As climate change increasingly affects forest ecosystems, detailed understanding of major effects is important to anticipate their consequences under future climate scenarios. The Mediterranean region is a prominent climate change hotspot, and evergreen cork oak (Quercus suber L.) woodlands are particularly climatically sensitive due to cork (bark) harvesting. Cork oak’s drought avoidance strategy is well-known and includes structural and physiological adaptations that maximise soil water uptake and transport and limit water use, potentially leading to reduced stem and cork growth. Trees’ responses to cope with water-limited conditions have been extensively described based on cork-rings width and, more recently, on cork-rings density, in dendroecological studies. However, so far, tree functional attributes and physiological strategies, namely photosynthetic metabolism adjustments affecting cork formation, have never been addressed and/or integrated on these previous cork-rings-based studies. In this study, we address the relation between carbon and oxygen stable isotopes of cork rings and precipitation and temperature, in two distinct locations of southwestern Portugal–the (wetter) Tagus basin peneplain and the (drier) Grândola mountains. We aimed at assessing whether the two climatic factors affect cork-ring isotopic composition under contrasting conditions of water availability, and, therefore, if carbon and oxygen signatures in cork can reflect tree functional (physiological and structural) responses to stressful conditions, which might be aggravated by climate change. Our results indicate differences between the study areas. At the drier site, the stronger statistically significant negative cork δ13C correlations were found with mean temperature, whereas strong positive cork δ18O correlations were fewer and found only with precipitation. Moreover, at the wetter site, cork rings are enriched in 18O and depleted in 13C, indicating, respectively, shallow groundwater as the water source for physiological processes related with biosynthesis of non-photosynthetic secondary tissues, such as suberin, and a weak stomatal regulation under high water availability, consistent with non-existent water availability constrains. In contrast, at the drier site, trees use water from deeper ground layers, depleted in 18O, and strongly regulate stomatal conductance under water stress, thus reducing photosynthetic carbon uptake and probably relying on stored carbon reserves for cork ring formation. These results suggest that although stable isotopes signatures in cork rings are not proxies for net growth, they may be (fairly) robust indicators of trees’ physiological and structural adjustments to climate and environmental changes in Mediterranean environments.