Assembly of acetylcholinesterase tetramers by peptidic motifs from the proline-rich membrane anchor, PRiMA: competition between degradation and secretion pathways of heteromeric complexes.
The membrane-bound form of acetylcholinesterase (AChE) constitutes the major component of this enzyme in the mammalian brain. These molecules are hetero-oligomers, composed of four AChE catalytic subunits of type T (AChE(T)), associated with a transmembrane protein of type 1, called PRiMA (proline-rich membrane anchor). PRiMA consists of a signal peptide, an extracellular domain that contains a proline-rich motif (14 prolines with an intervening leucine, P4LP10), a transmembrane domain, and a cytoplasmic domain. Expression of AChE(T) subunits in transfected COS cells with a truncated PRiMA, without its transmembrane and cytoplasmic domains (P(stp54) mutant), produced secreted heteromeric complexes (T4-P(stp54)), instead of membrane-bound tetramers. In this study, we used a series of deletions and point mutations to analyze the interaction between the extracellular domain of PRiMA and AChE(T) subunits. We confirmed the importance of the polyproline stretches and defined a peptidic motif (RP4LP10RL), which induces the assembly and secretion of a heteromeric complex with four AChE(T) subunits, nearly as efficiently as the entire extracellular domain of PRiMA. It is noteworthy that deletion of the N-terminal segment preceding the prolines had little effect. Interestingly, short PRiMA mutants, truncated within the proline-rich motif, reduced both cellular and secreted AChE activity, suggesting that their interaction with AChE(T) subunits induces their intracellular degradation.
Noureddine
Hiba
H
Laboratoire de Neurobiologie Cellulaire et Moléculaire, CNRS UMR 8544, Ecole Normale Supérieure, 46 Rue d'Ulm, 75005 Paris.
Schmitt
Claudine
C
Liu
Wangqing
W
Garbay
Christiane
C
Massoulié
Jean
J
Bon
Suzanne
S
D000110:Acetylcholinesterase / Q000737:chemistry* / Q000378:metabolism / Q000502:physiology; D020816:Amino Acid Motifs / Q000235:genetics; D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D020134:Catalytic Domain / Q000235:genetics; D002522:Chlorocebus aethiops; D006224:Cricetinae; D005110:Extracellular Space / Q000737:chemistry / Q000235:genetics / Q000502:physiology; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D051379:Mice; D008969:Molecular Sequence Data; D046912:Multiprotein Complexes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D016297:Mutagenesis, Site-Directed; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D010455:Peptides / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D021381:Protein Transport / Q000235:genetics; D051381:Rats; D017384:Sequence Deletion / Q000235:genetics; D015398:Signal Transduction / Q000235:genetics
D008565:Membrane Proteins; D046912:Multiprotein Complexes; D009419:Nerve Tissue Proteins; C508016:PRiMA1 protein, rat; D010455:Peptides; C486718:prima1 protein, mouse; D000110:Acetylcholinesterase