Expression and Nucleotide Sequence of an INS (3) P₁ Synthase Gene Associated with Low-Phytate Kernels in Maize (Zea mays L.)

Abstract

Most of the phosphorus (P) in maize (Zea mays L.) kernels is in the form of phytic acid. A low phytic acid (lpa) maize mutant, lpa1-1, displays levels reduced by 66%. A goal of genetic breeding is development of low phytic acid feedstocks to improve P nutrition of nonruminant animals and reduce the adverse environmental impacts of excess P in manure. The genetic basis of the lpa1-1 mutation is not known, but previous genetic mapping has shown both the mutant phenotype and the Ins (3) P(1) synthase (MIPS) gene, which encodes the first enzyme, myo-inositol phosphate synthase, in the phytic acid biosynthetic pathway, map to the same chromosomal region in maize. Research was conducted to determine the expression of the MIPS gene in lpa1-1 and wild-type kernels with similar genetic backgrounds and to ascertain if variation in the MIPS coding sequence could be inferred to be the basis of the mutation. MIPS enzyme activity determined by TLC was reduced 2-3-fold in mutant kernels. RT-PCR-based experiments using primers specific for the 1S-MIPS sequence indicated gene expression was reduced 50-60% in the mutant. Sequence analysis of the MIPS genomic sequence revealed 10 exons and 9 introns that are identical in both mutant and wild-type developing kernels. These findings support an association between reduced MIPS gene activity and low phytic acid content, but additional research should examine the promoter, the 5'UTR, or transcriptional controlling elements of the MIPS gene to ascertain the possible presence of a genetic lesion in those regions.

A paramutation phenomenon is involved in the genetics of maize low phytic acid1-241 (lpa1-241) trait

“…As independently found for both lpa1-1 and lpa1-241 alleles, mips1S gene expression was reduced in developing seeds (Pilu et al, 2003;Shukla et al, 2004), but in both alleles no molecular lesions in the gene coding region were found (Shukla et al, 2004;Pilu et al, 2005).…”

“…These results are consistent with the hypothesis that lpa1-241 mutant phenotype is due to epigenetic silencing of ZmMRP4 and, either directly or indirectly, mips1s gene. We may hypothesize that the reduced mips1s mRNA level independently found in developing seeds of lpa1-1 (Shukla et al, 2004) and in seeds and shoots of lpa1-241 mutants (Pilu et al, 2003), could be due to a metabolic feedback caused by an excess of myo-Ins phosphorylated intermediates or end products, such as phytic acid itself. In fact, MRP proteins in plants are vacuolar transporters involved in detoxification of both xenobiotics or endogenous substances (Klein et al, 2006), thus the loss of ZmMRP4 activity may cause a cytosolic accumulation of its substrate, which may in turn feedback inhibit mips1s transcription.…”

“…Although biochemical, mapping and gene expression data suggest the mips1s gene as a candidate for lpa1 mutation in maize (Raboy et al, 2000;Pilu et al, 2003;Shukla et al, 2004;Pilu et al, 2005), recent transposon mutagenesis experiments found that a novel gene designated ZmMRP4 (accession number EF86878), coding a multidrug resistance-associated-protein (MRP) mapping near the mips1s sequence, is the actual responsible gene for lpa1 mutation (Shi et al, 2007). MRP proteins represent a subfamily of ATP-binding cassette transmembrane transporters widespread in all eukaryotes, which in plants are involved in several functions, such as xenobiotic detoxification, organic ions transport, oxidative stress tolerance (Swarbreck et al, 2003) and even transpiration control (Klein et al, 2006).…”

An Arabidopsis Purple Acid Phosphatase with Phytase Activity Increases Foliar Ascorbate

“…Phytate is found in relatively large quantities in Arabidopsis seeds (24.5 6 3.5 mmol g 21 in dry seeds) and much less in other plant tissues and organs, including leaves, pollen, roots, and tubers (8-45 mmol g FW 21 ; Bentsink et al, 2003;Raboy, 2003). Phytate is synthesized from Glc-6-P through an initial conversion to Ins(3)P 1 by Ins(3)P 1 synthase (Shukla et al, 2004) or from myoinositol catalyzed by a myoinositol kinase (Shi et al, 2005). Phytate and inositol derivatives are important to plant signaling functions.…”

A maize defective-kernel mutant (longcell) characterized by tubular cells, severe morphological alterations and induction of cell death

“…LCD1 encodes a myo-inositol 1-phosphate synthase (MIPS), similar to ZmMIPS1 ( Shukla et al 2004). MIPS protein has been described in many organisms and is the key enzyme in the inositol biosynthetic pathway.…”