title: Rescue and Stabilization of Acetylcholinesterase in Skeletal Muscle by N-terminal Peptides Derived from the Noncatalytic Subunits.
abstract: The vast majority of newly synthesized acetylcholinesterase (AChE) molecules do not assemble into catalytically active oligomeric forms and are rapidly degraded intracellularly by the endoplasmic reticulum-associated protein degradation pathway. We have previously shown that AChE in skeletal muscle is regulated in part post-translationally by the availability of the noncatalytic subunit collagen Q, and others have shown that expression of a 17-amino acid N-terminal proline-rich attachment domain of collagen Q is sufficient to promote AChE tetramerization in cells producing AChE. In this study we show that muscle cells, or cell lines expressing AChE catalytic subunits, incubated with synthetic proline-rich attachment domain peptides containing the endoplasmic reticulum retrieval sequence KDEL take up and retrogradely transport them to the endoplasmic reticulum network where they induce assembly of AChE tetramers. The peptides act to enhance AChE folding thereby rescuing them from reticulum degradation. This enhanced folding efficiency occurs in the presence of inhibitors of protein synthesis and in turn increases total cell-associated AChE activity and active tetramer secretion. Pulse-chase studies of isotopically labeled AChE molecules show that the enzyme is rescued from intracellular degradation. These studies provide a mechanistic explanation for the large scale intracellular degradation of AChE previously observed and indicate that simple peptides alone can increase the production and secretion of this critical synaptic enzyme in muscle tissue.
authors.affiliation: Department of Cell Biology, University of Miami Miller School of Medicine, Miami, Florida 33136.
authors.forename: Carlos A
authors.initials: CA
authors.lastname: Ruiz
authors.affiliation: Department of Cell Biology, University of Miami Miller School of Medicine, Miami, Florida 33136.
authors.forename: Susana G
authors.initials: SG
authors.lastname: Rossi
authors.affiliation: Department of Cell Biology, University of Miami Miller School of Medicine, Miami, Florida 33136; Department of Neuroscience Program, University of Miami Miller School of Medicine, Miami, Florida 33136. Electronic address: rrotundo@miami.edu.
authors.forename: Richard L
authors.initials: RL
authors.lastname: Rotundo
mesh_terms: D000110:Acetylcholinesterase / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D030161:Avian Proteins / Q000235:genetics / Q000378:metabolism*; D019556:COS Cells; D020134:Catalytic Domain / Q000235:genetics*; D002522:Chlorocebus aethiops; D004625:Embryo, Nonmammalian; D058851:GPI-Linked Proteins / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D057809:HEK293 Cells; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D018482:Muscle, Skeletal / Q000166:cytology / Q000187:drug effects / Q000201:enzymology; D032446:Myoblasts / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology; D010455:Peptides / Q000138:chemical synthesis / Q000494:pharmacology*; D061251:Primary Cell Culture; D055503:Protein Multimerization; D055550:Protein Stability / Q000187:drug effects; D011784:Quail; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment
chemical_list: D030161:Avian Proteins; D058851:GPI-Linked Proteins; D010455:Peptides; D011994:Recombinant Proteins; C548244:ACHE protein, human; D000110:Acetylcholinesterase
keywords: ER quality control; enzyme processing; inactive precursors; membrane enzyme; protein folding; protein import; protein subunits