Sphingosine Kinase 1 (SPHK1) Is Induced by Transforming Growth Factor-β and Mediates TIMP-1 Up-regulation

Abstract

Transforming growth factor-␤ (TGF-␤) signaling plays a pivotal role in extracellular matrix deposition by stimulating collagen production and other extracellular matrix proteins and by inhibiting matrix degradation. The present study was undertaken to define the role of sphingosine kinase (SphK) in TGF-␤ signaling. TGF-␤ markedly up-regulated SphK1 mRNA and protein amounts and caused a prolonged increase in SphK activity in dermal fibroblasts. Concomitantly, TGF-␤ reduced sphingosine-1-phosphate phosphatase activity. Consistent with the changes in enzyme activity, corresponding changes in sphingolipid levels were observed such that sphingosine 1-phosphate (S1P) was increased (ϳ2-fold), whereas sphingosine and ceramide were reduced after 24 h of TGF-␤ treatment. Given the relatively early induction of SphK gene expression in response to TGF-␤, we examined whether SphK1 may be involved in the regulation of TGF-␤-inducible genes that exhibit compatible kinetics, e.g. tissue inhibitor of metalloproteinase-1 (TIMP-1). We demonstrate that decreasing SphK1 expression by small interfering RNA (siRNA) blocked TGF-␤-mediated up-regulation of TIMP-1 protein suggesting that up-regulation of SphK1 contributes to the induction of TIMP-1 in response to TGF-␤. The role of SphK1 as a positive regulator of TIMP-1 gene expression was further corroborated by using ectopically expressed SphK1 in the absence of TGF-␤. Adenovirally expressed SphK1 led to a 2-fold increase of endogenous S1P and to increased TIMP-1 mRNA and protein production. In addition, ectopic SphK1 and TGF-␤ cooperated in TIMP-1 upregulation. Mechanistically, experiments utilizing TIMP-1 promoter constructs demonstrated that the action of SphK1 on the TIMP-1 promoter is through the AP1-response element, consistent with the SphK1-mediated up-regulation of phospho-c-Jun levels, a key component of AP1. Together, these experiments demonstrate that SphK/S1P are important components of the TGF-␤ signaling pathway involved in up-regulation of the TIMP-1 gene. Transforming growth factor-␤ (TGF-␤)1 is a member of a large growth factor family with diverse functions in embryonic and adult tissues (1, 2). TGF-␤ signaling plays a pivotal role in extracellular matrix (ECM) deposition in fibroblasts by stimulating the production of collagens and other ECM proteins and inhibiting matrix degradation (3). The negative effects of TGF-␤ on ECM degradation are mediated via direct inhibition of the principal matrix-degrading proteinase, metalloproteinase-1 (MMP-1) (4), and stimulation of endogenous inhibitors of MMP-1, including TIMP-1 (5). In healthy tissues, a balance between matrix synthesis and degradation precisely controls ECM homeostasis, and shifting this balance can lead to pathological ECM turnover (6). Excessive matrix degradation occurs in rheumatoid arthritis, osteoarthritis, periodontitis, and tumor invasion (6). An imbalance between the tissue inhibitors of matrix metalloproteinases and matrix metalloproteinases is believed to be one of the mechanisms contributing to thes...

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“…It has been observed that TGF-β increases Sphk1 activity and upregulates mRNA and protein levels of Sphk1 in dermal fibroblasts [74]. Thus, it is hypothesized that crosstalk between TGF-β and S1P regulates MMP expression.…”

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