Rebun Island with Hamanaka and Funadomari among the 43 documented archaeological sites and the environmental archive stored in the Lake Kushu sediment proves to be one of the key areas to study the interplay between ecology, climate and human activities. This paper focuses on the potential of palaeobotanical records from Rebun Island for improving the chronological control and understanding of late Quaternary climate changes and habitation environments of northern hunter-gatherers in the Hokkaido Region of Japan. A set of 57 radiocarbon dates of the RK12 core (Lake Kushu) demonstrates that it represents a continuous environmental archive covering the last c. 17,000 years. The RK12 pollen record reflects distinct vegetation changes associated with the onset of the lateglacial warming about 15,000 cal. yr BP and the cold climate reversal after c. 13,000 cal. yr BP. The onset of the current Holocene interglacial after c. 11,700 cal. yr BP is marked by a major spread of trees. The middle Holocene (c. 8000–4000 cal. yr BP) is characterized by a major spread of deciduous oak in the vegetation cover reflecting a temperature increase. A decline of oak and spread of fir and pine is recorded at c. 2000 cal. yr BP. After c. 1100 cal. yr BP, arboreal pollen percentages decrease, possibly linked to intensified usage of wood during the Okhotsk and Ainu culture periods. The results of diatom analysis suggest marshy or deltaic environments at the RK12 coring site prior to c. 10,500 cal. yr BP and a brackish lagoon between c. 10,500 and 7000 cal. yr BP. A freshwater lake developed after 6500 cal. yr BP, likely reflecting sea level stabilization and formation of the sand bar separating the Kushu depression from the sea. Plant macrofossil analysis shows use of various wild plants and also domesticated barley during the Okhotsk and Ainu periods.