The body size of a univoltine carabid beetle Carabus tosanus on Shikoku Island, Japan, was clearly smaller in higher-altitude populations (subspecies), which possibly represents incipient speciation. To explore the determinants of altitudinal differences in body size in this species, we studied the degree of phenotypic plasticity by conducting rearing experiments at two constant temperatures and examined genetic differences through interpopulation crosses. At 15 °C, C. tosanus had a longer developmental period and a shorter adult body than at 20 °C. Nevertheless, variation in body size due to temperature effects (phenotypic plasticity) was small compared to the interpopulation differences, which suggests substantial genetic differences between populations (subspecies) at different altitudes. In F1 offspring from crosses between a low-altitude (subspecies tosanus) and a high-altitude population (subspecies ishizuchianus), adult body length was affected by the genotypes of both parents, with an interaction effect of parental genotype and offspring sex. Further analyses revealed that adult body length was affected by sex-linked factors in addition to autosomal factors. These genetic differences in body size may have resulted from adaptations to different altitudes and may be important for the process of incipient speciation because body size differences could contribute to premating reproductive isolation.