The paper presents a numerical activity directed at the analysis and optimization of internal combustion engine water cooling jackets, with particular emphasis on the fatigue-strength assessment and improvement. In the paper, full 3D-CFD and FEM analyses of conjugate heat transfer and load cycle under actual engine operation of a single bank of a current production V6 turbocharged Diesel engine are reported. A highly detailed model of the engine, made up of both the coolant galleries and the surrounding metal components, i.e. the engine head, the engine block, the gasket, the valve guides and valve seats, is used on both sides of the simulation process to accurately capture the influence of the cooling system layout under thermal and load conditions as close as possible to actual engine operations. Concerning the CFD side, a 50-50 binary mixture of ethylene-glycol and water is used in order to correctly reproduce the coolant behavior, while boundary conditions are derived from a combination of experimental measurements and a CFD-1D model of the whole engine. In order to find a proper CFD setup for the optimization of the thermal behavior of the engine, a preliminary comparison between experimental temperature distribution within the engine head and CFD forecasts is carried out. Eight thermocouples are used to measure the engine head local temperature at some critical locations. Among the many competing numerical sub-models involved in the CFD simulations, particular attention is devoted to the modeling of phase transition and vapor nuclei formation within the coolant galleries. Concerning the FEM side, thermo-mechanical analyses are carried out aiming at addressing the design optimization of the engine in terms of fatigue strength. In view of the wide range of thermal and load conditions, both high-cycle and low-cycle fatigue must be properly characterized by means of ad-hoc criteria. An energy-based criterion specifically suited for low-cycle fatigue regions is therefore superimposed to well-established S-N o ε-N criteria for the high cycle fatigue regions. The proposed methodology shows very promising results in terms of point-wise detection of possible engine failures ans proves to be an effective tool for the accurate thermo-mechanical characterization of internal combustion engines under actual life-cycle operations.