Prior literature has pointed out that organizational culture is an important factor affecting knowledge sharing and enhanced innovation in an organization. However, there is a lack of models linking organizational culture, knowledge sharing, and innovation at universities within developing countries, particularly Syria. The purpose of this study was to examine the mediating role of knowledge sharing on the relationship between organizational culture and innovation of the teaching staff in Syria public and private universities. The quantitative method was used in data collection. A random sampling technique was conducted which comprised of 334 valid responses to test the causal relationship between organizational culture, knowledge sharing, and innovation. PLS-SEM technique was used to analyze the direct and indirect relationships between the variables in this study. The results of the study indicated that there was a significant relationship between organizational culture and Knowledge sharing. The results show that there was a significant relationship between organizational culture and innovation. Findings had revealed that the indirect effect of organizational culture on innovation through knowledge sharing.