The role of environmental change in the late Pleistocene megafaunal extinctions remains a key question, owing in part to uncertainty about landscape changes at continental scales. We investigated the influence of environmental changes on megaherbivores using bone collagen nitrogen isotopes (n = 684, 63 new) as a proxy for moisture levels in the rangelands that sustained late Pleistocene grazers. An increase in landscape moisture in Europe, Siberia and the Americas during the Last Glacial-Interglacial Transition (LGIT; ~25-10 kyr bp) directly affected megaherbivore ecology on four continents, and was associated with a key period of population decline and extinction. In all regions, the period of greatest moisture coincided with regional deglaciation and preceded the widespread formation of wetland environments. Moisture-driven environmental changes appear to have played an important part in the late Quaternary megafaunal extinctions through alteration of environments such as rangelands, which supported a large biomass of specialist grazers. On a continental scale, LGIT moisture changes manifested differently according to regional climate and geography, and the stable presence of grasslands surrounding the central forested belt of Africa during this period helps to explain why proportionally fewer African megafauna became extinct during the late Pleistocene.