The Reinforcement Sensitivity Theory, first proposed by Gray and later revised, describes three motivational systems: Behavioral Inhibition System (BIS), Behavioral Activation System (BAS), and Fight-Flight-Freeze System (FFFS). Studies have shown that high BIS and FFFS activation are positively related to anxiety symptoms, yet the relationship between BAS and anxiety remains unclear. Research data have also suggested that anxiety symptoms occur with the loss of perceived control. Thus, although studies on the direct effect of locus of control (LOC) on trait anxiety have accumulated for many years, the issue of how LOC may mediate the relationship between BIS/BAS/FFFS sensitivity and anxiety has not been addressed. This study aimed to explore the mediating role of LOC orientation on trait anxiety among young adults in association with these three motivational systems. Cross-sectional data were obtained from 422 volunteers. The BIS/BAS Scale, Trait Anxiety Inventory, and Rotter’s Internal-External LOC Scale were applied. A series of mediation analyses were performed to estimate total, indirect, and direct effects. The results showed that BIS and FFFS positively predicted trait anxiety. In addition, LOC positively predicted trait anxiety and BIS. The results of the mediation analyses indicated that LOC functioned as a partial mediator between BIS and trait anxiety. This finding revealed that a high BIS level, one of the motivational systems, was associated with external LOC, which in turn contributed to reporting high trait anxiety in young adults. Hence, BIS and external LOC orientation could be suggested as risk factors for trait anxiety. As the external LOC orientation of individuals with high punishment sensitivity increased, their trait anxiety levels also increased. Therefore, it was suggested that it might be useful to be aware that LOC orientations of individuals with BIS sensitivity may pose a risk for trait anxiety.