Department of Earth and Environmental Sciences, University of Pavia, Via Taramelli 24, 27100, Pavia, Italy.
The Armenise-Harvard Laboratory of Structural Biology, Department of Biology and Biotechnology "L. Spallanzani", University of Pavia, Via Ferrata 9A, 27100, Pavia, Italy.
Sci Rep. 2023 Aug 29;13(1):14164. doi: 10.1038/s41598-023-41012-9.
The evolution of chemical signals is subject to environmental constraints. A multicomponent signal may combine semiochemical molecules with supporting compounds able to enhance communication efficacy. Carbonic anhydrases (CAs) are ubiquitous enzymes catalysing the reversible hydration of carbon dioxide, a reaction involved in a variety of physiological processes as it controls the chemical environment of the different tissues or cellular compartments, thus contributing to the overall system homeostasis. CA-IV isoform has been recently identified by mass spectrometry in the femoral gland secretions (FG) of the marine iguana, where it has been hypothesized to contribute to the chemical stability of the signal, by regulating blend pH. Lizards, indeed, use FG to communicate by delivering the waxy secretion on bare substrate, where it is exposed to environmental stressors. Therefore, we expect that some molecules in the mixture may play supporting functions, enhancing the stability of the chemical environment, or even conferring homeostatic properties to the blend. CA-IV may well represent an important candidate to this hypothesized supporting/homeostatic function, and, therefore, we can expect it to be common in FG secretions of other lizard species. To evaluate this prediction and definitely validate CA identity, we analysed FG secretions of eight species of wall lizards (genus Podarcis), combining mass spectrometry, immunoblotting, immunocytochemistry, and transmission electron microscopy. We demonstrate CA-IV to actually occur in the FG of seven out of the eight considered species, providing an immunochemistry validation of mass-spectrometry identifications, and localizing the enzyme within the secretion mass. The predicted structure of the identified CA is compatible with the known enzymatic activity of CA-IV, supporting the hypothesis that CA play a signal homeostasis function and opening to new perspective about the role of proteins in vertebrate chemical communication.
化学信号的进化受到环境的限制。一个多成分的信号可能会将半化学分子与能够增强通讯效果的支持化合物结合起来。碳酸酐酶（CA）是一种普遍存在的酶，能够催化二氧化碳的可逆水合反应，该反应参与了多种生理过程，因为它控制着不同组织或细胞区室的化学环境，从而有助于整个系统的内稳态。最近，通过质谱法在海洋鬣蜥的股腺分泌物（FG）中鉴定出 CA-IV 同工型，据推测，它通过调节混合物的 pH 值，有助于信号的化学稳定性。蜥蜴确实通过将蜡状分泌物涂在裸露的基质上来进行 FG 通讯，在那里它会受到环境胁迫的影响。因此，我们预计混合物中的某些分子可能具有支持功能，增强化学环境的稳定性，甚至赋予混合物内稳态特性。CA-IV 很可能是这种假设的支持/内稳态功能的重要候选者，因此，我们可以预期它会存在于其他蜥蜴物种的 FG 分泌物中。为了评估这一预测并明确验证 CA 的身份，我们分析了 8 种壁蜥（Podarcis 属）的 FG 分泌物，结合了质谱、免疫印迹、免疫细胞化学和透射电子显微镜。我们证明 CA-IV 实际上存在于 8 种被考虑的物种中的 7 种 FG 中，提供了对质谱鉴定的免疫化学验证，并在分泌质量内定位了该酶。鉴定出的 CA 的预测结构与 CA-IV 的已知酶活性兼容，支持 CA 发挥信号内稳态功能的假设，并为脊椎动物化学通讯中蛋白质的作用开辟了新的视角。
