Habitat fragmentation has been implicated as one of the major cause of diversity decline in dendritic ecological networks such as river ecosystems. Previous studies have shown that river size and local environmental quality regulate fish assemblages in fragmented habitats. Fish assemblages in main stem are influenced by not only habitat size and local environmental conditions but also distance to and characteristics of nearest tributaries, but few studies have considered river network structure to evaluate effects of habitat fragmentation on the assemblages. Thus, I examined effects of fragmentation by dams on river fish assemblages by focusing on ecosystem size, quality, and river network structure in the Kiso, Nagar and Ibi River, central Japan. Fragmented ecosystem size and water quality were important factors for explaining fish assemblages, and these effects did not vary among species. River network structure, such as distance to nearest tributaries, influenced fish assemblages, but the effects were dependent on species. Pelagic fishes were more influenced by the network structure than benthic and sedentary ones. Our findings suggest that connectivity and complexity of river networks should be considered to understand metacommunity structure in fish assemblages.