Holm oak (Quercus ilex subsp. rotundifolia Lam.) is an evergreen tree that experiences a physiological deterioration of the photosynthetic apparatus associated with age-dependent changes in net photosynthesis (AN), stomatal conductance (gs), mesophyll conductance (gm) and chloroplast surface area exposed to intercellular air space (Sc/S). However, it remains unclear what the time course is and what physiological factors are associated with this phenomenon in old non-senescent leaves of evergreen trees with long leaf lifespan. We evaluated the evolution of anatomical, biochemical, hydraulic and photosynthetic traits during three consecutive vegetative periods in current, 1- and 2-year-old leaves of holm oak. Current-year leaves reached their maximum photosynthetic activity during early summer after the coordinated maturation of morphoanatomical traits and the ability for CO2 diffusion and fixation in the chloroplasts. Thereafter, we found a gradual downregulation of photosynthetic activity throughout the leaf lifespan that was explained by the reduction in (1) gs and gm, (2) electron transport rate (Jflu), and (3) carboxylation capacity of Rubisco (Vcmax). Ultimately, 2-year-old leaves experienced during summer an abrupt decrease in leaf hydraulic conductance (Kleaf), which indicated a coordination between AN and Kleaf that might be mediated by cell-level anatomical traits such as Sc/S. These findings first demonstrated that leaf hydraulic limitations (i.e., Kleaf) are associated with the dismantling of the photosynthetic apparatus in older leaves of evergreen tree species with long leaf lifespan such as holm oak. Contrary to expectations, bulk leaf ABA did not appear to play a main role as a signaling molecule in this process.