The CCAAT Enhancer-binding Protein (C/EBP)β and Nrf1 Interact to Regulate Dentin Sialophosphoprotein (DSPP) Gene Expression during Odontoblast Differentiation

TLDR

Odontoblast terminal differentiation involves synthesis of type I collagen and noncollagenous proteins, with DSP and DPP—encoded by the DSPP gene—serving as specific markers. The study aims to uncover how the interaction between transcription factors Nrf1 and C/EBPβ regulates tissue‑specific expression of the DSPP gene. Immunoprecipitation, chromatin immunoprecipitation, and domain‑deletion analyses revealed that Nrf1 and C/EBPβ interact via their bZIP domains, repress DSPP transcription through a shared repression domain that requires Nrf1 phosphorylation at Ser599. In undifferentiated odontoblasts, Nrf1 and C/EBPβ jointly repress DSPP promoter activity, whereas loss of their interaction in differentiated cells elevates DSPP transcription, establishing a novel regulatory mechanism.

Abstract

Terminal differentiation of odontoblasts, the principal cells in dentin formation, proceeds by synthesis of type I collagen and noncollagenous proteins. DSP and DPP are specific markers for terminally differentiated odontoblasts and are encoded by a single gene DSPP (dentin sialophosphoprotein). In an attempt to understand the molecular mechanisms required for tissue-specific expression of the DSPP gene, we have identified a novel interaction between two bZIP transcription factors, Nrf1 and the CCAAT enhancer-binding protein (C/EBP)beta. This interaction was confirmed by both immunoprecipitation and chromatin immunoprecipitation assays. In undifferentiated odontoblasts, Nrf1 and C/EBPbeta repress DSPP promoter activity individually and synergistically by cooperatively interacting with each other. This mutual interaction is facilitated by the bZIP domains in both the proteins. The repression domain in both Nrf1 and C/EBPbeta was determined, and deletion of this domain abolished transcriptional repression. In fully differentiated odontoblasts, the loss of interaction between Nrf1 and C/EBPbeta results in an increased DSPP transcription. Further, this interaction was found to be dependent on phosphorylation at Ser(599) of Nrf1. Thus, the physical interaction between Nrf1 and C/EBPbeta provide a novel mechanism for the transcriptional regulation of DSPP in odontoblasts.