Matrix metalloproteinase-1 induces cleavage of exogenous alphab-crystallin transduced by a cell-penetrating peptide

Seung Won Yang, Seung Min Lee, Eun Young Choi, Kyung Hye Lee, Soo Hyuk Kim, Min-Jeong Shin, Ye Sun Han, Seok Min Kang, Ji Hyung Chung

Research output: Contribution to journalArticle

Abstract

Cell-penetrating peptides (CPPs), including TAT-CPP, have been used to deliver exogenous proteins into living cells. Although a number of proteins fused to TAT-CPP can be delivered into various cells, little is known about the proteolytic cleavage of TAT-fusion proteins in cells. In this study, we demonstrate that a small heat shock protein (sHSP), alphaB-crystallin (αB-crystallin), delivered by TAT-CPP is susceptible to proteolytic cleavage by matrix metalloproteinase-1 (MMP-1) in cardiac myoblast H9c2 cells. Recombinant TAT-αB-crystallin was efficiently transduced into H9c2 cells. For a few hours following protein transduction, generation of a 14-kDa fragment, a cleavage band of TAT-αB-crystallin, increased in a time-dependent manner. This fragment was observed only in detergent-insoluble fractions. Interestingly, treatment with MMP inhibitors blocked the cleavage of TAT-αB-crystallin. In test tubes, recombinant MMP-1 processed TAT-αB-crystallin to generate the major cleavage fragment 14-kDa, as observed in the cells treated with TAT-αB-crystallin. The N-terminal sequences of the 14-kDa fragment were identified as Leu-Arg-Ala-Pro-Ser-Trp-Phe, indicating that this fragment is generated by cleavage at Phe54-Leu55 of αB-crystallin. The MMP-1-selective inhibitor abolished the production of 14-kDa fragments in cells. In addition, the cleaved fragment of TAT-αB-crystallin was significantly reduced in cells transfected with MMP-1 siRNA. Moreover, the enzymatic activity of MMP-1 was markedly increased in TAT-αB-crystallin-treated cells. TAT-αB-crystallin has a cytoprotective effect on H9c2 cells under hypoxic insult, moreover, MMP-1-selective inhibitor treatment led to even increased cell viability. These results suggest that MMP-1 is responsible for proteolytic cleavage of TAT-αB-crystallin during its intracellular transduction in H9c2 cells.

Original languageEnglish
Pages (from-to)2454-2462
Number of pages9
JournalJournal of Cellular Biochemistry
Volume112
Issue number9
DOIs
Publication statusPublished - 2011 Sep 1

Fingerprint

Cell-Penetrating Peptides
Matrix Metalloproteinase 1
Crystallins
Cells
Cardiac Myoblasts
Proteins
Small Heat-Shock Proteins
Matrix Metalloproteinase Inhibitors
Detergents
Small Interfering RNA

Keywords

  • Alphab-crystallin
  • Cell-penetrating peptide
  • MMP-1
  • Proteolysis

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

Cite this

Matrix metalloproteinase-1 induces cleavage of exogenous alphab-crystallin transduced by a cell-penetrating peptide. / Yang, Seung Won; Lee, Seung Min; Choi, Eun Young; Lee, Kyung Hye; Kim, Soo Hyuk; Shin, Min-Jeong; Han, Ye Sun; Kang, Seok Min; Chung, Ji Hyung.

In: Journal of Cellular Biochemistry, Vol. 112, No. 9, 01.09.2011, p. 2454-2462.

Research output: Contribution to journalArticle

Yang, Seung Won ; Lee, Seung Min ; Choi, Eun Young ; Lee, Kyung Hye ; Kim, Soo Hyuk ; Shin, Min-Jeong ; Han, Ye Sun ; Kang, Seok Min ; Chung, Ji Hyung. / Matrix metalloproteinase-1 induces cleavage of exogenous alphab-crystallin transduced by a cell-penetrating peptide. In: Journal of Cellular Biochemistry. 2011 ; Vol. 112, No. 9. pp. 2454-2462.
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AU - Lee, Kyung Hye

AU - Kim, Soo Hyuk

AU - Shin, Min-Jeong

AU - Han, Ye Sun

AU - Kang, Seok Min

AU - Chung, Ji Hyung

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