Abstract

Identification and characterization of TGF�²-dependent and -independent cis-regulatory modules in the C4ST-1/CHST11 locus

Chondroitin-4-sulfotransferase-1(C4ST-1)/carbohydrate sul­fotransferase 11 (CHST11) is a Golgi-bound enzyme involved in the biosyn­thesis of the glycosaminoglycan chondroitin sulfate. The sulfation pattern of chondroitin is tightly regulated during development, injury and disease, with the temporal and spatial expression of chondroitin sulfotransferase genes be­lieved to be a crucial determinant of the fine balance of chondroitin sulfation. We have previously identified mouse C4st-1 as a target gene of ligands of the TGFβ superfamily of growth factors, which could positively regulate C4st-1 expression in a number of cell types. We have also shown that a gene trap loss-of-function mutation in C4st-1 leads to severe skeletal abnormali­ties during mouse embryogenesis. In addition, we described a highly specific temporal and spatial expression pattern of C4st-1 during mouse embryogen­esis. However, the transcriptional regulatory mechanisms that control C4st-1 gene expression remain unexplored. In order to gain knowledge on the transcriptional regulation of C4ST-1, we used a bioinformatical approach to identify conserved putative long-range cis-regulatory modules in a region of 120 kb spanning the 5’ end of the C4ST-1 gene. Luciferase reporter assays in human HEK293T and mouse NmuMG cells identified a functional C4ST-1 promoter, as well as a number of cis-regulatory modules able to positively and negatively regulate C4ST-1 expression. Moreover, we identified TGFβ- responsive regulatory modules that can function in a cell type-specific fash­ion. Taken together, our results identify TGFβ-dependent and -independent cis-regulatory modules of the C4ST-1 gene.

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