Over 80% of the vascular flowering plants are capable of forming mutualistic symbioses with arbuscular mycorrhizal (AM) fungi. These associations develop in the roots, where in exchange for carbon, the fungal symbionts transfer phosphorus and nitrogen from the soil to the plant. In the symbiosis, the AM fungi inhabit the root cortical cells where they form branched hyphae called arbuscules. The arbuscules are responsible for nutrient delivery to the root cells. Previous studies in Medicago truncatula indicated that a symbiosis-specific phosphate transporter, MtPT4, is essential for symbiotic phosphate transport. In this project the investigators will further analyze MtPT4 and in addition, begin to characterize the roles of a second phosphate transporter, MtPT8, and a novel ammonium transporter, MtAMT2. Single, double and triple mutant plants lacking these transporters will be generated and they will be used to determine the extent to which symbiotic phosphate and nitrogen transport regulate the AM symbiosis. Experiments will address the underlying mechanisms and signaling pathways involved. A second aspect of the project focuses on protein targeting in the symbiosis. The MtPT4 protein is located exclusively on a specialized membrane called the peri-arbuscular membrane, which surrounds the arbuscule. Using immunological approaches, and live cell imaging of MtPT4 tagged with fluorescent proteins, the investigators will analyze the mechanisms underlying protein targeting to the peri-arbuscular membrane. The AM symbiosis is formed by almost all vascular flowering plants but the proteins that mediate symbiotic phosphate and ammonium transport, the mechanisms that regulate maintenance of key symbiotic interfaces, and the signaling pathways that integrate regulatory mechanisms with the mineral nutrient status of the plant, are all largely unknown. The data generated in these experiments will advance our understanding of these aspects of the symbiosis. The project includes opportunities for undergraduate students to obtain experience with confocal microscopy.
Over 80% of the vascular flowering plants are capable of forming mutualistic associations with soil fungi, called arbuscular mycorrhizal (AM) fungi. These associations (called AM symbioses) develop in the roots, where the fungus inhabits the root cells and delivers phosphate from the soil to the plant. The transfer of phosphate from the fungus to the plant occurs through specialized structures called arbuscules. Phosphate is an essential mineral nutrient that plants require for growth but it can be difficult for plants to obtain. Consequently, the additional phosphate delivered by the fungal partner, is beneficial for plant health and growth. In this project, we investigated aspects of symbiotic phosphate transport in a model legume plant, Medicago truncatula during AM symbiosis. The goals were to determine the extent to which symbiotic phosphate transport regulates the AM symbiosis, and to assess the underlying mechanisms and signaling pathways involved. Also to determine how phosphate transporters are directed to the appropriate membrane within the cell during symbiosis. The data generated has provided several new insights. First, the activities of both phosphate and nitrogen transporters at the symbiotic membrane influence survival of the fungus in the root and consequently maintenance of the association. Through analysis of M. truncatula mutants, we were able to demonstrate that the signaling pathways involved in signaling the plants nutrient status during AM symbiosis differ from those used during symbiosis with rhizobia. The project also provided a new understanding of the cell biology of the colonized root cells and of the mechanisms that underlie phosphate transporter trafficking in the cell. Studies of tagged phosphate transporter proteins and phosphate transporter mutants showed that the cell alters its default secretion pathways to send phosphate transporters to the symbiotic membrane that surrounds the arbuscule. This was an unexpected finding which changes the direction of current thinking about how the cell populates the symbiotic membrane with the appropriate proteins. It also provides new approaches to manipulate the symbiosis. The project findings have several broader impacts. Phosphorous (P) is essential for plant growth and is one of the mineral nutrients that limits crop production worldwide. P-based fertilizers are costly and fertilizer runoff contributes to the contamination of aquatic ecosystems. In addition, the source material for P-based fertilizers is a non-renewable resource. Consequently, there are strong economic and ecological incentives for increasing the efficiency of P-fertilizer usage and modifying agricultural practices to improve sustainability. Understanding the AM symbiosis to enable its use in agriculture and land management practices is one part of the solution. The project provided training for scientists at all levels of their careers, including several undergraduate minority students.
