The Error-related Negativity (ERN) currently appears as a physiological measure in relation to three RDoC constructs: Cognitive Control, Sustained Threat, and Reward Learning. We propose a conceptual model in which variance in the ERN reflects individual differences in the degree to which errors are evaluated as threatening. We also discuss evidence for the placement of the ERN in the ‘Sustained Threat’ construct, as well as evidence that the ERN may more specifically reflect sensitivity to endogenous threat. Following this, we present data from a sample of 515 adolescent females demonstrating larger ERN in relation to self-reported checking behaviors, but only in older adolescents, suggesting that sensitivity to internal threat and the ERN-checking relationship may follow a developmental course as adolescents develop behavioral control. In contrast, depressive symptoms were linked to smaller ERN, and this association was invariant with respect to age. Collectively, these data suggest that the magnitude of the ERN is sensitive both to specific anxiety-related processes and depression, in opposing directions that may reflect variation in internal threat sensitivity. We discuss directions for future research, as well as ways in which findings for the ERN complement and challenge aspects of the current RDoC matrix.