Induction of Pro-Fibrotic CLIC4 in Dermal Fibroblasts by TGF-β/Wnt3a Is Mediated by GLI2 Upregulation

Abstract

Chloride intracellular channel 4 (CLIC4) is a recently discovered driver of fibroblast activation in Scleroderma (SSc) and cancer-associated fibroblasts (CAF). CLIC4 expression and activity are regulated by TGF-β signalling through the SMAD3 transcription factor. In view of the aberrant activation of canonical Wnt-3a and Hedgehog (Hh) signalling in fibrosis, we investigated their role in CLIC4 upregulation. Here, we show that TGF-β/SMAD3 co-operates with Wnt3a/β-catenin and Smoothened/GLI signalling to drive CLIC4 expression in normal dermal fibroblasts, and that the inhibition of β-catenin and GLI expression or activity abolishes TGF-β/SMAD3-dependent CLIC4 induction. We further show that the expression of the pro-fibrotic marker α-smooth muscle actin strongly correlates with CLIC4 expression in dermal fibroblasts. Further investigations revealed that the inhibition of CLIC4 reverses morphogen-dependent fibroblast activation. Our data highlights that CLIC4 is a common downstream target of TGF-β, Hh, and Wnt-3a through signalling crosstalk and we propose a potential therapeutic avenue using CLIC4 inhibitors

The Beta secretase BACE1 drives fibroblasts activation in Systemic Sclerosis through the APP/β-catenin/Notch signalling axis

“…As BACE1 KO has no effect on α-SMA expression on healthy fibroblasts, we therefore assessed the effects of BACE1 inhibition and KO on healthy fibroblasts stimulated with pro-fibrotic stimuli such as the morphogens. Morphogen signalling (TGF-β, Wnt-3a and Hedgehog) is deregulated in SSc fibroblasts and play an important role in SSc disease progression (2, 3, 4). The morphogens enhance the expression of several pro-fibrotic genes in fibroblasts, including α-SMA (3).…”

“…Morphogen signalling (TGF-β, Wnt-3a and Hedgehog) is deregulated in SSc fibroblasts and play an important role in SSc disease progression (2, 3, 4). The morphogens enhance the expression of several pro-fibrotic genes in fibroblasts, including α-SMA (3). To determine if the morphogens play a role in regulating BACE1 expression in dermal fibroblasts, we stimulated healthy dermal fibroblasts with TGF-β, Wnt-3a or SAG (smoothened agonist that activates the hedgehog pathway) and assessed BACE1 protein levels.…”

“…We have previously shown co-operation between the Wnt3a and TGF-β signalling pathways in dermal fibroblasts. β-catenin inhibitors blocked the ability of TGF-β to induce signalling (3).…”

“…The fibrosis is driven by activated fibroblasts (myofibroblasts) characterised by increased expression of α-SMA and secretion of extracellular matrix proteins. Several pathways have been implicated in the activation of fibroblasts in SSc, including the TGF-β, Wnt/β-catenin and sonic hedgehog (SHh) pathways (1,2, 3, 4).…”

Role of the hedgehog signaling pathway in rheumatic diseases: An overview

“…It is associated with increased activation of TGF-β ( 99 ). Investigations revealed the expression of CLIC4 in normal dermal fibroblasts was driven by TGF-β/SMAD3 together with Wnt3a/β-catenin and Smo/Gli signaling, and inhibition of Gli reduced CLIC4 expression ( 86 ). The hedgehog acyltransferase (HHAT) catalyses the attachment of palmitate onto Shh ( 100 ).…”