Analysis of regulatory mechanisms governing the developmental specific expression of intracisternal A-particle genes
The endogenous retrovirus, intracisternal A-particle (IAP), is expressed at unique stages during mouse embryogenesis, and is also transcriptionally activated during the in vitro differentiation of F9 cells. The current analysis of the cis-acting DNA elements and trans-acting protein factors that control IAP expression during F9 differentiation reveals a unique dual regulatory mechanism. An enhancer element, the IAP upstream enhancer (IUE) from $-$186 to $-$167 within the IAP LTR, is identified by transient transfection assays and found to be active in both undifferentiated and differentiated cell types. Band shift, methylation interference, and UV cross-linking analyses further reveal that a ubiquitous 65 Kd protein factor, the IAP upstream enhancer binding protein (IUEB), binds to the sequence GATGGCGC within the IUE. Site-specific methylation within the IUEB binding site strongly inhibits both IUEB binding and IUE transcriptional activity suggesting that methylation may play a role in specifically regulating IAP expression. Further transfection assays reveal that the IAP basal promoter, from $-$50 to +1, directs differentiation-specific expression and is repressed by the adenovirus E1A gene, consistent with the presence of an E1A-like activity in F9 cells. Band shift and UV cross-linking analyses identify a protein factor(s), consisting of 40 and 60 Kd polypeptides present in both undifferentiated and differentiated cells, that binds specifically from $-$50 to $-$18. This suggests that these DNA-binding polypeptides either alone or with additional cellular components direct differentiation-specific and E1A-responsive expression from the IAP basal promoter.
Lamb, Bruce Timothy, "Analysis of regulatory mechanisms governing the developmental specific expression of intracisternal A-particle genes" (1991). Dissertations available from ProQuest. AAI9125698.