Identification of differentially expressed genes associated with 5-methyltryptophan resistance in rice mutants

Abstract

5-Methyltryptophan (5MT), a tryptophan analog, resistant M 4 rice mutants with high free amino acid contents were obtained through in vitro mutagenesis. To evaluate the 5MT resistance mechanism, a cDNA library was constructed by using the leaves and roots of the 5MT resistant mutant plants. Expressed sequenced tags (ESTs) of 1 019 randomly selected clones were analyzed and then assembled 588 unigens. A total of 389 unigenes had significant homologies with known protein sequences at the NCBI database and the remaining 199 unigenes were designated unidentified genes. These unigens were grouped into 13 categories according to their putative functions. Of the 233 randomly selected clones, 25 were identified as differentially expressed genes between 5MT resistant and 5MT sensitive wild type plants. For further study of the differential expression of the genes, expression patterns of 12 genes related to various biological functions were evaluated in response to 5MT treatment in both the resistant plants and sensitive plants. All of the tested 12 genes exhibited higher expression levels in mutant plants than wild type plants under the 5MT inhibition. These expression patterns of the 12 genes suggested that the genes related to 5MT resistance in the rice mutants have a variety of functions, and yield remarkably diverse expression patterns upon 5MT treatment. Many genes that were identified tend to be related to defense and stress responses, suggesting "cross-talking" between biotic/abiotic stresses including the 5MT treatment. Therefore, 5MT resistant mutants might be of value for identifying genes related to plant defenses and stresses.

Physico-chemical characterization and transcriptome analysis of 5-methyltryptophan resistant lines in rice

“…Mutants and transgenic rice with elevated Trp content have also been characterized extensively [4, 12–16]. 5-Methyltryptophan (5MT) can fit into an allosteric site of AS, the key feedback player in the Trp biosynthetic branch of the shikimate pathway, in the same way as Trp [17]. Even a very sensitive enzyme may fail to completely differentiate between Trp and 5MT.…”

“…However, 5MT cannot replace Trp in cellular protein synthesis, thus arresting cellular growth. Trp over-synthesizing mutants are known to be resistant to growth inhibition [17].…”