Biochemical evidence of the interactions of membrane type-1 matrix metalloproteinase (MT1-MMP) with adenine nucleotide translocator (ANT)

Potential implications linking proteolysis with energy metabolism in cancer cells

Ilian A. Radichev, Albert G. Remacle, Nor Eddine Sounni, Sergey A. Shiryaev, Dmitri Rozanov, Wenhong Zhu, Natalya V. Golubkova, Tatiana I. Postnova, Vladislav S. Golubkov, Alex Y. Strongin

Research output: Contribution to journalArticle

9 Citations (Scopus)

Abstract

Invasion-promoting MT1-MMP (membrane type-1 matrix metalloproteinase) is a key element in cell migration processes. To identify the proteins that interact and therefore co-precipitate with this proteinase from cancer cells, we used the proteolytically active WT (wild-type), the catalytically inert E240A and the C-end truncated (tailless; ΔCT) MT1-MMP-FLAG constructs as baits. The identity of the pulled-down proteins was determined by LC-MS/MS (liquid chromatography tandem MS) and then confirmed by Western blotting using specific antibodies. We determined that, in breast carcinoma MCF cells (MCF-7 cells), ANT (adenine nucleotide translocator) efficiently interacted with the WT, E240A and ΔCT constructs. The WT and E240A constructs also interacted with α-tubulin, an essential component of clathrin-mediated endocytosis. In turn, tubulin did not co-precipitate with the ΔCT construct because of the inefficient endocytosis of the latter, thus suggesting a high level of selectivity of our test system. To corroborate these results, we then successfully used the ANT2-FLAG construct as a bait to pull-down MT1-MMP, which was naturally produced by fibrosarcoma HT1080 cells. We determined that the presence of the functionally inert catalytic domain alone was sufficient to cause the proteinase to interact with ANT2, thus indicating that there is a non-proteolytic mode of these interactions. Overall, it is tempting to hypothesize that by interacting with pro-invasive MT1-MMP, ANT plays a yet to be identified role in a coupling mechanism between energy metabolism and pericellular proteolysis in migrating cancer cells.

Original languageEnglish (US)
Pages (from-to)37-47
Number of pages11
JournalBiochemical Journal
Volume420
Issue number1
DOIs
StatePublished - May 15 2009
Externally publishedYes

Fingerprint

Proteolysis
Matrix Metalloproteinase 14
Adenine Nucleotides
Energy Metabolism
Cells
Neoplasms
Tubulin
Endocytosis
Precipitates
Peptide Hydrolases
Clathrin
Matrix Metalloproteinase 1
Fibrosarcoma
Liquid chromatography
MCF-7 Cells
Liquid Chromatography
Cell Movement
Catalytic Domain
Proteins
Western Blotting

Keywords

  • Adenine nucleotide translocator (ANT)
  • Energy metabolism
  • Membrane type-1 matrix metalloproteinase(MT1-MMP)
  • Mitochondrion
  • Pericellular proteolysis
  • Protein-protein interaction

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology
  • Medicine(all)

Cite this

Biochemical evidence of the interactions of membrane type-1 matrix metalloproteinase (MT1-MMP) with adenine nucleotide translocator (ANT) : Potential implications linking proteolysis with energy metabolism in cancer cells. / Radichev, Ilian A.; Remacle, Albert G.; Sounni, Nor Eddine; Shiryaev, Sergey A.; Rozanov, Dmitri; Zhu, Wenhong; Golubkova, Natalya V.; Postnova, Tatiana I.; Golubkov, Vladislav S.; Strongin, Alex Y.

In: Biochemical Journal, Vol. 420, No. 1, 15.05.2009, p. 37-47.

Research output: Contribution to journalArticle

Radichev, Ilian A. ; Remacle, Albert G. ; Sounni, Nor Eddine ; Shiryaev, Sergey A. ; Rozanov, Dmitri ; Zhu, Wenhong ; Golubkova, Natalya V. ; Postnova, Tatiana I. ; Golubkov, Vladislav S. ; Strongin, Alex Y. / Biochemical evidence of the interactions of membrane type-1 matrix metalloproteinase (MT1-MMP) with adenine nucleotide translocator (ANT) : Potential implications linking proteolysis with energy metabolism in cancer cells. In: Biochemical Journal. 2009 ; Vol. 420, No. 1. pp. 37-47.
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