Amino Acids

, Volume 41, Issue 2, pp 351–361 | Cite as

Quantitative proteome analysis of the 20S proteasome of apoptotic Jurkat T cells

  • Frank Schmidt
  • Burkhardt Dahlmann
  • Hanne K. Hustoft
  • Christian J. Koehler
  • Margarita Strozynski
  • Alexander Kloß
  • Ursula Zimny-Arndt
  • Peter R. Jungblut
  • Bernd Thiede
Original Article

Abstract

Regulated proteolysis plays important roles in cell biology and pathological conditions. A crosstalk exists between apoptosis and the ubiquitin–proteasome system, two pathways responsible for regulated proteolysis executed by different proteases. To investigate whether the apoptotic process also affects the 20S proteasome, we performed three independent SILAC-based quantitative proteome approaches: 1-DE/MALDI-MS, small 2-DE/MALDI-MS and large 2-DE/nano-LC–ESI–MS. Taking the results of all experiments together, no quantitative changes were observed for the α- and β-subunits of the 20S proteasome except for subunit α7. This protein was identified in two protein spots with a down-regulation of the more acidic protein species (α7a) and up-regulation of the more basic protein species (α7b) during apoptosis. The difference in these two α7 protein species could be attributed to oxidation of cysteine-41 to cysteine sulfonic acid and phosphorylation at serine-250 near the C terminus in α7a, whereas these modifications were missing in α7b. These results pointed to the biological significance of posttranslational modifications of proteasome subunit α7 after induction of apoptosis.

Keywords

Apoptosis 5-Fluorouracil 20S proteasome Phosphorylation SILAC 

Abbreviations

19S-Reg

19S regulator complex

5-FU

5-Fluorouracil

H/L

Heavy to light

PARP1

Poly (ADP-ribose) polymerase-1

SILAC

Stable isotope labeling with amino acids in cell culture

Supplementary material

726_2010_575_MOESM1_ESM.xls (42 kb)
Supplementary material 1 (XLS 42 kb)

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Copyright information

© Springer-Verlag 2010

Authors and Affiliations

  • Frank Schmidt
    • 1
    • 2
  • Burkhardt Dahlmann
    • 3
  • Hanne K. Hustoft
    • 1
  • Christian J. Koehler
    • 1
  • Margarita Strozynski
    • 1
  • Alexander Kloß
    • 3
  • Ursula Zimny-Arndt
    • 4
  • Peter R. Jungblut
    • 4
  • Bernd Thiede
    • 1
  1. 1.The Biotechnology Centre of OsloUniversity of OsloOsloNorway
  2. 2.Interfaculty Institute for Genetics and Functional GenomicsUniversity of GreifswaldGreifswaldGermany
  3. 3.Institute of BiochemistryCharité-UniversitätsmedizinBerlinGermany
  4. 4.Max Planck Institute for Infection Biology, Core Facility Protein AnalysisBerlinGermany

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