Identification and comparison of orthologous cell types from primate embryoid bodies shows limits of marker gene...

The identification of cell types remains a major challenge. Even after a decade of single-cell RNA sequencing (scRNA-seq), reasonable cell type annotations almost always include manual non-automated steps. The identification of orthologous cell types across species complicates matters even more, but at the same time strengthens the confidence in the assignment. Here, we generate and analyze a dataset consisting of embryoid bodies (EBs) derived from induced pluripotent stem cells (iPSCs) of four primate species: humans, orangutans, cynomolgus, and rhesus macaques. This kind of data includes a continuum of developmental cell types, multiple batch effects (i.e. species and individuals) and uneven cell type compositions and hence poses many challenges. We developed a semi-automated computational pipeline combining classification and marker-based cluster annotation to identify orthologous cell types across primates. This approach enabled the investigation of cross-species conservation of gene expression. Consistent with previous studies, our data confirm that broadly expressed genes are more conserved than cell type-specific genes, raising the question of how conserved, inherently cell type-specific, marker genes are. Our analyses reveal that human marker genes are less effective in macaques and vice versa, highlighting the limited transferability of markers across species. Overall, our study advances the identification of orthologous cell types across species, provides a well-curated cell type reference for future in vitro studies and informs the transferability of marker genes across species.