A DNA element that regulates expression of an endogenous retrovirus during F9 cell differentiation is E1A dependent.
The retinoic acid-induced differentiation of F9 cells into parietal endoderm-like cells activates transcription of the endogenous mouse retrovirus, the intracisternal A-particle (IAP). To investigate the elements that control IAP gene differentiation-specific expression, we used methylation interference, Southwestern (DNA-protein), and transient-transfection assays and identified the IAP-proximal enhancer (IPE) element that directs differentiation-specific expression. We find that the IPE is inactive in undifferentiated F9 cells and active in differentiated parietal endoderm-like PYS-2 cells. Three proteins of 40, 60, and 68 kDa bind to the sequence GAGTAGAC located between nucleotides -53 and -47 within the IPE. The 40- and 68-kDa proteins from both the undifferentiated and differentiated cells exhibit similar DNA-binding activities. However, the 60-kDa protein from differentiated cells has greater binding activity than that from undifferentiated cells, suggesting a role for this protein in F9 differentiation-specific expression of the IAP gene. The IAP gene is negatively regulated by the adenovirus E1A proteins, and the E1A sequence responsible for repression is located at the N terminus, between amino acids 2 and 67. The DNA sequence that is the target of E1A repression also maps to the IPE element. Colocalization of the differentiation-specific and E1A-sensitive elements to the same protein-binding site within the IPE suggests that the E1A-like activity functions in F9 cells to repress IAP gene expression. Activation of the IAP gene may result when the E1A-like activity is lost or inactivated during F9 cell differentiation, followed by binding of the 60-kDa positive regulatory protein to the enhancer element.
Lamb
B T
BT
Wistar Institute of Anatomy and Biology, Philadelphia, Pennsylvania 19104.
Satyamoorthy
K
K
Solter
D
D
Basu
A
A
Xu
M Q
MQ
Weinmann
R
R
Howe
C C
CC
D017500:Adenovirus E1A Proteins / Q000378:metabolism*; D001483:Base Sequence; D002454:Cell Differentiation / Q000235:genetics*; D004279:DNA, Viral / Q000235:genetics*; D004742:Enhancer Elements, Genetic; D015967:Gene Expression Regulation, Viral; D007426:Genes, Intracisternal A-Particle / Q000235:genetics*; D008969:Molecular Sequence Data; D014157:Transcription Factors / Q000378:metabolism; D014162:Transfection; D014407:Tumor Cells, Cultured
D017500:Adenovirus E1A Proteins; D004279:DNA, Viral; D014157:Transcription Factors