Enhancing adoptive cell therapies to boost their anti-tumour effectiveness is a major focus of the cancer immunotherapy field. One approach gaining interest is overexpressing the transcription factor, AP-1, in CAR T cells to overcome regulatory processes like exhaustion. AP-1 is a family of dimeric TFs, primarily composed of Jun and Fos proteins, which is assumed to play a crucial role in T cell differentiation through both promoting cytokine production and suppressing regulatory processes. However, there is surprisingly little functional evidence to support these assumptions. In fact, given the large number of AP-1 family members, no study has fully knocked out AP-1 to assess its contribution to CD8+ T cell differentiation in vivo. We recently developed a novel CRISPR-Cas9 gene editing method enabling rapid generation of knock-out (KO) T cells. Using this technique, we optimised an approach to fully ablate all Jun proteins (i.e. cJun, JunB and JunD) in naïve CD8+ T cells, thereby eliminating all potential AP-1 complexes. During acute viral infection, AP-1 KO CD8+ T cells display a survival defect compared to control cells, with the surviving cells also having decreased expression of immune memory markers and increased expression of exhaustion markers. Unlike exhausted cells, though, KO cells retained cytokine effector function, suggesting that AP-1 may be fully dispensable for T cell cytokine production. KO cells also failed to acquire features of peripheral tolerance, a regulatory state that AP-1 was proposed to suppress. Interestingly, KO of JunB alone was sufficient to recapitulate the AP-1 KO cell phenotype, with KO of c-Jun and/or JunD having little impact on T cell differentiation. Thus, contrary to current dogma, AP-1 represses aspects of the exhausted fate but does not impact peripheral tolerance or acquisition of T cell effector functions. Moreover, only JunB-containing AP-1 complexes appear functionally important in normal effector T cell differentiation.