We evaluated the impacts of domestic and agricultural rainwater harvesting (RWH) systems in three watersheds within the Albemarle-Pamlico river basin (southeastern U.S.) using life cycle assessment (LCA) and life cycle cost assessment. Life cycle impact assessment (LCIA) categories included energy demand, fossil fuel, metals, ozone depletion, global warming, acidification, smog, blue and green water use, ecotoxicity, eutrophication, and human health effects. Building upon previous LCAs of near-optimal domestic and agricultural RWH systems in the region, we scaled functional unit LCIA scores for adoption rates of 25%, 50%, 75%, and 100% and compared these to conventional municipal water and well water systems. In addition to investigating watershed-scale impacts of RWH adoption, which few studies have addressed, potential life cycle cost savings due to reduced cumulative energy demand were scaled in each watershed for a more comprehensive analysis. The importance of managing the holistic water balance, including blue water (surface/ground water), green water (rainwater) use, and annual precipitation and their relationship to RWH are also addressed. RWH contributes to water resource sustainability by offsetting surface and ground water consumption and by reducing environmental and human health impacts compared to conventional sources. A watershed-wide RWH adoption rate of 25% has a number of ecological and human health benefits including blue water use reduction ranging from 2–39 Mm3, cumulative energy savings of 12–210 TJ, and reduced global warming potential of 600–10,100 Mg CO2 eq. Potential maximum lifetime energy cost savings were estimated at $5M and $24M corresponding to domestic RWH in Greens Mill and agricultural RWH in Back Creek watersheds.