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1. Spatial variation in species composition within and among communities may be caused by deterministic, niche-based species sorting in response to underlying environmental heterogeneity as well as by stochastic factors such as dispersal limitation and variable species pools. An important goal in ecology is to reconcile deterministic and stochastic perspectives of community assembly and to assess the contribution of each class of processes to community dynamics and structure. 2. We present an 8-year field experiment of grassland secondary succession that documents plant community differentiation in response to an experimental gradient of nitrogen (N) fertilization, factorially crossed with a manipulation of the available species pool achieved using a multi-species seed sowing treatment. We evaluate the hypothesis, adapted from meta-community theory, that seed availability limits the contribution of niche-based, species sorting to patterns of community variation along an environmental gradient. 3. The magnitude of species sorting and community differentiation observed in response to N fertilization (measured as N effect size on multivariate community composition) became progressively more distinct and more statistically significant over time in experimental plots as succession proceeded. However, this response was significantly more pronounced among plots that had been exposed to experimentally enriched propagule pools. 4. Synthesis. Our findings support the hypothesis that dispersal limitations and species pools can mediate the contribution of niche-based, species-environment sorting to plant community development and limit the extent to which underlying resource gradients become deterministically expressed in patterns of vegetation composition. We suggest that continued habitat destruction and fragmentation in the former prairie landscape where this study was conducted would further reduce native species pools and habitat connectivity, diminishing opportunities for species-environment sorting and compromising the capacity of these grassland systems to respond to environmental change.