Submitted to: Planta
Publication Type: Peer reviewed journal
Publication Acceptance Date: 2/18/2002
Publication Date: 7/1/2002
Citation: McConn,M.M., Nakata,P.A. 2002. Calcium oxalate crystal morphology mutants from Medicago truncatula. Planta. 215(3):380-386. Interpretive Summary: Plants acquire crystals of calcium oxalate which have many different shapes. We wanted to figure out, from a genetic point of view, how and why different types of plants developed calcium oxalate crystals that reveal these different shapes under the microscope. In order to do that, we examined leaves from a chemically mutagenized plant population and looked for alterations in calcium oxalate crystal formation. We chose a plant called Medicago truncatula, which is like alfalfa in that it is a forage crop, which means that farmers let their animals graze on this plant in the fields. We performed what is called a mutant screen with a mutant Medicago truncatula plant population. We looked at each plant to find altered crystal shapes. We isolated the plants with altered crystal shapes and tried to genetically characterize them. We found seven different classes of calcium oxalate-defective mutants that showed altered crystals whose shapes were different from those of the original plant. Then we noticed that those mutant plants' crystal shapes were like those found in other legumes. We used clover, broadbean and cowpea as examples of different legumes, for comparison purposes. We were able to match those plants' crystal shapes to those of the mutants in the Medicago plant. After pondering these facts based on our genetic analysis, we formulated the theory that these mutants were generated by simple point mutations. Just one base-pair change was sufficient to alter the shape of the crystal, and these crystals now matched those of other plants. This fact suggested to us that over many years, these plants developed different-shaped crystals by these simple point mutations, which contributed to the evolution of the plants.
Technical Abstract: Plants accumulate crystals of calcium oxalate in a variety of shapes and sizes. Each plant forms a crystal or set of crystals with a specific morphology. The mechanism(s) through which a plant defines the morphology of its crystals remains unknown. To gain insight into the mechanisms regulating crystal shapes, we conducted a mutant screen to identify the genetic determinants. A single leaf was harvested from individual Medicago truncatula plants that had been chemically mutagenized. Each leaf was visually inspected, using crossed-polarized light microscopy, for alterations in crystal shape and size. Seven different crystal morphology defective mutants were identified. Six mutants were recessive and one dominant. Genetic analysis of the 6 recessive mutants suggested that each mutant was affected at a different locus. The growth and development of the crystal morphology mutants appeared similar to wild type. This observation coupled with the finding that several of the mutants had drastically altered the amount of calcium they partition into the oxalate crystal, questions current hypotheses regarding crystal function. Comparisons between the mutant crystals and those present in other legumes indicated the likelihood that simple point mutations contributed to the evolution of the variations in prismatic crystal shapes commonly observed in these plants today. The availability of crystal morphology mutants provides the opportunity to investigate aspects of crystal shape and size that have been recalcitrant by other approaches.