Background. Determining differences in plant-mycorrhizal fungal associations across spatial gradients in plant ranges over time can only provide data on how the identity of symbiotic partners changes, not the effect of that identity on the outcome of symbiosis. To test how plants perform with different mycorrhizal fungal symbionts, I will manipulate the identity of mycorrhizal fungal symbionts in two common gardens, one at the northern portion of plant ranges in upper Michigan and one near the southern part of plant ranges in Knoxville, Tennessee. The northern common garden is at the leading edge of three of the EM and two of the AM plant species and will simulate range expansion of all ten focal plant species. The southern common garden is at the trailing edge of four of the EM plant species and three of the AM plant species and will simulate the environmental stress of plants remaining in place at the trailing edge of their current range. As plants move, they will either encounter new mycorrhizal fungal symbionts or co-disperse with their previous mycorrhizal fungal symbionts. Therefore, each common garden will contain a fully factorial combination of plant and mycorrhizal fungal symbiont soils in local soil. To account for the fact that plant species have different range distributions, I will include distance from geographic and/or environmental range center as a covariate in all statistical models. Approach. Common garden design. The northern common garden will be constructed at the University of Michigan Biological Station (UMBS) in Pellston, MI (see letter of support). The UMBS covers 13,000 acres of forests and has long been used to study many of the focal plant species in the proposed research (Sakai 1990, McCarthy-Neumann and Ibanez 2012, Lee and Ibanez 2021). The southern common garden will take place on the University of Tennessee, Knoxville (UTK) campus. At each common garden, I will plant seeds of each focal plant species from the leading, center, or trailing edges of their ranges into trenched and lined plots in the ground. Each plant seedling will receive belowground fungal symbionts from their own leading, center, or trailing edge, or a control with no mycorrhizal fungi and bacteria only. Belowground fungal symbionts will always be sourced from the same location for each plant manipulation (i.e., leading edge mycorrhizal fungal symbionts will be placed on seeds collected from the same location when paired). All treatments will be replicated four times for 360 plants per common garden, 720 total (10 plant species x 3 plant sources x 3 fungal symbiont sources x 4 replicates = 360 plants per common garden). Common garden setup. Plant seeds will be collected at maturity from leading, center, and trailing populations in 2024. Seeds will be surface sterilized and stored at 4°C. Fungal symbionts will be collected from rhizosphere soil and plant roots of focal tree species following Kivlin and Hawkes (2018). Spores will be isolated from soil using the sucrose floatation method (Ianson and Allen 1986). Surface-sterilized roots will be ground into a slurry and applied to rhizosphere soils. Seedlings and mycorrhizal fungal inocula will be planted in the ground in trenched plots that will allow recovery of belowground biomass at the end of the experiment. Seedlings will be grown in local soil in each respective common garden that has not been sterilized. These conditions mimic plants and mycorrhizal fungi shifting ranges into new locations that already contain an intact soil microbiome. Common garden measurements. Plant survival, growth, and reproduction will be measured monthly on each plant in each common garden during the growing season for four years (2025-2028). At the end of four years, I will harvest each tree and measure plant biomass, root:shoot ratios, and plant C/N. I will also measure fungal colonization rates and fungal composition in roots as in AIMS I and II. Finally, I will measure soil C/N and potential C fractions (Mineral-Associated Organic Matter [MAOM] and Particulate Organic Matter [POM]) under each plant to understand the ecosystem consequences of shifting plant and mycorrhizal fungal ranges. All methods for these measurements have been honed in the Kivlin Lab over the last five years (see Kivlin et al. 2018, Kivlin et al. 2021). Expected outcomes. I expect plant growth, survival, and reproduction to be highest when plants are grown with mycorrhizal fungal symbionts from the center of their ranges since fungal symbiont specificity is often highest in range centers (e.g., Cobian et al. 2019). Alternatively, if less beneficial mycorrhizal fungal symbionts accumulate at the center of a plant’s range, leading edge soils should decrease negative interactions between plants and fungal symbionts, thereby increasing plant growth, survival, and reproduction (e.g., McCarthy-Neumann and Ibanez 2012). Trailing edges may promote refugia of plant and mycorrhizal fungal symbiont diversity (Love et al. in review). If this is the case, trailing edge mycorrhizal fungal symbionts may increase plant growth, survival, and reproduction more than center or leading edge fungal communities. Mitigating risk. Finding seeds for all ten tree species may be challenging in natural systems, as seed production varies among plant taxa throughout the growing season and from year to year. If seeds cannot be collected from sites sampled in AIM I, I will purchase seeds from commercial vendors from multiple locations (north and south) to ensure high genetic variation in seed source. Additionally, it is well known that seedling mycorrhizal fungal associations can be different in composition (Husband et al. 2002), colonization magnitude, and effects on plant growth (Hawkes et al. 2013) compared to adult plants. While this experiment will be conducted for four years, even juvenile plants may not reflect trends in adults. However, since plants will disperse as seeds as plant ranges shift poleward, the seedling and juvenile plant interactions with mycorrhizal fungi are the crucial establishment phase that will demonstrate how successfully plant and mycorrhizal fungal interactions will persist with climate change. If composition of mycorrhizal fungi is significantly different in seedlings compared to adults in AIMS I and II, I will collect mycorrhizal fungi from seedlings in my twenty core sites to ensure that treatment bias or experimental error did not influence the outcome of the common garden experiments. Finally, establishment success of mycorrhizal fungal inocula is not guaranteed. However, I have experience with these inoculation methods on tree seedlings (see Kivlin and Hawkes 2018) and previous studies have shown persistence of mycorrhizal fungal inocula in natural systems for decades following transplantation (Selosse et al. 1998).