Abstract
Activator protein-1 (AP-1) regulates diverse gene responses triggered by environmental cues and virus-induced cellular stress. Although many signaling events leading to AP-1 activation have been described, the fundamental features underlying binding site selection and factor recruitment of dimeric AP-1 complexes to their target genes remain mostly uncharacterized. Using recombinant full-length human AP-1 dimers formed between c-Jun and Fos family members (c-Fos, FosB, Fra-1, Fra-2) for DNA binding and transcriptional analysis, we found that each of these AP-1 complex exhibits differential activity for distinct non-consensus AP-1 sites present in human papillomavirus (HPV), and each AP-1 complex is capable of activating transcription from in vitro-reconstituted HPV chromatin in a p300- and acetyl-CoA-dependent manner. Transcription from HPV chromatin requires AP-1-dependent and contact-driven recruitment of p300. Acetylation of dimeric AP-1 complexes by p300 enhances AP-1 binding to DNA. Using a human C-33A cervical cancer-derived cell line harboring the episomal HPV type 11 genome, we illustrate binding site selectivity recognized by c-Jun, JunB, JunD, and various Fos family members in a combinatorial and unique pattern, highlighting the diversity and importance of non-canonical binding site recognition by various AP-1 family proteins.
Highlights
Biochemical activity of dimeric Activator protein-1 (AP-1) family proteins remains to be characterized
Using an in vitro chromatin transcription system reconstituted with purified HeLa core histones, recombinant nucleosome assembly protein 1 (NAP-1) histone chaperon, and the ATP-utilizing chromatin assembly and remodeling factor (ACF), we have demonstrated that a specific form of AP-1 complex, c-Jun/c-Fos, is able to switch on transcription from an human papillomavirus (HPV) type 11 (HPV-11) upstream regulatory region (URR)-containing chromatin template [25]
Our successful purification of homodimeric c-Jun/c-Jun and heterodimeric c-Jun/Fos complexes made it possible for the first time to directly compare the functional properties of these dimeric AP-1 complexes in recognizing non-canonical AP-1 sequence elements found in a native HPV genome and define their transcriptional activity from in vitro-reconstituted HPV chromatin that faithfully recapitulates nucleosomal phasing seen in vivo [25]
Summary
Biochemical activity of dimeric AP-1 family proteins remains to be characterized. Results: 1) Binding to canonical and non-canonical sequence elements in HPV-11 is different among dimeric AP-1 family members. 2) p300-mediated acetylation is important for AP-1-dependent HPV chromatin transcription. With our recent adaptation of a bacterial co-expression system that enables us to purify full-length recombinant human AP-1 dimers [26], we are able to define the role of each AP-1 complex in HPV chromatin-dependent transcription and reveal novel AP-1 sites that have not yet been characterized because of their deviation from the consensus TRE All of these non-canonical AP-1 sites are present in different types of HPVs in various combinations, indicating the existence of an intricate interplay between cis-elements and trans-acting factors in generating the regulatory circuit diversity to modulate HPV transcription in response to constantly changeable cellular environments
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