B-cell lymphoma 2 (Bcl-2) proteins are central, conserved regulators of apoptosis. Bcl-2 family function is regulated by binding interactions between the Bcl-2 homology 3 (BH3) motif in pro-apoptotic family members and the BH3 binding groove found in both the pro-apoptotic effector and anti-apoptotic Bcl-2 family members. A novel motif, the reverse BH3 (rBH3), has been shown to interact with the anti-apoptotic Bcl-2 homolog MCL1 (Myeloid cell leukemia 1) and have been identified in the p53 homolog p73, and the CDK4/6 (cyclin dependent kinase 4/6) inhibitor p18INK4c, (p18, cyclin-dependent kinase 4 inhibitor c). To determine the conservation of rBH3 motif, we first assessed conservation of MCL1's BH3 binding groove, where the motif binds. We then constructed neighbor-joining phylogenetic trees of the INK4 and p53 protein families and analyzed sequence conservation using sequence logos of the rBH3 locus. This showed the rBH3 motif is conserved throughout jawed vertebrates p63 and p73 sequences and in chondrichthyans, amphibians, mammals, and some reptiles in p18. Finally, a potential rBH3 motif was identified in mammalian and osteichthyan p19INK4d (p19, cyclin dependent kinase 4 inhibitor d). These findings demonstrate that the interaction between MCL1 and other cellular proteins mediated by the rBH3 motif may be conserved throughout jawed vertebrates.