The VP16 accessory protein HCF is a family of polypeptides processed from a large precursor protein

Wilson, Angus C.; LaMarco, Kelly; Peterson, Michael Gregory; Herr, Winship

doi:10.1016/0168-9525(93)90037-i

Cited by 91 publications

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“…One function of the spacer region in HCF-1 is to specify proteolytic processing (Wilson et al, 1993b;Kristie et al, 1995;Wilson et al, 1995b;Vogel and Kristie, 2000). To determine if the dHCF protein is also processed, we transiently transfected Drosophila SL2 cells with an expression vector encoding full-length dHCF tagged at the N-terminus with the T7 epitope and at the C-terminus with a FLAG epitope (illustrated schematically in Fig.…”

Section: Proteolytic Processing Of Dhcfmentioning

confidence: 99%

“…In addition, HCF-1 contains its own activation domain and this is required for optimal transactivation by the VP16-induced complex ( The arrangement of functional domains in human HCF-1 is shown in Figure 1A. HCF-1 is translated as a 230-300-kDa precursor and processed into N-and C-terminal subunits through autocatalytic proteolytic cleavage at a series of six HCF PRO repeats located near the center of the precursor polypeptide (Wilson et al, 1993b;Kristie et al, 1995;Wilson et al, 1995b;Vogel and Kristie, 2000). The resulting subunits remain stably associated via two matched pairs of interaction domains: termed SAS1N-SAS1C and SAS2N-SAS2C .…”

mentioning

confidence: 99%

“…The SAS2 self-association domains flank the HCF PRO repeat region and have been mapped with less precision. The majority of HCF-1 molecules in actively proliferating cells have undergone processing suggesting this is the active form of the protein (Wilson et al, 1993b;Kristie et al, 1995;Wilson et al, 1995b). …”

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Molecular cloning of Drosophila HCF reveals proteolytic processing and self‐association of the encoded protein

Mahajan

Johnson

Wilson

2002

Journal Cellular Physiology

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HCF-1 functions as a coactivator for herpes simplex virus VP16 and a number of mammalian transcription factors. Mature HCF-1 is composed of two subunits generated by proteolytic cleavage of a larger precursor at six centrally-located HCF PRO repeats. The resulting N-and Cterminal subunits remain tightly associated via two complementary pairs of self-association domains: termed SAS1N-SAS1C and SAS2N-SAS2C. Additional HCF proteins have been identified in mammals (HCF-2) and Caenorhabditis elegans (CeHCF). Both contain wellconserved SAS1 domains but do not undergo proteolytic processing. Thus, the significance of the cleavage and self-association of HCF-1 remains enigmatic. Here, we describe the isolation of the Drosophila HCF homologue (dHCF) using a genetic screen based on conservation of the SAS1 interaction. The N-terminal β-propeller domain of dHCF supports VP16-induced complex formation and is more similar to mammalian HCF-1 than other homologues. We show that fulllength dHCF expressed in Drosophila cells undergoes proteolytic cleavage giving rise to tightly associated N-and C-terminal subunits. As with HCF-1, the SAS1N and SAS1C elements of dHCF are separated by a large central region, however, this sequence lacks obvious homology to the HCF PRO repeats required for HCF-1 cleavage. The conservation of HCF processing in insect cells argues that formation of separate N-and C-terminal subunits is important for HCF function.HCF-1, also known as C1 factor, is a broadly-expressed nuclear protein involved in the regulation of cellular and viral gene expression. HCF-1 was first identified through its role in transcriptional activation by the herpes simplex virus (HSV) VP16 protein (Gerster and Roeder, 1988;Kristie et al., 1989). Association of HCF-1 with VP16 (also known as α-TIF or Vmw65) leads to the assembly of a multiprotein-DNA complex that includes the POU protein Oct-1 and the TAATGARAT element, a VP16-responsive DNA sequence present in all viral immediate-early (IE) gene promoters (reviewed in (Herr, 1998)). Nuclear localization of VP16 is dependent on association with HCF-1, which contributes a nuclear localization signal (NLS) located at its C-terminus (LaBoissière et al., 1999;Scarr et al., 2000;Wilson et al., 2000). In addition, HCF-1 contains its own activation domain and this is required for optimal transactivation by the VP16-induced complex ( The arrangement of functional domains in human HCF-1 is shown in Figure 1A. HCF-1 is translated as a 230-300-kDa precursor and processed into N-and C-terminal subunits through autocatalytic proteolytic cleavage at a series of six HCF PRO repeats located near the center of the precursor polypeptide (Wilson et al., 1993b;Kristie et al., 1995;Wilson et al., 1995b;Vogel and Kristie, 2000). The resulting subunits remain stably associated via two matched pairs of interaction domains: termed SAS1N-SAS1C and SAS2N-SAS2C . The SAS1C domain corresponds to a pair of fibronectin type 3 (Fn3) repeats, which interact with a 43-residue sequence (SAS1N) that is locate...

