The ability to control the interfacial properties in metal-oxide thin films through surface defect engineering is vital to fine tune their optoelectronic properties and thus their integration in novel optoelectronic devices. This is exemplified in photovoltaic devices based on organic, inorganic or hybrid technologies, where precise control of the charge transport properties through the interfacial layer is highly important for improving device performance. In this work, we study the effects of in situ annealing in nearly stoichiometric MoO x (x~3.0) thin-films deposited by reactive sputtering. We report on a work function increase of almost 2 eV after inducing in situ crystallization of the films at 500°C, resulting in the formation of a single crystalline α-MoO 3 overlaid by substoichiometric and highly disordered nanoaggregates. The surface nanoaggregates possess various electronic properties, such as a work function ranging from 5.5 eV up to 6.2 eV, as determined from low-energy electron microscopy studies. The crystalline underlayer possesses a work function greater than 6.3 eV, up to 6.9 eV, characteristic of a very clean and nearly defect-free MoO 3. By combining electronic spectroscopies together with structural characterizations, this work addresses a novel method for tuning, and correlating, the optoelectronic properties and microstructure of device-relevant MoO x layers.