Creating breakthrough innovation (BI) is a crucial endeavor for contemporary companies to survive given the highly uncertain and competitive environment. Several scholars have explored the factors that facilitate BI in established companies and identified that human resource management (HRM) practices have significant impact on innovation. However, as the relationship between HRM practices and innovation includes several causal links at the organizational and individual-level, innovation management scholars have not fully examined each linkage. To fill this gap, this study examines the impact of HRM practices on the individual-level BI behavior by employing the Ability-Motivation-Opportunity (AMO) framework as a theoretical lens. Specifically, we developed four hypotheses that were tested using empirical analysis, with survey data from 307 individuals involved in BI projects at large companies in Japan. Our data analysis reveals that innovation talent development and BI role clarity impact employees’ BI behavior and senior management support moderates the impact of extrinsic rewards. These findings contribute to the existing literature by elaborating the underlying mechanism of the AMO framework and provide practical implications for the design of HRM systems for BI success.