Significance: Membrane fusion is essential in multiple cell processes, including neuronal communication. Numerous proteins are recruited to the presynaptic space to execute a highly controlled process, resulting in the liberation of neurotransmitters. Many of these proteins share C2 domains as common structural motifs, regulated by their ability to bind Ca ²⁺ , phospholipids, and other proteins, endowing them with properties to fine-tune a wide variety of vesicle release modes. Here, by solving the structures of Rabphilin-3A (Rph3A) C2B–SNAP25 and Rph3A C2B–PIP ₂ complexes, we revealed a membrane-binding mode in which the Rph3A-C2 domains operate in cooperation with PIP ₂ /Ca ²⁺ and SNAP25, adopting a conformation able to promote membrane bending, suggesting a model to explain how Rph3A regulates various steps of the vesicle fusion process.

Protein kinase C (PKC) comprises a family of highly related serine/threonine protein kinases involved in multiple signaling pathways, which control cell proliferation, survival, and differentiation. The role of PKCα in cancer has been studied for many years. However, it has been impossible to establish whether PKCα acts as an oncogene or a tumor suppressor. Here, we analyzed the importance of PKCα in cellular processes such as proliferation, migration, or apoptosis by inhibiting its gene expression in a luminal A breast cancer cell line (MCF-7). Differential expression analysis and phospho-kinase arrays of PKCα-KD vs. PKCα-WT MCF-7 cells identified an essential set of proteins and oncogenic kinases of the JAK/STAT and PI3K/AKT pathways that were down-regulated, whereas IGF1R, ERK1/2, and p53 were up-regulated. In addition, unexpected genes related to the interferon pathway appeared down-regulated, while PLC, ERBB4, or PDGFA displayed up-regulated. The integration of this information clearly showed us the usefulness of inhibiting a multifunctional kinase-like PKCα in the first step to control the tumor phenotype. Then allowing us to design a possible selection of specific inhibitors for the unexpected up-regulated pathways to further provide a second step of treatment to inhibit the proliferation and migration of MCF-7 cells. The results of this study suggest that PKCα plays an oncogenic role in this type of breast cancer model. In addition, it reveals the signaling mode of PKCα at both gene expression and kinase activation. In this way, a wide range of proteins can implement a new strategy to fine-tune the control of crucial functions in these cells and pave the way for designing targeted cancer therapies.