title: Competitive Inhibition of the Endoplasmic Reticulum Signal Peptidase by Non-cleavable Mutant Preprotein Cargos.
abstract: Upon translocation across the endoplasmic reticulum (ER) membrane, secretory proteins are proteolytically processed to remove their signal peptide by signal peptidase (SPase). This process is critical for subsequent folding, intracellular trafficking, and maturation of secretory proteins. Prokaryotic SPase has been shown to be a promising antibiotic target. In contrast, to date, no eukaryotic SPase inhibitors have been reported. Here we report that introducing a proline immediately following the natural signal peptide cleavage site not only blocks preprotein cleavage but also, in trans, impairs the processing and maturation of co-expressed preproteins in the ER. Specifically, we find that a variant preproinsulin, pPI-F25P, is translocated across the ER membrane, where it binds to the catalytic SPase subunit SEC11A, inhibiting SPase activity in a dose-dependent manner. Similar findings were obtained with an analogous variant of preproparathyroid hormone, demonstrating that inhibition of the SPase does not depend strictly on the sequence or structure of the downstream mature protein. We further show that inhibiting SPase in the ER impairs intracellular processing of viral polypeptides and their subsequent maturation. These observations suggest that eukaryotic SPases (including the human ortholog) are, in principle, suitable therapeutic targets for antiviral drug design.
authors.affiliation: Department of Endocrinology and Metabolism, Tianjin Medical University General Hospital, Tianjin 300052, China,; Division of Metabolism, Endocrinology, and Diabetes, University of Michigan Medical School, Ann Arbor, Michigan 48105.
authors.forename: Jingqiu
authors.initials: J
authors.lastname: Cui
authors.affiliation: Division of Metabolism, Endocrinology, and Diabetes, University of Michigan Medical School, Ann Arbor, Michigan 48105,; Beijing Institute of Pharmacology and Toxicology, Beijing 100850, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Chen
authors.affiliation: Division of Metabolism, Endocrinology, and Diabetes, University of Michigan Medical School, Ann Arbor, Michigan 48105.
authors.forename: Jinhong
authors.initials: J
authors.lastname: Sun
authors.affiliation: Division of Metabolism, Endocrinology, and Diabetes, University of Michigan Medical School, Ann Arbor, Michigan 48105.
authors.forename: Huan
authors.initials: H
authors.lastname: Guo
authors.affiliation: Division of Metabolism, Endocrinology, and Diabetes, University of Michigan Medical School, Ann Arbor, Michigan 48105.
authors.forename: Rachel
authors.initials: R
authors.lastname: Madley
authors.affiliation: Division of Metabolism, Endocrinology, and Diabetes, University of Michigan Medical School, Ann Arbor, Michigan 48105.
authors.forename: Yi
authors.initials: Y
authors.lastname: Xiong
authors.affiliation: Division of Metabolism, Endocrinology, and Diabetes, University of Michigan Medical School, Ann Arbor, Michigan 48105.
authors.forename: Xingyi
authors.initials: X
authors.lastname: Pan
authors.affiliation: Division of Gastroenterology, Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, Michigan 48109.
authors.forename: Hongliang
authors.initials: H
authors.lastname: Wang
authors.affiliation: Division of Gastroenterology, Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, Michigan 48109.
authors.forename: Andrew W
authors.initials: AW
authors.lastname: Tai
authors.affiliation: Department of Biochemistry, Case Western Reserve University, Cleveland, Ohio 44106. Electronic address: michael.weiss@case.edu.
authors.forename: Michael A
authors.initials: MA
authors.lastname: Weiss
authors.affiliation: Division of Metabolism, Endocrinology, and Diabetes, University of Michigan Medical School, Ann Arbor, Michigan 48105.
authors.forename: Peter
authors.initials: P
authors.lastname: Arvan
authors.affiliation: Department of Endocrinology and Metabolism, Tianjin Medical University General Hospital, Tianjin 300052, China,; Division of Metabolism, Endocrinology, and Diabetes, University of Michigan Medical School, Ann Arbor, Michigan 48105.
authors.forename: Ming
authors.initials: M
authors.lastname: Liu
mesh_terms: D004721:Endoplasmic Reticulum / Q000201:enzymology*; D057809:HEK293 Cells; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism*; D011498:Protein Precursors / Q000378:metabolism*; D059748:Proteolysis; D012697:Serine Endopeptidases / Q000378:metabolism*
chemical_list: D008565:Membrane Proteins; D011498:Protein Precursors; D012697:Serine Endopeptidases; C026515:type I signal peptidase
keywords: enzyme inhibitor; mutant; protein processing; protein synthesis; protein trafficking; secretory protein; signal peptidase; viral polypeptide