Hydrologic variability and connectivity influence stream-fish assemblage structure, but other correlated habitat variables often confound attempts to document such relationships. By their nature, dams decrease stream connectivity, but upstream influences on tributary fish assemblages are infrequently documented and poorly understood. We sampled stream fishes seasonally for one year to compare headwater assemblages of the unimpounded East Fork (nine sites) and the impounded West Fork (12 sites) of the San Jacinto River in Texas. Using multiple regression and canonical ordination, we partitioned variation in species' distributions into that explained by instream structural, hydrologic, and physicochemical factors, drainage features, and season. In addition to effects of temporal variation of connectivity (as related to base flow and season) and other environmental factors, spatial patterns of species distribution between tributaries in the East and West Fork indicated upstream effects of impoundment caused by two probable mechanisms. Lentic habitats of impounded lower reaches in the West Fork apparently reduced movement by fluvial specialists among streams and, thus, reduced their opportunity to recolonize dry reaches compared to populations in East Fork streams. Moreover, West Fork assemblages had more macrohabitat-generalist species, which were abundant in the littoral zone of the impoundment and able to recolonize or tolerate environmental conditions in intermittent reaches. We caution that upstream effects of impoundment could be mistakenly attributed to other factors if they are not explicitly considered in species-environment studies.