A New Class of Mutants Deficient in Dodecamerization of Aminopeptidase 1 and Vacuolar Transport

A New Class of Mutants Deficient in Dodecamerization of Aminopeptidase 1 and Vacuolar Transport

Vacuolar aminopeptidase 1 is transported to the vacuole by cytoplasmic double-membrane vesicles, the nonclassic Cvt pathway. The cytosolic protein dodecamerizes and is enclosed in a double-membrane vesicle, which is transported to and fuses with the vacuole releasing a single-membrane autophagic bod...

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Bibliographic Details
Published in:The Journal of biological chemistry Vol. 276; no. 15; pp. 11606 - 11614
Main Authors: Andrei-Selmer, Cornelia, Knüppel, Angelika, Satyanarayana, Chitkala, Heese, Claudia, Schu, Peter V.
Format: Journal Article
Language:English
Published: United States Elsevier Inc 13-04-2001
American Society for Biochemistry and Molecular Biology
Subjects:
Aminopeptidases - chemistry
Aminopeptidases - genetics
Aminopeptidases - metabolism
autophagy
Biological Transport
Biopolymers
Mutation
Protein Conformation
vacuolar aminopeptidase 1
Vacuoles - enzymology
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Summary:Vacuolar aminopeptidase 1 is transported to the vacuole by cytoplasmic double-membrane vesicles, the nonclassic Cvt pathway. The cytosolic protein dodecamerizes and is enclosed in a double-membrane vesicle, which is transported to and fuses with the vacuole releasing a single-membrane autophagic body into the vacuolar lumen. This is degraded and the precursor sequence of aminopeptidase 1 is removed. This pathway resembles autophagy, and most proteins identified to function in the Cvt pathway are also required for autophagy and vice versa. The cytosolic precursor protein and the matured vacuolar protein form a homododecameric complex, and only this complex has enzymatic activity. We developed a new genetic screen to isolate mutants in the biogenesis of vacuolar aminopeptidase 1 based on its enzymatic activity. The sensitivity of this assay made it possible for us to search for mutants under conditions where autophagy is down-regulated, and we describe two new mutants defective in the biogenesis pathway of vacuolar aminopeptidase 1. Mutants are defective in dodecamerization of pApe1p and in Cvt vesicle formation. Complex assembly and transport vesicle formation appear to be linked processes. This mechanism can control the potentially harmful cytoplasmic proteolytic activity and could be the driving force for this nonclassic mechanism of vacuolar enzyme transport.
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ISSN:0021-9258
1083-351X
DOI:10.1074/jbc.M003846200