Eukaryotic initiation factor 6 modulates myofibroblast differentiation at transforming growth factor-β1 transcription level via H2A.Z occupancy and Sp1 recruitment

Abstract

Eukaryotic initiation factor 6 (eIF6) is a pivotal regulator of ribosomal function, participating in translational control. Previously our data suggested that eIF6 acts as a key binding protein of P311 (a hypertrophic scar-related protein; also known as NREP). However, a comprehensive investigation of its functional role and the underlying mechanisms in modulation of myofibroblast (a key effector of hypertrophic scar formation) differentiation remains unclear. Here, we identified that eIF6 is a novel regulator of transforming growth factor-β1 (TGF-β1) expression at transcription level, which plays a key role in myofibroblast differentiation. Mechanistically, this effect is associated with eIF6 altering the occupancy of the TGF-β1 promoter by H2A.Z (Swiss-Prot P0C0S6) and Sp1. Accordingly, modulation of eIF6 expression in myofibroblasts significantly affects their differentiation via the TGF-β/Smad signaling pathway, which was verified in vivo by the observation that heterozygote eIF6 +/− mice exhibited enhanced TGF-β1 production coupled with increased α-smooth muscle actin (α-SMA) + myofibroblasts after skin injury. Overall, our data reveal a novel transcriptional regulatory mechanism of eIF6 that acts on facilitating Sp1 recruitment to TGF-β1 promoter via H2A.Z depletion and thus results in increased TGF-β1 transcription, which contributes to myofibroblast differentiation.

Involvement of eIF6 in external mechanical stretch–mediated murine dermal fibroblast function via TGF-β1 pathway

“…We recently reported that eukaryotic initiation factor 6 (eIF6), which is involved in ribosomal biogenesis and translational control by regulating the binding of the 40S and 60S ribosomal subunits, regulates fibrosis via the TGF-β1 pathway in animal models16, human hypertrophic scarring15, and chronic kidney disease27. eIF6 is also involved in regulating MMP-2/TIMP-2 ratios to balance the degradation and deposition of the extracellular matrix28.…”

“…Because TGF-β pathway activity is intimately involved in fibrosis and collagen deposition, and eIF6 inversely modulates the expression of TGF-β1, mainly at the transcription level16, eIF6 deficiency may result in increased collagen I deposition. This prediction is supported by our results in which the level of TGF-β1 mRNA expression, as well as p-Smad protein expression, is increased in cells containing reduced levels of eIF6, suggesting the activation of TGF-β1signal transduction.…”

“…Pure breed C57BL/6 male mice (wild type, eIF6 +/+ , purchased from the Third Military Medical University, China) and eIF6 heterozygous male mice (eIF6 deficient, eIF6 +/− , purchased from Fondazione Centro San Raffaele del Monte Tabor, Italy) aged 8~10-week weighing 20~22 g were backcrossed to C57BL/6 mice as described in the literature16. Mice were anesthetized with an intraperitoneal injection of 1% (w/v) sodium pentobarbital (0.01 ml/g body weight).…”

“…Our earlier study showed that eIF6 regulates TGF-β1 expression at the transcriptional level by facilitating occupancy of the TGF-β1 promoter by Sp1 rather than H2A.Z16. In vivo studies also show that a heterozygous eIF6 +/− mouse exhibits enhanced TGF-β1 production after skin injury, coupled with increased numbers of α-smooth muscle actin (α-SMA) + myofibroblasts16. We hypothesized that eIF6 plays a role in maintaining fibroblast homeostasis under external mechanical loading through TGF-β pathway regulation.…”

“…eIF6 is also involved in the development of hypertrophic scars, and is therefore a potential target for the control of hypertrophic scarring15. Our earlier study showed that eIF6 regulates TGF-β1 expression at the transcriptional level by facilitating occupancy of the TGF-β1 promoter by Sp1 rather than H2A.Z16. In vivo studies also show that a heterozygous eIF6 +/− mouse exhibits enhanced TGF-β1 production after skin injury, coupled with increased numbers of α-smooth muscle actin (α-SMA) + myofibroblasts16.…”