<p>Despite the critical role teachers play in AI education, research on their understanding of generative AI tools remains limited, particularly concerning their ethical assessment knowledge. This study addresses the gap by proposing the Generative AI Technological Pedagogical Content Knowledge (GenAI-TPACK) framework, which incorporates ethical knowledge as an essential component of university teachers’ professional development. Grounded in the Technological Pedagogical Content Knowledge (TPACK) model, we utilize structural equation modeling to investigate the interrelationships among Generative AI Technological Knowledge, pedagogical knowledge, and ethical assessment knowledge. Our findings indicate that university teachers’ effective use of generative AI is contingent upon their technological knowledge and engagement with these tools. Furthermore, while technical knowledge supports evaluative decision-making, it is insufficient on its own for successful integration into teaching practices. Instead, a holistic approach that combines technological and pedagogical knowledge enhances teachers’ capacities to utilize generative AI effectively. The study also reveals a positive correlation between ethical assessment knowledge and both pedagogical and content knowledge, contributing to a comprehensive understanding of the GenAI-TPACK framework. This research underscores the importance of integrating ethical considerations into AI education, offering valuable insights for the professional development of educators and the enhancement of teacher training programs.</p>