Thymosin ß4 Enhances MMP-1 Promoter Activity by Involving C-Jun Interaction With Histone Acetyltransferase (HAT) and Swi/Snf Chromatin Remodeling Factors

Abstract

Purpose:: The interaction of transcription factors with chromatin modifier and remodeling factors determine the transcriptional levels of eukaryotic genes. Previously, we demonstrated that Tß₄ enhances c-Jun and MMP-1 transcription in corneal epithelial cells and promotes epithelial cell migration and wound healing. The purpose of this study is to investigate whether Tß₄ modulation of MMP-1 promoter function involves communications between c-Jun, CBP/p300 and ATPase subunits of the SWI/SNF complex.

Methods:: Immortalized human corneal epithelial cells (HCET) were cultured in KGM and transiently transfected with the following plasmids: 1) luciferase reporter with 500bp of the 5’ promoter region of the MMP-1 gene; 2) expression vectors encoding the transcription factors c-Jun, chromatin remodeling factors (Brg-1 and BRM), HATs (CBP and p300), and their inhibitors (E1A and Twist). Transfected cells were treated either with basic medium (KBM) alone; KBM plus TSA, a histone deacetylase (HDAC) inhibitor, with and without Tß₄ for 24 hours. Collected cell lysates were used to assay luciferase activity.

Results:: MMP-1 promoter activity only marginally increased in cells over-expressing CBP, p300, Brg-1, BRM, or those treated with TSA or Tß₄. Introducing c-Jun into HCET stimulated MMP-1 promoter activity by 25-35 fold over controls. Co-transfection of Jun with p300 or BRM further enhanced MMP-1 promoter activity by 50 and 100 fold, respectively. These synergistic effects were markedly reduced by replacing p300 and BRM with enzyme-activity deficient mutants, while over-expression of the HATs inhibitors E1A and Twist did not influence these stimulatory effects. MMP-1 promoter activity was stimulated as high as 175-300 fold by Tß₄ in the presence of over-expression of c-Jun-p300 or c-Jun-BRM.

Conclusions:: c-Jun plays a critical role in recruiting chromatin modifier and remodeling factors into the MMP-1 promoter region during transcription. Our data suggests that Tß₄ may target the c-Jun-related transcriptional complex thereby modulating MMP-1 promoter activation as well as chromatin remodeling. Since Tß₄ promotes corneal epithelial cell migration and wound healing, these findings suggest a possible mechanism by which Tß₄ affects the transcription of the pro-migratory molecule, MMP-1.

Keywords: cornea: epithelium • gene/expression • transcription