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This systematic review examines whether negative affect symptoms (i.e., anxiety, depression, and internalizing symptoms more broadly) predict subsequent adolescent substance use after controlling for co-occurring externalizing symptoms. Following PRISMA procedures, we identified 61 studies that tested the association of interest. Findings varied depending on the type of negative affect symptom and to some extent on the substance use outcome. The most consistent associations were evident for depressive symptoms, particularly as predictors of substance use composite scores. No clear association between anxiety and substance use or between internalizing symptoms and substance use was evident, and indeed these associations were as often negative as positive. Mixed findings regarding the depression–substance use association, however, also call for greater attention to potential moderating factors that may help define who, when, and in what context depression serves as an important risk factor for later substance use above and beyond risk associated with externalizing symptoms.