Interferon (IFN)‐induced signalling pathways have essential functions in innate immune responses. In response to type I IFNs, filamin B tethers RAC1 and a Jun N‐terminal kinase (JNK)‐specific mitogen‐activated protein kinase (MAPK) module—MEKK1, MKK4 and JNK—and thereby promotes the activation of JNK and JNK‐mediated apoptosis. Here, we show that type I IFNs induce the conjugation of filamin B by interferon‐stimulated gene 15 (ISG15). ISGylation of filamin B led to the release of RAC1, MEKK1 and MKK4 from the scaffold protein and thus to the prevention of sequential activation of the JNK cascade. By contrast, blockade of filamin B ISGylation by substitution of Lys 2467 with arginine or by knockdown of ubiquitin‐activating enzyme E1‐like (UBEL1) prevented the release of the signalling molecules from filamin B, resulting in persistent promotion of JNK activation and JNK‐mediated apoptosis. These results indicate that filamin B ISGylation acts as a negative feedback regulatory gate for the desensitization of type I IFN‐induced JNK signalling.