Non-synonymous gene polymorphisms in the secretory signal peptide of human TGF-beta1 affect cellular synthesis but not secretion of TGF-beta1.

We have demonstrated that gene polymorphisms within the N-terminal leader sequence of TGF-beta1 contribute to the outcome of hepatic fibrogenesis. In addition, the polymorphism at codon 25 affects TGF-beta1 production in peripheral blood leukocytes. Therefore, it is general assumed that these polymorphisms influence cellular secretion of this cytokine. In the present study, we analysed if this widespread hypothesis is true. We cloned FLAG-tagged CMV-driven human full-length TGF-beta1 expression constructs of the different allelic variations (i.e. 10Leu/25Arg, 10Pro/25Pro and 10Pro/25Arg) and transfected them into the immortal hepatic stellate cell line LX-2 and Chinese Hamster Ovary cells. Surprisingly, the allelic variants carrying a proline either in codon 10 or 25 showed overall reduced expression as assessed by Western blot and quantitative ELISA. We conclude that the allelic variations within the signal sequence influence the expression and not secretion of TGF-beta1. Detailed RNA structure prediction analysis further suggests that the individual variants form different secondary structures.