Mutation of Inositol 1,3,4-trisphosphate 5/6-kinase6 Impairs Plant Growth and Phytic Acid Synthesis in Rice.

Meng Jiang,Yang Liu,Jianzhong Huang,Qingyao Shu,Yuanyuan Tan,Yanhua Liu

doi:10.3390/plants8050114

Abstract

Inositol 1,3,4-trisphosphate 5/6-kinase (ITPK) is encoded by six genes in rice (OsITPK1-6). A previous study had shown that nucleotide substitutions of OsITPK6 could significantly lower the phytic acid content in rice grains. In the present study, the possibility of establishing a genome editing-based method for breeding low-phytic acid cultivars in rice was explored, in conjunction with the functional determination of OsITPK6. Four OsITPK6 mutant lines were generated by targeted mutagenesis of the gene’s first exon using the CRISPR/Cas9 method, one (ositpk6_1) with a 6-bp in-frame deletion, and other three with frameshift mutations (ositpk6_2, _3, and _4). The frameshift mutations severely impaired plant growth and reproduction, while the effect of ositpk6_1 was relatively limited. The mutant lines ositpk6_1 and _2 had significantly lower levels (−10.1% and −32.1%) of phytic acid and higher levels (4.12- and 5.18-fold) of inorganic phosphorus compared with the wild-type (WT) line. The line ositpk6_1 also showed less tolerance to osmotic stresses. Our research demonstrates that mutations of OsITPK6, while effectively reducing phytic acid biosynthesis in rice grain, could significantly impair plant growth and reproduction.

Highlights

Myo-inositol-1,2,3,4,5,6-hexakisphosphate (IP6 ), known as phytic acid (PA), is the main storage form of phosphorous (P) (65–80%) in cereal and legume seeds, accounting for ~1.5% of the dry weigh [1]
ITPK6 is a unique gene in the ITPK gene family, and knowledge of its function has so far been very limited
The identification and characterization in rice of two itpk6 mutant lines were reported in a study, which is the only study on the function of ITPK6 in all organisms [30]

Summary

Introduction

Myo-inositol-1,2,3,4,5,6-hexakisphosphate (IP6 ), known as phytic acid (PA), is the main storage form of phosphorous (P) (65–80%) in cereal and legume seeds, accounting for ~1.5% of the dry weigh [1]. The PA biosynthetic pathway is still not well defined, but a number of genes involved in its biosynthesis or transport have already been cloned in several plants. Mutations of these genes could result in low-phytic-acid (lpa) grains in rice [4,5,6,7,8,9,10,11,12,13,14] and other plants, e.g., wheat [15] and maize [3,16,17]. 12 genes have been identified that catalyze the production of intermediate inositol polyphosphates in seeds [18]

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Results

Conclusion