Cooperative Activation of Human Papillomavirus Type 8 Gene Expression by the E2 Protein and the Cellular Coactivator p300

Abstract

The E2 proteins of papillomaviruses (PV) bind to the coactivator CBP/p300 as do many other transcription factors, but the precise role of CBP/p300 in E2-specific functions is not yet understood. We show that the E2 protein of human PV type 8 (HPV8) directly binds to p300. Activation of HPV8 gene expression by low amounts of HPV8 E2 was stimulated up to sevenfold by coexpression of p300. The interaction between E2 and p300 may play a role in differentiation-dependent activation of PV gene expression, since we can show that the expression level of p300 increases during keratinocyte differentiation. Surprisingly, sequence-specific binding of E2 to its recognition sites within the regulatory region of HPV8 is not necessary for this cooperation, indicating that E2 can be recruited to the promoter via protein-protein interaction. HPV8 E2 binds via its N-terminal activation domain (AD), its C-terminal DNA binding domain (DBD), and its internal hinge region to p300 in vitro. Transient-transfection assays revealed that the AD is necessary and sufficient for cooperative activation with p300. However, we provide evidence that the interaction of the hinge and the DBD of HPV8 E2 with p300 may contribute. Our data suggest an important role of p300 in regulation of HPV8 gene expression and reveal a new mechanism by which E2 may be recruited to a promoter to activate transcription without sequence specific DNA binding.Papillomaviruses (PV) infect the basal cells of the skin or the mucosa, causing proliferative lesions like warts or dysplasias. PV require differentiating keratinocytes to replicate their DNA. The expression of the structural proteins is restricted to some of the most-differentiated keratinocytes. This cell tropism is due to the involvement of transcription factors specifically expressed in these cells. In addition to ubiquitously expressed and keratinocyte-specific cellular transcription factors, viral gene expression is also modulated by the viral E2 protein. The E2 protein binds via its carboxy (C)-terminal DNA binding domain (DBD) and dimerization domain to the 12-bp palindromic sequence ACCN 6 GGT mostly located within the regulatory region, also called the long control region or noncoding region (NCR) of the PV genome. In the case of bovine PV type 1 (BPV1), the long control region contains 12 E2 binding sites, which mediate a strong activation of several BPV1 promoters by E2 (48-50). Human PV (HPV) types infecting the genital mucosa contain four E2 binding sites in conserved positions. Here, only a moderate activation by E2 could be detected, and E2 repressed HPV gene expression in most cases. This repression is mediated by binding to two promoter-proximal E2 binding sites, as revealed by transient transfections of cervical carcinoma cell lines and immortalized skin keratinocytes (10,52,58,59). Repression of HPV gene expression by the E2 activator protein occurs when E2 binding sites are overlapping with the binding sites for cellular transcription factors necessary for promoter activity. In contrast to nat...

Interaction between the HPV E7 oncoprotein and the transcriptional coactivator p300

“…E2 interacts with the CBP KIX domain, which is required for stable interaction with CREB (Kwok et al, 1994), and this interaction appears to enhance the ability of E2 to activate viral gene expression. This interaction may also play a role in the differentiation-dependent activation of viral gene expression, since recent studies have shown that the level of p300 expression increases during keratinocyte differentiation (Mu¨ller et al, 2002). It is therefore intriguing that the interaction between E7 and p300 results in abrogation of E2-p300 coactivator function.…”

The Human Papillomavirus Type 8 E2 Protein Suppresses β4-Integrin Expression in Primary Human Keratinocytes

“…Terminal differentiation of keratinocytes, however, is required to complete the viral life cycle. Recently, it has been shown that E2 binds to and cooperates with cellular transcription factors, which are involved in keratinocyte differentiation (17,23,27,28,32). In noninfected epithelia, differentiation is triggered by loss of integrin-mediated anchorage to extracellular matrix (1,24,49).…”

“…Moreover, E2 proteins from different HPVs cooperate with cellular transcription factors involved in keratinocyte differentiation. These include C/EBP factors, p300, and p53 (17,23,27,28,32). It has recently been demonstrated that the E2 protein of the genital high-risk virus HPV18 even induces apoptosis in primary keratinocytes lacking other viral proteins (6).…”

The Role of E2 Proteins in Papillomavirus DNA Replication

“…This association is driving the E2-mediated tethering of viral genomes to host chromatin, and at the same time is retaining the viral genomes in transcriptionally active regions of the nucleus to escape silencing (Jang et al, 2009;Kurg et al, 2005). The transactivation domain mediates functional interactions with cellular transcription factors Sp1 and AP1, histone acetylase complexes containing CBP/p300 and pCAF, and with nucleosome assembly protein hNAP1 (Lee et al, 2002a;Lee et al, 2000;Li et al, 1991;Müller et al, 2002;Rehtanz et al, 2004;Thierry et al, 1992). E2-dependent activities are also modulated by interactions with Brm, a chromatin remodeling protein associated with SWI/SNF ATP-dependent chromatin remodeling complex, and with EP400, a component of the NuA4/TIP60 histone acetylase complex, and SMCX, also known as histone demethylase JARID1C , and Tax1BP1 (Kumar et al, 2007;Smith et al, 2010;Wang et al, 2009b).…”