Streamflow characteristics are driven by specific flow-generation mechanisms, which are in turn determined by the biophysical properties of catchments. They provide important environmental services for society and ecosystems, regulating water supply and quality, flood mitigation, and the biological diversity of aquatic ecosystems. This study investigates how the drivers of streamflow characteristics vary at the level of regional management (regional [104 km2] and continental scales [107 km2]) in eastern Australia. Three hydrological signatures were used to represent streamflow characteristics: runoff coefficient, baseflow index, and zero flow ratio. Long-term streamflow data and 24 spatially distributed biophysical properties from 354 catchments in eastern Australia were analysed with random forest and generalized additive beta regression models to determine the dominant drivers of streamflow characteristics. We found that the main drivers of streamflow characteristics cannot be generalized from region to region and that specific biophysical variables govern their spatial variability. However, some important drivers such as the dryness index and the fraction of photosynthetically active radiation from vegetation explain the variability of streamflow characteristics at both regional and continental scales with differing importance. Our findings also suggest that soil properties have a significant effect on streamflow characteristics at regional scales. However, the relative importance of these soil properties varies among regions depending on the streamflow characteristics. This paper demonstrates that the drivers of streamflow characteristics are scale and region dependent, and biogeographically different regions have specific mechanisms governing streamflow. It opens an avenue to better connect the management perspectives of ecology and hydrology.