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Section: Proteolytic Processing Of Dhcfmentioning

confidence: 99%

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Molecular cloning of Drosophila HCF reveals proteolytic processing and self‐association of the encoded protein

Mahajan

Johnson

Wilson

2002

Journal Cellular Physiology

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“…Activators have been classified as acidic, glutamine-rich, proline-rich and serine/threonine-rich, depending on the preponderance of amino acids in their TAD 3 . An archetypal acidic activator is the herpes simplex virus protein 16 (VP16), which activates the expression of immediate early viral genes during infection [4][5][6][7][8] . VP16 exerts its activating function through a C-terminal TAD that includes residues 413-490 (refs.…”

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Structure and VP16 binding of the Mediator Med25 activator interaction domain

Vojnic

Mourão

Seizl

et al. 2011

Nat Struct Mol Biol

113

161

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4 0 4 VOLUME 18 NUMBER 4 APRIL 2011 nAture structurAl & moleculAr biologyActivator proteins regulate eukaryotic RNA polymerase II (Pol II) transcription in response to developmental and environmental signals. They bind to the DNA recognition sites of target genes with a sequence-specific DNA-binding domain, and recruit components of the Pol II machinery through a transcriptional activation domain (TAD) 1,2 . Activators have been classified as acidic, glutamine-rich, proline-rich and serine/threonine-rich, depending on the preponderance of amino acids in their TAD 3 . An archetypal acidic activator is the herpes simplex virus protein 16 (VP16), which activates the expression of immediate early viral genes during infection 4-8 . VP16 exerts its activating function through a C-terminal TAD that includes residues 413-490 (refs. 9-13). The VP16 TAD has been intensely used for studying transcriptional activation. Usually, the VP16 TAD is fused with its N terminus to the DNA-binding domain of the yeast transcription factor Gal4. The resulting Gal4-VP16 activator fusion protein stimulates transcription from promoters that contain Gal4-binding sites in the yeast and mammalian transcription systems in vivo 14,15 and in vitro 16,17 . This indicates that basic mechanisms of transcription activation are conserved amongst eukaryotes. The VP16 TAD targets basal Pol II transcription factors, including TFIIA, TFIIB, TFIID, the TFIIH subunit Tfb1/p62 (yeast/human) and the Mediator co-activator [18][19][20][21][22][23] . The VP16 TAD binds the Mediator subunit Med25 (also called Arc92) [24][25][26][27] , which is specific to higher eukaryotes. Med25 consists of two domains, the activator interaction domain (ACID) 24 that binds the VP16 TAD, and a 'von Willebrand factor type A' domain that anchors Med25 to Mediator 24 . Mediator generally conveys regulatory information by forming a bridge between activators and the basal Pol II machinery 28,29 . Whereas information on the core Mediator structure is emerging, structural information about its more peripheral activator-binding domains is limited to the KIX domain in subunit Med15 (or Arc105) 30,31 .The VP16 TAD is intrinsically flexible, whereas the VP16 core domain (residues 49-402) forms a stable structure 23,[32][33][34] . The TAD contains two functional subdomains, H1 (residues 410-452) and H2 (residues 453-490), which activate transcription independently 35,36 . Both H1 and H2 contain acidic amino acids, but specific bulky hydrophobic and aromatic residues are required for their function [36][37][38][39] . H2 has been proposed to adopt an α-helical conformation when bound to TFIIB 40 . NMR analysis revealed a nine-residue amphipathic α-helix in H2 that docks onto a PH fold in Tfb1 21 . On the basis of these and other results it is assumed that acidic TADs are flexible in their free state, but become transiently structured upon interaction with their target proteins.Here we report the NMR structure of the Mediator Med25 ACID and analyze its structural and functional interaction wi...

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“…16 VP16-mediated transcriptional activation occurs via interaction of VP16 and two cellular factors, Oct-1 [17][18][19] and HCF, 20,21 and subsequent binding of the complex to TAATGARAT elements found within HSV IE promoter regions. 22 This interaction results in robust up-regulation of IE gene expression.…”

Section: Figure 6 Amplicon Stock-mediated Cytotoxicity Is Not Increasmentioning

confidence: 99%

Expression of vhs and VP16 during HSV-1 helper virus-free amplicon packaging enhances titers

et al. 2001

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Recently developed helper virus-free methods of herpes simplex virus (HSV) amplicon vector packaging provide stocks that are virtually devoid of the cytotoxic component normally associated with traditional helper virus-based packaging methods. These approaches involve cotransfection of amplicon plasmid DNA with either a five-cosmid set or a bacterial artificial chromosome (BAC) that contains the HSV genome without its cognate pac signals. Helper virus-free amplicon packaging produces low-titer stocks (Ͻ10 5 expressing particles/ml) that exhibit a high frequency of pseudotransduction. In an effort to enhance amplicon titers, we introduced in trans a genomic copy of the virion host shutoff (vhs) protein-encoding gene UL41 into both cosmid-

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The VP16 accessory protein HCF is a family of polypeptides processed from a large precursor protein

Cited by 91 publications

References 0 publications

Molecular cloning of Drosophila HCF reveals proteolytic processing and self‐association of the encoded protein

Molecular cloning of Drosophila HCF reveals proteolytic processing and self‐association of the encoded protein

Structure and VP16 binding of the Mediator Med25 activator interaction domain

Expression of vhs and VP16 during HSV-1 helper virus-free amplicon packaging enhances titers

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