This study used a short-term longitudinal design with theoretically derived preregistered hypotheses and analyses to examine the role of temperament in the development of forms (i.e., physical and relational) and functions (i.e., proactive and reactive) of aggressive behavior in early childhood (N = 300, M age = 44.70 months, SD = 4.38, 44% girls). Temperament was measured via behavioral reports of emotional dysregulation, fearlessness/daring, and rule internalization/empathy and, in a subsample that completed a physiological assessment, via skin conductance and respiratory sinus arrhythmia. Emotion dysregulation generally served as a risk factor for all subtypes of aggression, with evidence of stronger associations with reactive as compared to proactive functions of relational aggression for girls. Daring predicted increases in physical aggression, especially among boys, and rule internalization predicted decreases in relational aggression, especially among girls. Rule internalization mediated longitudinal associations between daring and proactive relational aggression for girls. Some evidence also emerged supporting associations between adaptive functioning (i.e., high empathy, high respiratory sinus arrhythmia) and proactive functions of aggression. Findings highlight distinct temperamental risk factors for physical versus relational aggression and provide partial support for gender-linked theories of the development of aggression.