N-Terminal Hemagglutinin Tag Renders Lysine-Deficient APOBEC3G Resistant to HIV-1 Vif-Induced Degradation by Reduced Polyubiquitination

Wang, Yudi; Shao, Qiujia; Yu, Xianghui; Kong, Wei; Hildreth, James E. K.; Liu, Bindong

doi:10.1128/jvi.01925-10

Cited by 28 publications

(35 citation statements)

References 81 publications

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“…2A), as we previously showed that the N-terminus of A3G is an essential ubiquitin acceptor site [27]. When an HA-tag was fused to the N-terminus of lysine-free A3G (HAA3G20R), the A3G mutant was found to be resistant to Vif-induced degradation and its polyubiquitination modification was drastically reduced [27]. Therefore, using HAA3G20R as a template, we generated single arginine to lysine point reversions in A3G positions 297, 301, 303 and 334 (referred to as HAA3G297_K, HAA3G301_K, HAA3G303_K and HAA3G334_K, respectively).…”

Section: Resultsmentioning

confidence: 95%

“…Each revertant mutant harbors an HA-tag (YDYVPDYA) fused to the N-terminus (schematic shown in Fig. 2A), as we previously showed that the N-terminus of A3G is an essential ubiquitin acceptor site [27]. When an HA-tag was fused to the N-terminus of lysine-free A3G (HAA3G20R), the A3G mutant was found to be resistant to Vif-induced degradation and its polyubiquitination modification was drastically reduced [27].…”

Section: Resultsmentioning

confidence: 99%

“…We also sought to determine the essential lysine residues for A3G polyubiquitination/degradation by using our lysine-free A3G mutant, HAA3G20R (schematic shown in Fig. 3A), which was previously reported to be resistant to Vif-induced degradation [27]. Based on this lysine-free construct, revertant mutants were created by substituting arginine to lysine at positions 297, 301, 303 and 334 (HAA3G4K) (Fig.…”

Section: Discussionmentioning

confidence: 99%

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Differential Contributions of Ubiquitin-Modified APOBEC3G Lysine Residues to HIV-1 Vif-Induced Degradation

Turner

Shao

Wang

et al. 2016

Journal of Molecular Biology

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Apolipoprotein B mRNA-editing enzyme-catalytic polypeptide-like 3G (A3G) is a host restriction factor that impedes HIV-1 replication. Viral integrity is salvaged by HIV-1 virion infectivity factor (Vif), which mediates A3G polyubiquitination and subsequent cellular depletion. Previous studies have implied that A3G polyubiquitination is essential for Vif-induced degradation. However, the contribution of polyubiquitination to the rate of A3G degradation remains unclear. Here we show that A3G polyubiquitination is essential for degradation. Inhibition of ubiquitin-activating enzyme E1 by PYR-41 or blocking the formation of ubiquitin chains by over-expressing the lysine to arginine mutation of ubiquitin K48 (K48R) inhibited A3G degradation. Our A3G mutagenesis study showed that lysine residues 297, 301, 303 and 334 were not sufficient to render lysine free A3G sensitive to Vif mediated degradation. Our data also confirms that Vif could induce ubiquitin chain formation on lysine residues interspersed throughout A3G. Notably, A3G degradation relied on the lysine residues involved in polyubiquitination. Although A3G and the A3G C-terminal mutant interacted with Vif and were modified by ubiquitin chains, the latter remained more resistant to Vif-induced degradation. Furthermore, the A3G C-terminal mutant, but not the N-terminal mutant, maintained potent antiviral activity in the presence of Vif. Taken together, our results suggest that the location of A3G ubiquitin modification is a determinant for Vif-mediated degradation, implying that in addition to polyubiquitination, other factors may play a key role in the rate of A3G degradation.

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Section: Resultsmentioning

confidence: 95%

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

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Differential Contributions of Ubiquitin-Modified APOBEC3G Lysine Residues to HIV-1 Vif-Induced Degradation

Turner

Shao

Wang

et al. 2016

Journal of Molecular Biology

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“…Some of these include APOBEC3G, TRIM5α, and tetherin (57, 58). APOBEC3G is incorporated into virus particles in an infected cell and when a new cell is infected it deaminates cytosines in the viral genome and inhibits reverse transcriptase (59). This results in the generation of mutations during reverse transcription essentially mutating the virus out of existence.…”

Section: Weak Iop Responsesmentioning

confidence: 99%

Prospects for Lentiviral Vector Mediated Prostaglandin F Synthase Gene Delivery in Monkey EyesIn vivo

