Transcription factors Zic1 and Zic2 bind and transactivate the apolipoprotein E gene promoter

We have used the yeast one-hybrid system to identify transcription factors that bind to specific sequences in proximal regions of the apolipoprotein E gene promoter. The sequence between -163 and -124, that has been previousl y defined as a functional promoter element, was used as a bait to screen a human brain cDNA library. Ten cDNA clones that encoded portions of the huma n Zic1 (five clones) and Zic2 (five clones) transcription factors were isol ated. Electrophoretic mobility shift assays confirmed the presence of a bin ding site for Zic1 and Zic2 in the -136/-125 region. Displacement of bindin g with oligonucleotides derived from adjacent sequences within the APOE pro moter revealed the existence of two additional Zic-binding sequences in thi s promoter. These sequences were identified by electrophoretic mobility shi ft assays and mutational analysis in regions -65/-54 and -185/-174. Cotrans fection of Zic1 and Zic2 expression vector and different APOE promoter-luci ferase reporter constructs in U87 glioblastoma cell line showed that the th ree binding sites partially contributed to the trans-stimulation of the luc iferase reporter. Ectopic expression of Zic1 and Zic2 in U87 cells also tra ns-stimulated the expression of the endogenous gene, increasing the amount of apolipopro tein E produced by glial cells. These data indicate that Zic proteins might contribute to the transcriptional activity of the apolipopro tein E gene and suggest that apolipoprotein E could mediate some of the dev elopmental processes in which Zic proteins are involved.