Ap-1 Recognizes Sequence Elements on Hiv-1 LTR in Human Epithelial Tumor-Cell Lines

Abstract

Abstract. Investigation of the nucleotide sequence of the HIV-1 LTR showed the presence of four novel short DNA regions which are homologous to the recognition site for the cellular transcription factor AP-1. Four short oligonucleotide hybrids containing these potential AP-1 sites were constructed and used in gel retardation assays and in competition experiments in order to determine the role of the AP-1 protein in the regulation of HIV-1 expression. The breast MDA MB 468 and cervical HeLa tumor cell lines, which are known to overexpress the AP-1 protein were used in a gel retardation assay as a control to study the affinity of the AP-1 to synthesized oligonucleotide sequences. We have observed specific binding of nuclear factor AP-1 to three of these oligonucleotide hybrids. These results demonstrate the presence of three novel AP-1 binding sites on HIV-1 LTR, one of which was found within the TAR element and in the Tat protein binding region. Moreover, they suggest that AP-1 could be contributing to HIV-1 transcriptional regulation through its interaction with the AP-1 binding sites of HIV-1 LTR.

Human Lung, Bladder and Head and Neck Tumors as Compared to Their Adjacent Normal-Tissues Have Elevated Ap-1 Activity and Recognize Sequence Elements of Hiv-1 LTR

“…The AP-1 activity is elevated in 9/12 lung tumors, in 9/14 bladder tumors and in 7/7 head and neck carcinoma tumors as compared to normal human tissue. These results are in good agreement with our previous study (34) and suggest that these new HIV-1 LTR putative AP-1 sites have the same binding effect as nuclear extracts from HeLa and MDA MB 468 tumor cell lines.…”

“…This is not surprising since EjAP-1 contains the AP-1 binding consensus from the E 3 promoter (37), while lab and 4ab are highly homologous (71.5%) to the same sequence (34). However, the non- III II I III I III III II II I II III "All tissue specimens were obtained from lung cancer patients.…”

“…Four short DNA regions which are putative AP-1 binding sites were identified in the HIV-1 LTR (34). From these four sequences, region 2 shows the highest homology (85.8%) to the AP-1 consensus (5'-TGAGTCA-3') and therefore the corresponding oligonucleotide hybrid 2ab was used in our previous studies to test the affinity of the AP-1 protein in MDA MB 468 and HeLa tumor cells.…”

“…DNA oligonucleotides were as described (34). The oligonucleotides were purified as described (34) and the double stranded oligonucleotides were 5' end-labelled using γ 32 Ρ-ΑΤΡ and T4 polynucleotide kinase and end filled using the Klenow fragment of DNA polymerase according to Maniatis et al (36).…”

Binding of Wild-Type and Mutant Forms of P53 Protein From Human Tumors to a Specific Dna-Sequence of the First Intron of the H-Ras Oncogene

“…The pentanucleotide p53 binding repeats are underlined. SP-1 oligonucleotide was used as a p53-unrelated oligonucleotide competitor (32). Radioactive end labelling was performed using T4 polynucleotide kinase (Boehringer) and [γ- DNA binding assay.…”