Peatlands have stored ca. one-third of the global soil carbon stock because of low rates of plant residue decomposition due to oxygen limitation resulting from waterlogging which inhibits microbial activities. Thus, thanks to the continuous accumulation organic matter (OM), peatlands are suitable archives for the reconstruction of past environmental changes. The combination of natural climatic variations and human impacts (particularly during the late Holocene) represents a challenge for reconstructing past environmental changes from peat archives, and requires the combination of several proxies. Within the PEATWARM project (ANR-07-VUL-010), we investigated a 4 m peat profile collected in a bog located in Frasne mire (French Jura Mountains) and covering the last 7,400 years BP. We analysed peat OM composition (lignin-derived phenolic monomers, monosaccharides, lipids), testate amoebae, branched glycerol dialkyl glycerol tetraethers (GDGTs) and pollen. Major changes in OM sources and environmental conditions were revealed at 2.5 m depth (ca. 5,000 yrs BP). Below 2.5 m depth, higher contents of lignin-derived phenolic monomers were found suggesting a predominance of vascular plants (e.g. sedges). In this deeper section, the peat composition is also characterised by (i) higher percentages of amorphous OM, (ii) lower percentages of mucilage and fungal hyphae, and (iii) lower cellulosic sugar contents and C/N ratio. In addition, at this time (around 5,000 yrs BP), pollen analysis revealed a change in vegetation with especially the disappearance of Pinus. Taken together, these results indicate that a rapid shift of the ecosystem functioning occurred ca. 5,000 years ago from a minerotrophic fen to an ombrotrophic Sphagnum-dominated peatland.