Alpine areas are well known biodiversity hotspots, but their future may be threatened by expanding forest and changing human land use. Here, we reconstructed past vegetation, climate, and livestock over the past ~ 12,000 years from Lake Sulsseewli (European Alps), based on sedimentary ancient DNA, pollen, spores, chironomids, and microcharcoal. We assembled a highly-complete local DNA reference library (PhyloAlps, 3,923 plant species), and used this to obtain an exceptionally rich sedaDNA record of 366 plant taxa. The vegetation mainly responded to temperature during the first half of the Holocene, while human activity drove changes from 6 ka onwards. Land-use shifted from episodic grazing (Neolithic, Bronze Age) to agropastoral intensification (Medieval Age). This prompted a coexistence of species typically found at different elevational belts, thereby increasing plant richness to levels that characterise present-day alpine diversity. Our results indicate that traditional agropastoral activities should be maintained to prevent reforestation and preserve alpine plant biodiversity.