INTRODUCTION
Colour is one of the most striking and varied components of visual signals throughout the natural world. Many colours are produced by pigments, which can be directly or indirectly obtained from the diet, or alternatively, synthesised by the body (McGraw 2005). Carotenoids are the main class of dietary pigments and are the primary class of pigments producing yellow to red coloration in birds; however, the majority of vertebrates (ectothermic vertebrates – fish, reptiles, amphibians) can also produce yellow to red coloration using a biochemically distinct class of pigments called pteridines (Bagnara & Matsumoto 2006). Pteridines are synthesised de novo within pigment cells from abundant purine molecules (Bracher et al. 1998; Ziegler 2003; Braasch et al. 2007). Pteridines can be used instead of, or together with carotenoids to produce yellow-red colours and the two pigment classes can frequently be found together within xanthophore pigment cells (Bagnara & Hadley 1973; Bagnara & Matsumoto 2006). However, in sharp contrast to carotenoids, the evolutionary drivers of variation among species in pteridine pigments remain largely unknown.
What explains the use of carotenoid or pteridine pigments when both can produce yellow to red colours? One compelling but unsubstantiated possibility is that pteridines replace carotenoids in environments where carotenoid availability is limited. Carotenoid limitation is expected to alter the relative cost of acquiring and sequestering carotenoids compared to synthesising pteridines. Specifically, when carotenoids are rare, it may be metabolically cheaper to synthesise pteridines; whereas when carotenoids are abundant, it may be metabolically cheaper to acquire, transport and sequester carotenoids (Grether et al.2001; Grether et al. 2005).
Within the broad classes of carotenoids and pteridines, specific pigments have different hues, are acquired or metabolised in different ways and therefore have different costs and roles in colour production. Carotenoids are produced by plants and the most dominant carotenoids in angiosperms are yellow xanthophylls such as lutein (Heath et al.2013). Insect herbivores generally sequester carotenoids in proportion to the concentration found in the diet (Heath et al. 2013). Red ketocarotenoids, such as astaxanthin and canthaxanthin are comparatively rare in terrestrial ecosystems (primarily produced by microalgae and yeast), but some animals, including birds and turtles, can metabolically convert dietary yellow carotenoids to red ketocarotenoids (Lopeset al. 2016; Mundy et al. 2016; Twyman et al.2016). Due to the cost of metabolic conversion, or low dietary availability for terrestrial animals, ketocarotenoids are more strongly associated with measures of individual quality and sexual selection than dietary yellow carotenoids, particularly in birds (Weaver et al.2018). Pteridines similarly vary in colour from yellow (e.g. sepiapterin, xanthopterin) to red (e.g. drosopterin, erythropterin). Other pteridines (e.g. pterin, isoxanthopterin) are often assumed to be colourless but may take crystalline form, and by virtue of their crystallinity, contribute to integument coloration through reflection and scattering of light, rather than absorption (Oliphant & Hudon 1993; Palmer et al. 2018; Palmer et al. 2020). These other pteridines can be found in large quantities within xanthophores (Bagnara & Matsumoto 2006; McLean et al. 2017; McLean et al. 2019; Twomey et al. 2020b), suggesting that they may contribute to integument coloration. The different costs and roles in colour production for different types of carotenoids and pteridines influence expected associations with environmental factors and sexual selection. However, the ecological and evolutionary drivers of pigment variation remain unknown, with the exception of carotenoids in some groups of birds (Prum et al. 2012; Friedman et al. 2014b, a; Ligonet al. 2016).
Here, using an extensive dataset of concentrations of 5 carotenoid and 6 pteridine pigments, we test whether pigment concentrations are associated with environmental gradients indicative of carotenoid availability among 27 species of Australian agamid lizards (186 skin samples, 79 individuals, 28 populations with distinct coloration). Specifically, we use highly accurate liquid chromatography-mass spectrometry to quantify pigment concentrations in skin tissues of agamid lizards (McLean et al.2017; McLean et al. 2019). In addition to testing for associations with environmental gradients, we tested whether pteridines replace carotenoids with a similar hue (carotenoid-mimicry hypothesis) (Grether et al. 1999), resulting in a negative correlation between the concentrations of similarly coloured carotenoid and pteridine pigments. Since concentrations of different pigment types may depend on the strength of sexual selection, we simultaneously tested for relationships between pigment concentrations and proxies for the strength of sexual selection (sexual dichromatism and sexual size dimorphism). Additionally, we evaluated how carotenoid or pteridine concentrations covary with skin colour (hue, saturation, luminance). We show that environmental gradients can predict the use of biochemically different classes of pigment for integument coloration in vertebrates.