Abstract
The Epstein-Barr (EBV) virus induces a lytic state after infecting epithelial cells. Subsequently, there is infection of B lymphocytes with two types of cycles, latent and lytic. Apart from linkage of the EBV latent membrane protein-1 (LMP-1) with benign and malignant conditions of squamous epithelial cells, little is known about other EBV gene products that may be important in these processes as well as cellular transcriptional factors that regulate EBV gene expression in these epithelial cells. The EBV ED-L2 promoter, an early lytic cycle promoter, is located upstream of a transcription start site for a short open reading frame designated BNLF2 and just downstream of the BNLF1 (LMP-1) open reading frame. We have previously used the EBV ED-L2 promoter to target oncogenes in transgenic mice, resulting in tissue-specific expression in the tongue, esophagus, forestomach, and skin, all sharing stratifying squamous epithelia, alternatively called keratinocytes. In the present study, we have functionally dissected the ED-L2 promoter by making deletion constructs fused to the luciferase reporter gene with transient transfections into squamous and nonsquamous epithelial cell lines as well as B lymphocytes. A CACCC box-like cis-regulatory element has been identified that is located between -218 and -187 base pairs of the ED-L2 promoter that confers significant promoter activity only in squamous epithelial cells. This cis-regulatory element is active in a heterologous minimal herpes simplex virus thymidine kinase promoter reporter gene construct when transfected into squamous epithelial cells but not in nonsquamous epithelial cells. DNA gel mobility shift assays have led to the identification of DNA-protein complexes that bind the CACCC box-like element. One of these proteins is a novel transcriptional factor that is uniquely active in stratified squamous epithelial cells, designated as keratinocyte specific factor (KSF). KSF may be related to Sp1 but appears to be distinct from Sp1. In addition, KSF may interact with related or identical cis-regulatory elements found in human papillomavirus-11 E6 and cytokeratin K3 promoters that are active in keratinocytes. In aggregate, KSF may be important in the transcriptional regulation of viral and eukaryotic genes in keratinocytes.
Highlights
latent membrane protein-1 (LMP-1) does play a significant role, the repertoire of genes of Epstein-Barr virus (EBV) that are involved in producing the lytic cycle in epithelial cells remains to be elucidated, and the cellular transcriptional factors in the epithelial cells that govern the transcriptional regulation of these genes require further investigation
We have identified in the present study a CACCC box-like cis-regulatory element in the EBV-ED L2 promoter that interacts with several nuclear transcriptional factors, one of which appears to be keratinocyte-specific
The EBV ED-L2 Promoter Is Active in Keratinocytes Derived from Esophageal and Skin Stratified Squamous Epithelia— Since 782 bp of the EBV ED-L2 promoter proved to be sufficient in achieving gene expression in a tissue-specific fashion, this region was subjected to functional dissection by transient transfection of various cell lines with a series of plasmid constructs in which the full-length or serially deleted EBV ED-L2 promoter sequences were ligated to the luciferase reporter gene (Fig. 1A)
Summary
The first is a lytic infection where a high copy number of EBV genomes is initiated, followed by production of viral particles that can be released from the host cell. While much investigation has sought to elucidate EBV’s role in the immortalization and transformation of B lymphocytes, thereby leading to lymphoproliferative diseases [1] such as Burkitt’s lymphoma, Hodgkin’s disease, T-cell lymphoma, among others, very little is known about the molecular mechanisms underlying EBV’s lytic infection of oropharyngeal epithelial cells. LMP-1 transcripts are associated with nasopharyngeal carcinoma [6] In this context, LMP-1 does play a significant role, the repertoire of genes of EBV that are involved in producing the lytic cycle in epithelial cells remains to be elucidated, and the cellular transcriptional factors in the epithelial cells that govern the transcriptional regulation of these genes require further investigation. XhoI-L2 AS BamHI-L2 S4 BamHI-L2 S8 BamHI-L2 S7 BamHI-L2 S6 BamHI-L2 S9 BamHI-L2 S10 BamHI-L2 S11 BamHI-L2 S5 Luciferase AS
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
