Supplementary Material for: Elevated testosterone is required for male sexual behaviour and dynamic colour change in veiled chameleons

Introduction: Sex-specific genotype and early organization can influence the expression of sexually dimorphic traits in vertebrates. We tested these hypotheses in male-typical behaviour and rapid change to bright colouration in the veiled chameleon (Chamaeleo calyptratus) with XX/XY sex chromosomes. Methods: Hormonal manipulations included castration with and without testosterone replacement, and testosterone administration in females. Results: Long-term testosterone treatment induced male-typical sexual behaviour and an ability to switch to bright colouration in females, while castration suppressed these traits in males. These observations document that elevated testosterone alone is sufficient for the expression of these traits in both males and females. Surprisingly, high testosterone levels led to indiscriminate courtship behaviour, with frequent mating attempts directed at conspecifics regardless of their sex and testosterone level in both home cages and neutral arenas. This unexpected behaviour suggests that visual cues, such as body and head-casque size, may not reliably guide sex recognition during short distance encounters. Conclusion: The dependence of the male-typical sexual behaviour and colour change on the elevated androgen levels contrast sharply with earlier results on skeletal traits (body size and head-casque size), which are fully developed in castrated males, demonstrating that the ontogeny of the sex-typical phenotype involves different mechanisms in the emerging model species of chameleons.