Abstract
Background color matching is essential for camouflage and thermoregulation in ectothermic vertebrates, yet several key cellular-level questions remain unresolved. For instance, it is unclear whether the number of chromatophores or the activity of individual chromatophores plays a more critical role in this process. Using single-cell RNA sequencing (scRNA-seq), we investigated the cellular and molecular mechanisms underlying color change in Rhacophorus dugritei, which adapted to its background by displaying light-green skin on white and black skin on black within two days. We identified two types of chromatophores in their skin, both responsible for the observed color differences. Our findings reveal that morphological color change (MCC) is the dominant process, with the number of chromatophores being more influential in driving color change than the transcriptional activity of melanogenesis in individual cells. Additionally, melanophores from darker individuals exhibited increased activity in energy metabolism pathways, while those from lighter individuals showed stronger immune-related gene expression, suggesting that background adaptation involves more than just morphological changes. Overall, this study successfully applied single-cell sequencing technology to investigate skin pigmentation in a non-model organism. Our results suggest that MCC driven by chromatophore proliferation is a key mechanism of background adaptation, offering new insights into amphibian color adaptation and environmental adaptation in other vertebrates.