Enzyme‐Instructed Assemblies Enable Mitochondria Localization of Histone H2B in Cancer Cells

Abstract Presently, little is known of how the inter‐organelle crosstalk impacts cancer cells owing to the lack of approaches that can manipulate inter‐organelle communication in cancer cells. We found that a negatively charged, enzyme cleavable peptide (MitoFlag) enables the trafficking of histone protein H2B, a nuclear protein, to the mitochondria in cancer cells. MitoFlag interacts with the nuclear location sequence of H2B to block it from entering the nucleus. A protease on the mitochondria cleaves the Flag from the MitoFlag/H2B complex to form assemblies that retain H2B on the mitochondria and facilitate H2B entering the mitochondria. Adding NLS, replacing aspartic acid by glutamic acid residues, or changing the l ‐ to d ‐aspartic acid residue on MitoFlag abolishes the trafficking of H2B into mitochondria of HeLa cells. As the first example of the enzyme‐instructed self‐assembly of a synthetic peptide for trafficking endogenous proteins, this work provides insights for understanding and manipulating inter‐organelle communication in cells.