Abstract Background Differentiating dental pulp stem cells (DPSCs) into odontoblasts is a critical process for tooth self-repair and dentine‒pulp engineering strategies in the clinic. However, the mechanism underlying the regulation of DPSC odontoblastic differentiation remains largely unknown. Here, we demonstrated that BCL-2 interacting protein 3 (BNIP3)-dependent mitophagy is associated with importin subunit beta-1 (KPNB1)-activating transcription factor 4 (ATF4), which promotes DPSC odontoblastic differentiation. Methods The key genes involved in DPSC odontogenic differentiation were identified via bioinformatics. Stable silencing or overexpression of BNIP3 was performed to investigate its impact on DPSC differentiation in vitro (n ≥ 3). To explore the role of BNIP3 in vivo, tooth root fragments loaded with the hydrogel-transfected DPSC complex were implanted into nude mice (n ≥ 6). Dual-luciferase reporter assays and chromatin immunoprecipitation (ChIP) polymerase chain reaction (PCR) were conducted to explore the binding site of ATF4 to the BNIP3 promoter (n ≥ 3). Mitochondrial function experiments were performed to investigate the impact of ATF4-BNIP3 on mitochondria (n ≥ 3). Immunoprecipitation (IP) mass spectrometry (MS) was used to investigate the interaction between ATF4 and its binding protein, KPNB1. Plasmids containing wild-type (WT)/mutant (MUT)-nuclear localization signal (NLS) forms of ATF4 were constructed to determine the specific amino acid residues recognized by KPNB1 and their effects on DPSC odontoblastic differentiation (n ≥ 3). Results Compared with those in the control group, the levels of autophagy and mitophagy, especially BNIP3-dependent mitophagy, were greater in the DPSC odontoblastic differentiation group (P