Multiple activation of mitogen-activated protein kinases by purified independent CCN2 modules in vascular endothelial cells and chondrocytes in culture

Satoshi Kubota, H. Kawaki, S. Kondo, G. Yosimichi, M. Minato, Takashi Nishida, H. Hanagata, A. Miyauchi, Masaharu Takigawa

Research output: Contribution to journalArticle

18 Citations (Scopus)

Abstract

CCN2 consists of 4 distinct modules that are conserved among various CCN family protein members. From the N-terminus, insulin-like growth factor binding protein (IGFBP), von Willebrand factor type C repeat (VWC), thrombospondin type 1 repeat (TSP1) and C-terminal cysteine-knot (CT) modules are all aligned tandem therein. The multiple functionality of CCN2 is thought to be enabled by the differential use of these modules when interacting with other molecules. In this study, we independently prepared all 4 purified module proteins of human CCN2, utilizing a secretory production system with Brevibacillus choshinensis and thus evaluated the cell biological effects of such single modules. In human umbilical vascular endothelial cells (HUVECs), VWC, TSP and CT modules, as well as a full-length CCN2, were capable of efficiently activating the ERK signal transduction cascade, whereas IGFBP was not. In contrast, the IGFBP module was found to prominently activate JNK in human chondrocytic HCS-2/8 cells, while the others showed similar effects at lower levels. In addition, ERK1/2 was modestly, but significantly activated by IGFBP and VWC in those cells. No single module, but a mixture of the 4 modules provoked a significant activation of p38 MAPK in HCS-2/8 cells, which was activated by the full-length CCN2. Therefore, the signals emitted by CCN2 can be highly differential, depending upon the cell types, which are thus enabled by the tetramodular structure. Furthermore, the cell biological effects of each module on these cells were also evaluated to clarify the relationship among the modules, the signaling pathways and biological outcomes. Our present results not only demonstrate that single CCN2 modules were potent activators of the intracellular signaling cascade to yield a biological response per se, while also providing new insight into the module-wise structural and functional relationship of a prototypic CCN family member, CCN2.

Original languageEnglish
Pages (from-to)1973-1981
Number of pages9
JournalBiochimie
Volume88
Issue number12
DOIs
Publication statusPublished - Dec 2006

Fingerprint

Insulin-Like Growth Factor Binding Proteins
Endothelial cells
Chondrocytes
Mitogen-Activated Protein Kinases
Cell culture
Endothelial Cells
Chemical activation
CCN Intercellular Signaling Proteins
Cysteine
Thrombospondin 1
Signal transduction
von Willebrand Factor
p38 Mitogen-Activated Protein Kinases
Brevibacillus
Umbilicus
Molecules
Signal Transduction

Keywords

  • Angiogenesis
  • CCN family
  • CCN2
  • Chondrocyte
  • Connective tissue growth factor
  • Endothelial cell

ASJC Scopus subject areas

  • Biochemistry

Cite this

Multiple activation of mitogen-activated protein kinases by purified independent CCN2 modules in vascular endothelial cells and chondrocytes in culture. / Kubota, Satoshi; Kawaki, H.; Kondo, S.; Yosimichi, G.; Minato, M.; Nishida, Takashi; Hanagata, H.; Miyauchi, A.; Takigawa, Masaharu.

In: Biochimie, Vol. 88, No. 12, 12.2006, p. 1973-1981.

Research output: Contribution to journalArticle

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AU - Minato, M.

AU - Nishida, Takashi

AU - Hanagata, H.

AU - Miyauchi, A.

AU - Takigawa, Masaharu

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AB - CCN2 consists of 4 distinct modules that are conserved among various CCN family protein members. From the N-terminus, insulin-like growth factor binding protein (IGFBP), von Willebrand factor type C repeat (VWC), thrombospondin type 1 repeat (TSP1) and C-terminal cysteine-knot (CT) modules are all aligned tandem therein. The multiple functionality of CCN2 is thought to be enabled by the differential use of these modules when interacting with other molecules. In this study, we independently prepared all 4 purified module proteins of human CCN2, utilizing a secretory production system with Brevibacillus choshinensis and thus evaluated the cell biological effects of such single modules. In human umbilical vascular endothelial cells (HUVECs), VWC, TSP and CT modules, as well as a full-length CCN2, were capable of efficiently activating the ERK signal transduction cascade, whereas IGFBP was not. In contrast, the IGFBP module was found to prominently activate JNK in human chondrocytic HCS-2/8 cells, while the others showed similar effects at lower levels. In addition, ERK1/2 was modestly, but significantly activated by IGFBP and VWC in those cells. No single module, but a mixture of the 4 modules provoked a significant activation of p38 MAPK in HCS-2/8 cells, which was activated by the full-length CCN2. Therefore, the signals emitted by CCN2 can be highly differential, depending upon the cell types, which are thus enabled by the tetramodular structure. Furthermore, the cell biological effects of each module on these cells were also evaluated to clarify the relationship among the modules, the signaling pathways and biological outcomes. Our present results not only demonstrate that single CCN2 modules were potent activators of the intracellular signaling cascade to yield a biological response per se, while also providing new insight into the module-wise structural and functional relationship of a prototypic CCN family member, CCN2.

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