Lee

Rasmussen²,

Filla³

et al. 2014

Current Eye Research

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Currently, the most effective outflow drugs approved for clinical use are prostaglandin F2α analogues, but these require daily topical self-dosing and have various intraocular, ocular surface and extraocular side effects. Lentiviral vector-mediated delivery of the prostaglandin F synthase (PGFS) gene, resulting in long-term reduction of IOP, may eliminate off-target tissue effects and the need for daily topical PGF2α self-administration. Lentiviral vector-mediated delivery of the PGFS gene to the anterior segment has been achieved in cats and non-human primates. Although these results are encouraging, our studies have identified a number of challenges that need to be overcome for prostaglandin gene therapy to be translated into the clinic. Using examples from our work in non-human primates, where we were able to achieve a significant reduction in IOP (2 mm Hg) for 5 months after delivery of the cDNA for bovine PGF synthase, we identify and discuss these issues and consider several possible solutions.

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“…Similarly, the peroxisome proliferatoractivated receptor  co-activator 1 (PGC1) is primarily degraded through the nuclear N-terminus-dependent ubiquitin proteasome pathway (Trausch-Azar et al, 2010;Wang et al, 2011;Yang et al, 2009).…”

Section: N-terminal Ubiquitylationmentioning

confidence: 99%

Non-canonical ubiquitin-based signals for proteasomal degradation

Kravtsova‐Ivantsiv

Ciechanover

2012

Journal of Cell Science

193

155

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Commentary 539Introduction Ubiquitylation [also known as ubiquitination, as coined by the discoverers of this modification with regard to its connection to proteolysis (Wilkinson, 2005)] is a three-step enzymatic reaction that is carried out by several enzymes: the ubiquitin-activating enzyme (E1), a ubiquitin carrier protein (E2; also known as ubiquitin-conjugating enzyme, UBC) and a ubiquitin-protein ligase (E3). An additional component of the ubiquitylation machinery has been described. This E4 enzyme is involved in elongation of short ubiquitin chains (Koegl et al., 1999). However, the requirement for an E4 activity appears to be limited to a small subset of substrates. Ubiquitylation-dependent proteasomal degradation is involved in the regulation of numerous cellular processes, including cell cycle progression, apoptosis, DNA repair, the maintenance of cellular quality control, autophagy, the regulation of transcription and receptor-mediated endocytosis (Mayer et al., 2005;Mayer et al., 2006;Mayer et al., 2008). In general, modification by ubiquitin serves as a recognition element in trans, whereby different downstream effectors bind to the ubiquitin-modified protein to affect its fate and/or function. In the case of proteasomal degradation, the ubiquitylated protein is recognized by the 26S proteasome and subsequently degraded (Dikic et al., 2009;Su and Lau, 2009).The widely accepted canonical signal for proteasomal degradation is a polyubiquitin chain that is anchored to the -NH 2 group of a lysine residue(s) in the substrate by an isopeptide bond and is assembled through the formation of isopeptide bonds between the C-terminal residue of one ubiquitin moiety (glycine 76) and lysine 48 of the previously conjugated ubiquitin moiety (Chau et al., 1989). Recent studies have reported, however, that other types of ubiquitin chains can also be recognized by the proteasome (Figs 1, 2). These include an ester-based linkage that connects ubiquitin to a threonine or serine residue in the substrate, and a thiolester-based linkage whereby ubiquitin is bound to a cysteine residue in the substrate (McDowell et al., 2010;Tait et al., 2007;Vosper et al., 2009). Ubiquitin can also be conjugated to the -NH 2 group of the N-terminal residue of the substrate. Instead of using lysine 48 for the linkage, polyubiquitin chains can also be assembled through one of the six additional lysine residues in the molecule. Such homogenous chains based on, for example, lysine 63 (Saeki et al., 2009), or heterogeneous chains in which different ubiquitinubiquitin linkages are found, have also been reported to target proteins for proteasomal degradation. Linear chains, in which the ubiquitin links are attached to one another 'head-to-tail', and heterologous chains, in which the links are made of ubiquitin and a ubiquitin-like protein, such as small ubiquitin-like modifier (SUMO), have additionally been shown to target proteins for proteasomal degradation. Surprisingly, it has been demonstrated that the proteasome does not necessarily have ...

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N-Terminal Hemagglutinin Tag Renders Lysine-Deficient APOBEC3G Resistant to HIV-1 Vif-Induced Degradation by Reduced Polyubiquitination

Cited by 28 publications

References 81 publications

Differential Contributions of Ubiquitin-Modified APOBEC3G Lysine Residues to HIV-1 Vif-Induced Degradation

Differential Contributions of Ubiquitin-Modified APOBEC3G Lysine Residues to HIV-1 Vif-Induced Degradation

Prospects for Lentiviral Vector Mediated Prostaglandin F Synthase Gene Delivery in Monkey EyesIn vivo

Non-canonical ubiquitin-based signals for proteasomal degradation

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