Global change presents risks and opportunities for social-ecological systems worldwide. Key challenges for sustainability science are to identify plausible future changes in social-ecological systems and find ways to reach socially and environmentally desirable conditions. In this context, regional-scale studies are important, but to date, many such studies have focused on a narrow set of issues or applied a narrow set of tools. Here, we present a holistic approach to work through the complexity posed by cross-scale interactions, spatial heterogeneity, and multiple uncertainties facing regional social-ecological systems. Our approach is spatially explicit and involves assessments of social conditions and natural capital bundles, social-ecological system dynamics, and current development trends. The resulting understanding is used in combination with scenario planning to map how current development trends might be amplified or dampened in the future. We illustrate this approach via a detailed case study in southern Transylvania, Romania, one of Europe’s most significant biocultural refugia. Our goal was to understand current social-ecological dynamics and assess risks and opportunities for sustainable development. Our findings show that historical events have strongly shaped current conditions and current development trends in southern Transylvania. Moreover, although external drivers (including EU policies) set the general direction of regional development trajectories, local factors, including education, leadership, and the presence of bridging organizations, can enhance or counteract their effects. Our holistic approach was useful for generating an in-depth understanding of a regional social-ecological system and could be transferred to other parts of the world.