Proteomic analysis of dental enamel from 20 Homo naledi individuals shows no male markers

The Rising Star cave system excavations resulted in a high number of well-preserved skeletal specimens from multiple individuals of Homo naledi, showing a high degree of morphological homogeneity, including dental variation possibly consistent with a single-sex sample. Here, we report the paleoproteomic analysis of dental enamel proteins extracted via micro-destructive acid etching from 23 H. naledi specimens belonging to a minimum of 20 individuals. After excluding the possibility of technical bias, no convincing evidence supporting the confident identification of male individuals was detected in any of the investigated samples. We also detect no variability in the recovered proteome, and we observe two amino acid substitutions: a derived one in amelogenin X compared with Homo, and an ancestral one in COL17A1, also present in Paranthropus robustus. Our results further support the homogeneity of H. naledi fossils and show how to sustainably investigate extinct hominins.