Natural parks are comprised of preserved forested natural areas that are undergoing natural ecological processes. These areas can offer a refuge for local biodiversity and contribute substantially to ecosystem services in both rural areas with relatively low population densities, as well as high-density urban areas. Forested natural parks located in urban areas should experience more stressful environmental conditions than nearby rural areas, yet we know relatively little about how urbanization impacts tree communities within these important natural habitats. To better understand the impact of urbanization on forests, we investigated the species composition, abundance, and diversity of midstory and canopy trees as well as tree seedlings in urban and rural natural parks in and around Cleveland, Ohio. We found that both urban and rural natural parks have similar tree abundance, but midstory and canopy trees as well as tree seedling communities in the urban natural parks included higher abundances of stress-tolerant species compared to rural parks. In addition, this pattern was driven by changes in native tree species, as we observed low abundance of invasive species. More stress-resistant native species in urban areas include Quercus rubra and Prunus serotina, in contrast to rural natural parks which are dominated by Acer spp. and Fagus grandifolia. Lastly, we show that urban and rural natural parks have similar species diversity within plots, but we found higher variation in community composition among urban natural parks compared to among rural parks. Furthermore, Q. rubra and P. serotina were significantly larger in rural natural parks, indicating that both environmental stress and successional stage could drive compositional differences. Thus, we show that urbanization can have unexpected effects on plant community composition and diversity. Our study refutes the idea that these are degraded habitats, highlights the need to conserve them, and suggests that characterization of local variation in self-assembled urban tree communities will provide the most accurate picture of their management needs and potential ecosystem services.