Isolation and characterization of a low phytic acid rice mutant reveals a mutation in the rice orthologue of maize MIK.

Using a forward genetics approach, we isolated two independent low phytic acid (lpa) rice mutants, N15-186 and N15-375. Both mutants are caused by single gene, recessive non-lethal mutations, which result in approximately 75% (N15-186) and 43% (N15-375) reductions in seed phytic acid (inositol hexakisphosphate). High-performance liquid chromatography and GC-MS analysis of seed extracts from N15-186 indicated that, in addition to phytic acid, inositol monophosphate was significantly reduced whereas inorganic phosphorus and myo-inositol were greatly increased when compared with wild-type. The changes observed in N15-186 resemble those previously described for the maize lpa3 mutant. Analysis of N15-375 revealed changes similar to those observed in previously characterized rice lpa1 mutants (i.e. significant reduction in phytic acid and corresponding increase in inorganic phosphorus with little or no change in inositol phosphate intermediates or myo-inositol). Further genetic analysis of the N15-186 mutant indicated that the mutation, designated lpa N15-186, was located in a region on chromosome 3 between the microsatellite markers RM15875 and RM15907. The rice orthologue of maize lpa3, which encodes a myo-inositol kinase, is in this interval. Sequence analysis of the N15-186 allele of this orthologue (Os03g52760) revealed a single base pair change (C/G to T/A) in the first exon of the gene, which results in a nonsense mutation. Our results indicate that lpa N15-186 is a mutant allele of the rice myo-inositol kinase (OsMIK) gene. Identification and characterization of lpa mutants, such as N15-186, will facilitate studies on the regulation of phytic acid biosynthesis and accumulation and help address questions concerning the contribution of the inositol lipid-dependent and independent biosynthetic pathways to the production of seed phytic acid.

The rice OsLpa1 gene encodes a novel protein involved in phytic acid metabolism.

The rice low phytic acid 1 (lpa1) mutant was originally identified using a forward genetics approach. This mutant exhibits a 45% reduction in rice seed phytic acid with a molar-equivalent increase in inorganic phosphorus; however, it does not appear to differ significantly in productivity from its wild-type progenitor. A second lpa1 mutant was identified from additional screening for high seed inorganic phosphorus phenotypes. Using a positional cloning strategy, we identified a single candidate gene at the rice Lpa1 locus. Sequence analysis of the candidate gene from the lpa1 mutants revealed two independent mutations (a single base pair substitution and a single base pair deletion) that confirmed the identification of this candidate as the rice low phytic acid 1 gene, OsLpa1. The OsLpa1 gene has three splice variants. The location and nature of the two mutations suggests that these lesions only affect the translation of the predicted protein derived from the longest transcript. The proteins encoded by OsLpa1 do not have homology to any of the inositol phosphate metabolism genes recently characterized in plants, although there is homology to 2-phosphoglycerate kinase, an enzyme found in hyperthermophilic methanogens that catalyzes the formation of 2,3-bisphosphoglycerate from 2-phosphoglycerate. OsLpa1 represents a novel gene involved in phytic acid metabolism.