The role of neuron-restrictive silencer factor in modulating the Sp-1 mediated transactivation of human secretin receptor gene

Abstract

To understand further the cell-specific and developmental-specific expressions of the human secretin receptor (hSR), in the present study, we sought to investigate the functions of a putative neuron restriction silencer element NRSE (− 83 to − 67, relative to ATG) motif located downstream of the hSR core promoter. neuron restriction silencer element (NRSE, also known as repressor element-1 RE-1). Promoter assays coupled with scanning mutation analyses indicated that this NRSE is functional, while in vitro gel mobility shift studies and in vivo ChIP assays suggested the binding of NRSF with this motif. The silencing function of NRSF was further confirmed by over-expression and shRNA knock-down of NRSF. These studied showed an inverse relationship between the expression levels of NRSF and hSR in the cells. We recently demonstrated the hSR gene is controlled by two GC-boxes which are regulated by an in vivo ratio of Sp1 to Sp3, in the present study, we sought to investigate the interactions of NRSF and Sp proteins to regulate hSR gene. By co-immunoprecipitation assays, we found that NRSF could co-precipitate with Sp1 as well as Sp3 in PANC-1 cells. Interestingly, co-expressions of these factors showed that NRSF could suppress Sp1-mediated, but not Sp3-mediated, transactivation of hSR. Taken together, we propose here that the silencing effects of NRSF on hSR gene expression are mediated via the changing of the stochastic ratio of Sp1 to Sp3. The finding not only provided information regarding the cell-type specific expression pattern of hSR. It may also apply on other GC-boxes/NRSF regulating genes, including the secretin receptor gene family.