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Simple Summary: Our study confirmed that Bmp53 can directly induce cell apoptosis and regulate the morphology and development of larvae during pupation. Y2H-Seq data led to the development of a model for apoptosis regulation with Bmp53 as a central node and the prediction that the silkworm Mdm2-like protein may be a key component of the apoptotic pathway. These results provide the initial molecular foundation for elucidating apoptosis regulation in silkworms and assessing the various biological processes regulated by the Bmp53 interactome. The apoptosis pathway is highly conserved between invertebrates and mammals. Although genes encoding the classical apoptosis pathway can be found in the silkworm genome, the regulatory pathway and other apoptotic network genes have yet to be confirmed. Consequently, characterizing these genes and their underlying mechanisms could provide critical insights into the molecular basis of organ apoptosis and remodeling. A homolog of p53, a key apoptosis regulator in vertebrates, has been identified and cloned from Bombyx mori (Bmp53). This study confirmed via gene knockdown and overexpression that Bmp53 directly induces cell apoptosis and regulates the morphology and development of individuals during the metamorphosis stage. Furthermore, yeast two-hybrid sequencing (Y2H-Seq) identified several potential apoptotic regulatory interacting proteins, including the MDM2-like ubiquitination regulatory protein, which may represent an apoptosis factor unique to Bmp53 and which differs from that in other Lepidoptera. These results provide a theoretical basis for analyzing the various biological processes regulated by Bmp53 interaction groups and thus provide insight into the regulation of apoptosis in silkworms. The global interaction set identified in this study also provides a basic framework for future studies on apoptosis-dependent pupation in Lepidoptera. [ABSTRACT FROM AUTHOR]