Identification of autophagosome-associated proteins and regulators by quantitative proteomic analysis and genetic screens

Jörn Dengjel; Maria Høyer-Hansen; Maria O Nielsen; Tobias Eisenberg; Lea Mørch Harder; Søren Schandorff; Thomas Farkas; Thomas Kirkegaard; Andrea C Becker; Sabrina Schroeder; Katja Vanselow; Emma Lundberg; Mogens Møller Nielsen; Anders Riis Kristensen; Vyacheslav Akimov; Jakob Bunkenborg; Frank Madeo; Marja Jaattela; Jens S Andersen

doi:10.1074/mcp.M111.014035

Identification of autophagosome-associated proteins and regulators by quantitative proteomic analysis and genetic screens

Jörn Dengjel, Maria Høyer-Hansen, Maria O Nielsen, Tobias Eisenberg, Lea Mørch Harder, Søren Schandorff, Thomas Farkas, Thomas Kirkegaard, Andrea C Becker, Sabrina Schroeder, Katja Vanselow, Emma Lundberg, Mogens Møller Nielsen, Anders Riis Kristensen, Vyacheslav Akimov, Jakob Bunkenborg, Frank Madeo, Marja Jaattela, Jens S Andersen

Morphogenesis and Differentiation Program

86 Citations (Scopus)

Abstract

Autophagy is one of the major intracellular catabolic pathways, but little is known about the composition of autophagosomes. To study the associated proteins, we isolated autophagosomes from human breast cancer cells using two different biochemical methods and three stimulus types: amino acid deprivation or rapamycin or concanamycin A treatment. The autophagosome-associated proteins were dependent on stimulus, but a core set of proteins was stimulus-independent. Remarkably, proteasomal proteins were abundant among the stimulus-independent common autophagosome-associated proteins, and the activation of autophagy significantly decreased the cellular proteasome level and activity supporting interplay between the two degradation pathways. A screen of yeast strains defective in the orthologs of the human genes encoding for a common set of autophagosome-associated proteins revealed several regulators of autophagy, including subunits of the retromer complex. The combined spatiotemporal proteomic and genetic data sets presented here provide a basis for further characterization of autophagosome biogenesis and cargo selection.

Original language	English
Journal	Molecular & Cellular Proteomics
Volume	11
Issue number	3
Pages (from-to)	M111.014035
ISSN	1535-9484
DOIs	https://doi.org/10.1074/mcp.M111.014035
Publication status	Published - Mar 2012

Keywords

Amino Acids
Antibodies, Monoclonal
Antiviral Agents
Autophagy
Breast Neoplasms
Electrophoresis, Polyacrylamide Gel
Female
Genetic Testing
Green Fluorescent Proteins
Humans
Immunoprecipitation
Immunosuppressive Agents
Isotope Labeling
Lysosomes
Macrolides
Phagosomes
Proteins
Proteomics
Saccharomyces cerevisiae
Sirolimus
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Starvation
Tumor Cells, Cultured

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

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10.1074/mcp.M111.014035

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Dengjel, J., Høyer-Hansen, M., Nielsen, M. O., Eisenberg, T., Harder, L. M., Schandorff, S., Farkas, T., Kirkegaard, T., Becker, A. C., Schroeder, S., Vanselow, K., Lundberg, E., Nielsen, M. M., Kristensen, A. R., Akimov, V., Bunkenborg, J., Madeo, F., Jaattela, M., & Andersen, J. S. (2012). Identification of autophagosome-associated proteins and regulators by quantitative proteomic analysis and genetic screens. Molecular & Cellular Proteomics, 11(3), M111.014035. https://doi.org/10.1074/mcp.M111.014035

Dengjel, J, Høyer-Hansen, M, Nielsen, MO, Eisenberg, T, Harder, LM, Schandorff, S, Farkas, T, Kirkegaard, T, Becker, AC, Schroeder, S, Vanselow, K, Lundberg, E, Nielsen, MM, Kristensen, AR, Akimov, V, Bunkenborg, J, Madeo, F, Jaattela, M & Andersen, JS 2012, 'Identification of autophagosome-associated proteins and regulators by quantitative proteomic analysis and genetic screens', Molecular & Cellular Proteomics, vol. 11, no. 3, pp. M111.014035. https://doi.org/10.1074/mcp.M111.014035

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AU - Harder, Lea Mørch

AU - Schandorff, Søren

AU - Farkas, Thomas

AU - Kirkegaard, Thomas

AU - Becker, Andrea C

AU - Schroeder, Sabrina

AU - Vanselow, Katja

AU - Lundberg, Emma

AU - Nielsen, Mogens Møller

AU - Kristensen, Anders Riis

AU - Akimov, Vyacheslav

AU - Bunkenborg, Jakob

AU - Madeo, Frank

AU - Jaattela, Marja

AU - Andersen, Jens S

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N2 - Autophagy is one of the major intracellular catabolic pathways, but little is known about the composition of autophagosomes. To study the associated proteins, we isolated autophagosomes from human breast cancer cells using two different biochemical methods and three stimulus types: amino acid deprivation or rapamycin or concanamycin A treatment. The autophagosome-associated proteins were dependent on stimulus, but a core set of proteins was stimulus-independent. Remarkably, proteasomal proteins were abundant among the stimulus-independent common autophagosome-associated proteins, and the activation of autophagy significantly decreased the cellular proteasome level and activity supporting interplay between the two degradation pathways. A screen of yeast strains defective in the orthologs of the human genes encoding for a common set of autophagosome-associated proteins revealed several regulators of autophagy, including subunits of the retromer complex. The combined spatiotemporal proteomic and genetic data sets presented here provide a basis for further characterization of autophagosome biogenesis and cargo selection.

AB - Autophagy is one of the major intracellular catabolic pathways, but little is known about the composition of autophagosomes. To study the associated proteins, we isolated autophagosomes from human breast cancer cells using two different biochemical methods and three stimulus types: amino acid deprivation or rapamycin or concanamycin A treatment. The autophagosome-associated proteins were dependent on stimulus, but a core set of proteins was stimulus-independent. Remarkably, proteasomal proteins were abundant among the stimulus-independent common autophagosome-associated proteins, and the activation of autophagy significantly decreased the cellular proteasome level and activity supporting interplay between the two degradation pathways. A screen of yeast strains defective in the orthologs of the human genes encoding for a common set of autophagosome-associated proteins revealed several regulators of autophagy, including subunits of the retromer complex. The combined spatiotemporal proteomic and genetic data sets presented here provide a basis for further characterization of autophagosome biogenesis and cargo selection.

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KW - Genetic Testing

KW - Green Fluorescent Proteins

KW - Humans

KW - Immunoprecipitation

KW - Immunosuppressive Agents

KW - Isotope Labeling

KW - Lysosomes

KW - Macrolides

KW - Phagosomes

KW - Proteins

KW - Proteomics

KW - Saccharomyces cerevisiae

KW - Sirolimus

KW - Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization

KW - Starvation

KW - Tumor Cells, Cultured

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DO - 10.1074/mcp.M111.014035

M3 - Journal article

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SN - 1535-9476

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SP - M111.014035

JO - Molecular and Cellular Proteomics

JF - Molecular and Cellular Proteomics

IS - 3

ER -

Identification of autophagosome-associated proteins and regulators by quantitative proteomic analysis and genetic screens

Abstract

Keywords

UN SDGs

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