Amino acids necessary for DNA contact and dimerization imply novel motifs in the papillomavirus E2 trans-activator.

Prakash, Sadhana; Grossman, Steven R.; Pepinsky, R. Blake; Laimins, Laimonis A.; Androphy, Elliot J.

doi:10.1101/gad.6.1.105

Cited by 56 publications

(62 citation statements)

References 41 publications

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“…A cysteine at position 340 has been shown to be essential for DNA binding. Converting this cysteine into an arginine abolished activation of transcription by BPV1 E2 (43). This cysteine is conserved within all E2 proteins (16).…”

Section: E2 Directly Binds To the Hat Domain Of P300mentioning

confidence: 96%

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

Müller

Ritzkowsky

Steger

2002

J Virol

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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...

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Section: E2 Directly Binds To the Hat Domain Of P300mentioning

confidence: 96%

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

Müller

Ritzkowsky

Steger

2002

J Virol

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“…The full-length BPV-I E2 protein is a 48 kDa phosphoprotein (28) which dimerises spontaneously in the absence of DNA to form stable molecules (29,30). The E2 proteins of all papillomaviruses are organised into three domains, as shown in Figure 1: a well-conserved N-terminal domain involved in transactivation, a central hinge of variable sequence and length, and a C-terminal domain responsible for DNA binding and dimerisation (31).…”

Section: Introductionmentioning

confidence: 99%

“…Recent crystallographic data for the core BPV-l E2 DNA binding domain (C-terminal 85 amino acids) has revealed a novel dimeric DNA binding/dimerisation motif previously undescribed or unpredicted in a eucaryotic transcription factor (32). Two monomers interact to form an eight-stranded antiparallel (3-barrel which is stabilised by an extensive network of hydrogen bonds encompassing a central hydrophobic core where two conserved tryptophans critical for dimerisation (30) are in van der Waals contact. Target DNA is bent over the barrel to align a pair of sy.mmetrically disposed recognition helices with the major groove of the conserved nucleotides in each palindromic half site.…”

Section: Introductionmentioning

confidence: 99%

Kinetic and equilibrium binding studies of the human papillomavirus type-16 transcription regulatory protein E2 interacting with core enhancer elements

Sanders

Maitland

1994

Nucl Acids Res

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The human papillomaviruses (HPVs) are a family of DNA viruses which cause benign tumours of the skin and mucosa that infrequently progress to malignant carcinoma. The E2 open reading frame of HPV is thought to encode a papillomavirus-specific transcription factor which also has a role in viral replication. The E2 proteins of all papillomaviruses studied to date have been shown to bind specifically to the common conserved sequence ACC(N)6GGT found at multiple locations in their genomes. In the case of HPV-16, a 'high risk' genital papillomavirus, the E2 protein is thought to negatively regulate expression of the major viral transforming genes E6 and E7, which have been directly implicated in the oncogenic process. However, little information exists concerning the relative or absolute affinities of the native HPV-16 protein for its palindromic recognition sequences; moreover, interpretation of any transcription or replication phenomena attributed to this protein is more complicated in the absence of such data. Here we describe the overexpression, purification and characterisation of the C-terminal 89 amino acids of the protein encompassing the DNA binding/dimerisation domain. We show that the recombinant protein purified from E.coli by a combination of non-group-specific chromatography steps retains high biological activity and is able to bind to all sites in the HPV-16 genome with high affinity ( -8x 10-11 M). In addition, kinetic studies show that the E2 -DNA complexes are very stable, with half-lives ranging from 2.15 to greater than 240 min, and that nucleotides internal and external to the conserved palindrome appear to influence stability.

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“…E2 is a site-specific DNA binding protein that dimerizes spontaneously in the absence of DNA (McBride et al, 1989;Prakash et al, 1992) and interacts directly with 12 nucleotide pair palindromic sequences found at conserved locations in all PVs (Fig. 5).…”

Section: E2 Transcriptional Modulatormentioning

confidence: 99%

“…These two functional domains are separated by a somewhat poorly conserved central hinge region of variable length (Giri & Yaniv, 1988;McBride et al, 1991;Prakash et al, 1992). The crystal structure for the C-terminal DNA binding domain of BPV E2 (Hegde et al, 1992) and the NMR structure of HPV-31 E2 (Liang et al, 1996) bound to a double-stranded palindromic recognition sequence have been solved.…”

Section: E2 Transcriptional Modulatormentioning

confidence: 99%

Molecular Targets for Human Papillomaviruses: Prospects for Antiviral Therapy

Phelps

Barnes

Lobe

1998

Antivir Chem Chemother

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A substantial medical need exists for the development of antiviral medicines for the treatment of diseases associated with infection by human papillomaviruses (HPVs). HPVs are associated with various benign and malignant lesions including benign genital condyloma, common skin warts, laryngeal papillomas and anogenital cancer. Since treatment options are limited and typically not very satisfactory, the development of safe and effective antiviral drugs for HPV could have substantial clinical impact. In the last few years, exciting advances have been made in our understanding of papillomavirus replication and the effects that the virus has on growth of the host cell. Although still somewhat rudimentary, techniques have been developed for limited virion production in vitro offering the promise of more rapid advances in the dissection and understanding of the virus life cycle. Of the 8–10 HPV gene products that are made during infection, only one encodes enzymatic activities, the E1 helicase. Successful antiviral therapies have traditionally targeted viral enzymes such as polymerases, kinases and proteases. In contrast, macromolecular interactions which mediate the functions of E6, E7 and E2 are thought to be more difficult targets for small molecule therapy.

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Amino acids necessary for DNA contact and dimerization imply novel motifs in the papillomavirus E2 trans-activator.

Cited by 56 publications

References 41 publications

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

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

Kinetic and equilibrium binding studies of the human papillomavirus type-16 transcription regulatory protein E2 interacting with core enhancer elements

Molecular Targets for Human Papillomaviruses: Prospects for Antiviral Therapy

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