Drosophila melanogaster courtship, although stereotypical, continually changes based on cues received from the courtship subject. Such adaptive responses are mediated via rapid and widespread transcriptomic reprogramming, a characteristic now widely attributed to microRNAs (miRNAs), along with other players. Here, we conducted a large-scale miRNA knockout screen to identify miRNAs that affect various parameters of male courtship behavior. Apart from identifying miRNAs that impact male-female courtship, we observed that miR-957 mutants performed significantly increased male-male courtship and "chaining" behavior, whereby groups of males court one another. We tested the effect of miR-957 reduction in specific neuronal cell clusters, identifying miR-957 activity in Doublesex (DSX)-expressing and mushroom body clusters as an important regulator of male-male courtship interactions. We further characterized the behavior of miR-957 mutants and found that these males court male subjects vigorously, but do not elicit courtship. Moreover, they fail to lower courtship efforts toward females with higher levels of antiaphrodisiac pheromones. At the level of individual pheromones, miR-957 males show a reduced inhibitory response to both 7-Tricosene (7-T) and cis-vaccenyl acetate, with the effect being more pronounced in the case of 7-T. Overall, our results indicate that a single miRNA can contribute to the regulation of complex behaviors, including detection or processing of chemicals that control important survival strategies such as chemical mate-guarding, and the maintenance of sex- and species-specific courtship barriers.