This study investigated the relative influences of environmental, spatial, and historical factors, including the island-specific history of land connectivity, on bat assemblages in the Japanese Archipelago. We collected bat distribution data from 1408 studies and assigned them to Japan's First Standard Grid (approximately 6400 km2). Japanese bat assemblages were analyzed at two scales: the entire Japanese Archipelago comprised 16 islands and exclusively the four main islands. At first, we calculated taxonomic and functional total beta diversity (β total) by Jaccard pairwise dissimilarity and then divided this into turnover (β repl) and richness-difference (β rich) components. We conducted hierarchical clustering of taxonomic beta diversity to examine the influence of the two representative sea straits, Tsugaru and Tokara, which are considered biogeographical borders. Variation partitioning was conducted to evaluate the relative effects of the three factors on the beta diversity. Clustering revealed that the Tokara Strait bordered the two major clades; however, the Tsugaru Strait did not act as a biogeographical border for bats. In the variation partitioning, shared fraction between spatial and historical factors significantly explained taxonomic and functional β total and taxonomic β repl at the entire archipelago scale, but not at the four main islands scale extending only Tsugaru Strait but not Tokara Strait. Pure environmental factors significantly explained functional β total at both scales and taxonomic β total only at the four main islands scale. These results suggest that spatial and historical factors are more pronounced in biogeographical borders, primarily structuring assemblage composition at the entire archipelago scale, especially in taxonomic dimension. However, current environmental factors primarily shape the assemblage composition of Japanese bats at the main island scale. The difference in results between the two scales highlights that the primary processes governing assemblages of both dimensions depend on the quality of the dispersal barriers between terrestrial and aquatic barriers for bats.