Transcriptional profiling of wheat and wheat-rye addition lines to identify candidate genes for aluminum tolerance

Abstract

A large-scale expression profiling study was performed to investigate candidate genes associated with the two quantitative trait loci for aluminum (Al) tolerance (Alt1 and Alt2). They have been identified in rye and localized on chromosomes 6R and 3R, respectively. Materials employed were hexaploid wheat (cv. Chinese Spring), and two wheatrye addition lines (3R-AL and 6R-AL). Seedlings were treated with and without Al for 24 h to examine genes upregulated or down-regulated by Al. Measurements of root growth at different Al concentrations showed the Al tolerance was higher in 3R-AL than in 6R-AL and wheat. Initial transcriptomic results revealed that more genes changed expression (>10 fold) in the wheat and in the 6R-AL line (moderately tolerant) than in the 3R-AL line (highly tolerant). A method was developed to determine whether candidate genes are involved in Al tolerance or in responses to Al toxicity. Real-time quantitative PCRs were carried out in a subset of six genes with known function in near isogenic rye lines 389 (Al-sensitive) and 390 (Al tolerant). All six genes were up-regulated by Al in line 389 but not in line 390, indicating that they were involved in Al stress response but not in Al tolerance mechanisms. Subsequent analysis of Arabidopsis lines with knockout mutations in homologues of these six genes showed an Al sensitivity similar to the wild-type, providing more evidence towards their participation in the response to stress rather than to Al tolerance. Once the stress response genes were ruled out, the focus was turned to the identification of tolerance genes by studying transcripts up-regulated and down-regulated in the tolerant 3R line with respect to wheat and 6R line. Finally, a list of candidate genes that could be conferring increased tolerance was obtained.