The contribution of multidisciplinary approaches to the study of the lifestyles of ancient populations is now well established in the literature. Mobility, nutrition and physical activity are indicators that, when considered together, can document social organisations such as patrilocality, gender practices or social status. Our study is part of this dynamic scientific work and focuses on the Middle Neolithic human remains from the site of Pontcharaud (excavation 1986; 4300 − 3900 BCE; Auvergne, France). It aims to reconstruct patterns of behaviour, life history, and social organisation, particularly in light of the site’s unique funerary features, which remain a subject of debate. New data from bioanthropological investigations (72 individuals), stable isotope measurements (bone collagen δ34S from 55 individuals; 87Sr/86Sr from teeth enamel on 17 individuals) and genetic sex estimation (2 subadult individuals) were collated and integrated with contextual information, including funerary treatments, previously published isotopic data (δ13C, δ15N), and archaeological records. Stable isotope data suggest that some females were exposed to more variable environments compared to males. Strontium and sulphur isotope ratios also indicate differences in resource exploitation (water, plants, and animals from different origin or growth conditions) and suggest a different provenance for at least two females, one of whom was buried in a stone coffin. The seven adult and immature male individuals from the site’s only multiple grave yielded distinct stable isotope results (C, N, S and, to a lesser extent, Sr), suggesting different origins and/or animal protein intake, compared to individuals buried in single or double graves. Our multidisciplinary data reveal important variability in diet, provenance, and organisation within the Pontcharaud funerary group. We propose that the seven males from the multiple gathering may have been involved in a shared activity that exposed them to the same environment—perhaps as individuals engaged in a common economic or social practice.