Retrotransposon-Mediated Aluminum Tolerance through Enhanced Expression of the Citrate Transporter OsFRDL4

High aluminum (Al) tolerance of rice (Oryza sativa) is controlled by multiple tolerance genes, but the regulatory mechanisms underlying the differential expression of these genes are poorly understood. Here, we investigated the factors regulating the expression of OsFRDL4, a gene encoding a citrate efflux transporter involved in Al-induced citrate secretion from the roots. Analysis with chromosome segment substitution lines derived from cv Nipponbare (high OsFRDL4 expression) and cv Kasalath (low OsFRDL4 expression) revealed that the differential expression of OsFRDL4 is responsible for the quantitative trait locus for Al tolerance detected previously on chromosome 1. Comparison of the OsFRDL4 gene structure in cv Nipponbare and cv Kasalath showed that there was no difference in the position of the transcriptional start site, but a 1.2-kb insertion showing high similarity to the solo long terminal repeat of the retrotransposon was found in the promoter region of OsFRDL4 in cv Nipponbare. This insertion showed higher promoter activity and contained nine cis-acting elements for ALUMINUM RESISTANCE TRANSCRIPTION FACTOR1 (ART1). However, this insertion did not alter the spatial expression or cellular localization of OsFRDL4. Furthermore, this insertion was found in most japonica varieties but was largely absent from indica varieties or wild rice species. These results indicate that the 1.2-kb insertion in the OsFRDL4 promoter region in japonica subspecies is responsible for their higher expression level of OsFRDL4 due to the increased number of cis-acting elements of ART1. Our results also suggest that this insertion event happened at the initial stage of domestication of japonica subspecies.