Summary auto-generated
This study investigates transcriptional regulation of the Epstein-Barr virus BZLF1 Z promoter in human epithelial cells, specifically examining how epithelial cell differentiation affects viral gene expression. Using the SCC12F epithelial cell line, researchers identified a differentiation-responsive regulatory element within the BZLF1 promoter located at the ZII domain (TGACATCA sequence, positioned at -67 to -60 bp). Through gel mobility shift assays and supershift experiments, the authors demonstrated that the transcription factors CREB and ATF-1 specifically bind to the ZII element. Crucially, they found that ZII binding capacity increases approximately 3-fold when nuclear extracts are prepared from differentiated cells compared to undifferentiated cells. Immunoblot analysis confirmed that both CREB and ATF-1 protein abundance increases during SCC12F cell differentiation. The study also showed that mutation of the ZII binding site abolishes the promoter's responsiveness to differentiation signals. These findings establish a molecular mechanism linking epithelial cell differentiation status to EBV lytic cycle initiation through CREB and ATF-1-mediated transcriptional activation of BZLF1.
Key findings
- The ZII element (TGACATCA) within the BZLF1 Z promoter functions as the differentiation-responsive regulatory element in epithelial cells
- CREB and ATF-1 transcription factors specifically bind the ZII element as homodimers and heterodimers
- ZII-binding capacity increases approximately 3-fold in differentiated SCC12F cells compared to undifferentiated cells
- CREB and ATF-1 protein abundance increases during epithelial cell differentiation, correlating with enhanced promoter responsiveness
- Mutation of the ZII binding site eliminates differentiation-dependent BZLF1 promoter activation
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Abstract
Epstein--Barr virus (EBV) is a ubiquitous human herpesvirus and an important human pathogen. Initiation of the EBV lytic cycle is dependent upon transcription of the EBV BZLF1 gene. Our previous studies of transcriptional regulation of the BZLF1 Z promoter (Zp) in human SCC12F epithelial cells identified a region within Zp that is responsive to epithelial cell differentiation. In the present study, we localize this differentiation responsive element to the CREB/AP-1-like binding site (TGACATCA) between -67 to -60 bp within Zp, previously designated ZII, and furthermore show that homodimers and heterodimers of CREB and ATF-1 specifically bind ZII. Consistent with a regulatory role for CREB and ATF-1 in differentiation dependent BZLF1 expression, ZII was able to bind approximately 3-fold more CREB and ATF-1 when incubated with nuclear extract obtained from populations of SCC12F cells enriched for the differentiated phenotype than when incubated with extract obtained from populations enriched for the undifferentiated phenotype. In addition, CREB and ATF-1 were found to increase in abundance during SCC12F differentiation. These results indicate a regulatory role for CREB and ATF-1 in differentiation-dependent expression of BZLF1 in human epithelial cells.