive mutant star1. Plants were treated without or with Al (0: −Al or 100 µM AlCl₃: +Al), and cell wall fractions were extracted from roots and fractionated into different residues (endo-polygalacturonase (EPG)-soluble fraction as pectic fraction, trifluoroacetic acid (TFA)-soluble fraction as hemicellulosic fraction, and TFA-insoluble fraction as cellulosic fraction). After extraction of Al with HNO₃, Al content was determined by ICP-AES. Data represent means of independent biological replicates ± standard deviation (n = 5). Different letters in each panel indicate significant differences at p < 0.05 (Tukey’s test). ND, not detected.

Figure 2

Ca content in different cell wall fractions of WT and aluminum-sensitive mutant star1. Plants were treated without or with Al (0: −Al or 100 µM AlCl₃: +Al), and cell wall fractions were extracted from roots and fractionated into different residues (EPG-soluble fraction as pectic fraction and TFA-soluble fraction as hemicellulosic fraction). After extraction of Al with HNO₃, Al content was determined by ICP-AES. Data represent means of independent biological replicates ± standard deviation (n = 5). Different letters in each panel indicate significant differences at p < 0.05 (Tukey’s test).

Figure 3

Monosaccharide composition of TFA-soluble fractions of WT and aluminum-sensitive mutant star1. Monosaccharide composition of TFA-soluble fractions in roots from WT (cv Koshihikari) and star1. Plants were treated without or with Al (0: −Al or 100 µM AlCl₃: +Al). SD is given in parentheses. Different letters in each panel indicate significant differences at p < 0.05 (Tukey’s test). [Data represent means of independent biological replicates ± standard deviation. n = 6 (WT), and n = 6 (star1)].

Figure 4

Monosaccharide composition of TFA-soluble fractions of WT and OsXTH19-OX. Monosaccharide composition of TFA-soluble fractions in roots from WT (cv Nipponbare) and OsXTH19-OX. SD is given in parentheses. Plants were treated without or with Al (0: −Al or 100 µM AlCl₃: +Al). Different letters in each panel indicate significant differences at p < 0.05 (Tukey’s test). [Data represent means of independent biological replicates ± standard deviation. n = 6 (WT), and n = 7 (OsXTH19-OX)].

Figure 5

Pectin staining in the roots of the WT and OsXTH19-OX seedlings treated without or with Al (0 or 100 µM AlCl₃). Roots were stained with 0.01% ruthenium red for 5 min after saponification (0.1 N NaOH, 1 min). The experiments were performed at least five times with similar results. Bars = 0.1 mm.

Figure 6

Effects of Al on root growth of WT and OsXTH19-OX. Plants were treated with Al (0, 50, 100 µM AlCl₃). Seedling root length was measured before and after Al treatment, and amount of root elongation was determined. Root elongation differed significantly between WT and OsXTH19-OX rice under 100 µM Al treatment (p < 0.05, Student’s t-test). Data are means ± SDs, n = 12. Different letters in each panel indicate significant differences at p < 0.05 (Tukey’s test).

Figure 7

Al content in different cell wall fractions of WT and OsXHT19-OX. Plants were treated without or with Al (0: −Al or 100 µM AlCl₃: +Al), and cell wall fractions were extracted from roots and fractionated into different residues (endo-polygalacturonase (EPG)-soluble fraction as pectic fraction, trifluoroacetic acid (TFA)-soluble fraction as hemicellulosic fraction, and TFA-insoluble fraction as cellulosic fraction). After extraction of Al with HNO₃, Al content was determined by inductively coupled plasma atomic emission spectrometry. Data represent means of independent biological replicates ± standard deviation (n = 5). Different letters in each panel indicate significant differences at p < 0.05 (Tukey’s test).