miR-iab-4-3p fine-tunes JNK signaling via a negative feedback loop targeting kay in Drosophila

The c-Jun N-terminal kinase (JNK) signaling pathway governs multiple developmental and physiological processes in Drosophila, yet its post-transcriptional regulation remains largely unexplored. Here, we identify miR-iab-4-3p as a novel negative regulator of JNK signaling. Overexpression of miR-iab-4 rescues the eye and wing defects caused by ectopic activation of the JNK pathway. Genetic epistasis analyses position miR-iab-4 downstream of the kinase Hep but upstream of the AP-1 transcription factor complex. Mechanistically, miR-iab-4-3p directly binds to the 3'-untranslated region (UTR) of kay, which encodes the AP-1 component Kay, thereby suppressing Kay expression and attenuating JNK transcriptional output. Furthermore, we demonstrate that AP-1 directly activates miR-iab-4-3p transcription, uncovering a negative feedback loop wherein JNK-induced AP-1 drives miR-iab-4-3p expression, which in turn represses Kay to limit signaling activity. Our findings reveal a post-transcriptional layer of JNK regulation and establish miR-iab-4-3p as a key node in maintaining JNK signal homeostasis via a negative feedback circuit.