Submitted to: Functional Plant Biology
Publication Type: Peer reviewed journal
Publication Acceptance Date: 9/1/2006
Publication Date: 4/1/2007
Citation: Nakata, P.A., McConn, M. 2007. Genetic evidence for differences in the pathways of druse and prismatic calcium oxalate crystal formation in Medicago truncatula. Functional Plant Biology. 34:332-338. Interpretive Summary: Calcium oxalate crystals have been shown to be important in protecting plants from various environmental stresses (e.g., insects, metals, and other ions). From a human heath perspective oxalate is an antinutrient and potential toxin. Thus, efforts to manipulate oxalate biosynthesis to improve the nutritional quality of edible plants will also have to take into account the beneficial effects it has toward protecting the plant. It is currently thought that one primary ascorbate-utilizing pathway of oxalate production occurs in plants. The genetic, biochemical, and cellular methodologies used in this studies, however, revealed the possibility that more than one pathway of oxalate production occurs in the model legume, Medicago truncatula. Microscopic examination of calcium oxalate formation in wild type and mutant M. truncatula plants indicated that there are independent pathways of calcium oxalate formation for each crystal type (prismatic and druse) present in M. truncatula leaves. Measurement of ascorbate levels in the wild type and mutant plants, along with ascorbate induction studies revealed that ascorbate may be the substrate in the formation of only one of the crystal types (druse). Therefore the possibility exists that there may be another pathway for the formation of the other crystal type (prismatic). The authors offer a working model depicting independent pathways of calcium oxalate formation. In this model two scenarios exist. The pathways may be "different" in that they use different precursors (e.g., ascorbate and some other precursor) and different enzymes or the pathway may be "parallel" in that they use the same precursor (e.g., ascorbate), but have cell-specific isozymes (encoded by different nuclear genes) responsible for the conversion of ascorbate into oxalate. Overall, the knowledge gained from this study is crucial for designing suitable strategies that improves the nutritional value of the edible plant while maintaining adequate protection of the plant from environmental factors.
Technical Abstract: Current evidence supports a single pathway utilizing ascorbic acid as the precursor in oxalate biosynthesis. In this study, we address the possibility that more than one pathway of oxalate biosynthesis and calcium oxalate formation occurs in Medicago truncatula. Like wildtype, developing leaves of the calcium oxalate defective (cod) 4 mutant contains prismatic crystal along the vascular strand, but this mutant also hyper-accumulates druse crystals within the mesophyll cells. A second mutant, cod5, fails to accumulate prismatic crystals along the vascular strand, but is capable of wildtype druse crystal accumulation in maturing leaves. To assess whether a single pathway of oxalate biosynthesis and calcium oxalate formation occurred in M. truncatula we generated and characterized the cod4/cod5 double mutant. Microscopic examination of the cod4/cod5 revealed that the double mutant exhibited both cod4 and cod5 mutant crystal phenotypes simultaneously suggesting that the two crystal types may form through independent pathways. Measured ascorbic acid levels and ascorbate induction studies were also consistent with the acid as precursor to oxalate in druse crystal formation but not necessarily prismatic crystal formation. Based on these findings, we propose a working model depicting possible independent pathways of oxalate biosynthesis and calcium oxalate formation.