Primate lentiviral virion infectivity factors are substrate receptors that assemble with cullin 5–E3 ligase through a HCCH motif to suppress APOBEC3G

Abstract

Cullin-Ring E3 ubiquitin ligases target substrates for ubiquitindependent, proteasome-mediated degradation and regulate critical cellular processes. These cullins assemble with cellular substrate receptor proteins through specific adaptor molecules. F-box-and BC-box-containing receptors use Skp1, ElonginB, and ElonginC as adaptors to recruit Cul1͞Cul7 and Cul2͞Cul5, respectively. At present, the determinants of Cul2 vs. Cul5 specificity for the BC-box-containing receptors are poorly defined. Here, we demonstrate that primate lentiviral Vif (virion infectivity factor) proteins represent previously uncharacterized substrate receptor proteins that contain divergent BC-box motifs. These molecules selectively assemble with a Cul5-E3 ligase to suppress the antiviral activity of autologous cytidine deaminase APOBEC3G. A previously unrecognized Hx 5Cx17-18Cx3-5H motif that is highly conserved among all primate lentiviral Vif proteins was found to be critical for the selective assembly and activity of Vif-Cul5-E3 ligase. Non-primate lentiviral Vif proteins, which lack this HCCH motif, displayed reduced interaction with Cul5. These data suggest that in addition to target protein specificity, substrate receptor proteins play important roles in cullin selection and functional assembly of cullinRing E3 ligases. The discovery of these viral substrate receptor molecules that recruit Cul5 through distinct mechanisms from cellular proteins may facilitate the identification of additional cellular factors that regulate cellular functions through Cul5-E3 ligase. Motifs in Vif that are absent from cellular proteins could also be targets for the development of innovative therapeutics. cullin-E3 ubiquitin ligase ͉ HIV-simian immunodeficiency virus virion infectivity factorT argeted protein degradation is one mechanism by which critical processes such as mitosis and the cell cycle are regulated (1). The specificity of protein degradation is mediated by members of the E3 ubiquitin ligase family, including the cullin-based E3 ligases. Cullins form a scaffold on which other components of the E3 ligase organize to bring the substrate into close proximity with the E2 ubiquitin-conjugating enzyme (2-5). E3 ligases are substratespecific, and cullin-based E3 ligases display striking similarities. In SCF (Skp1-Cul1-F-box) complexes, the adaptor protein Skp1 bridges the interaction between the Cul1 and substrate receptor proteins through specific interaction with an F-box (2-5). These substrate receptor proteins bind substrates through distinct protein-protein interaction domains (e.g

Cytidine deaminase APOBEC3B interacts with heterogeneous nuclear ribonucleoprotein K and suppresses hepatitis B virus expression

“…Lentiviruses such as HIV-1 and SIV encode the Vif protein, which assembles with cellular proteins Cul5, ElonginB and ElonginC to form an E3 ubiquitin ligase (Yu et al, 2003;Mehle et al, 2004;Kobayashi et al, 2005;Liu et al, 2005;Luo et al, 2005) that induces polyubiquitination and degradation of multiple APOBEC3 molecules (Conticello et al, 2003;Marin et al, 2003;Sheehy et al, 2003;Stopak et al, 2003;Yu et al, 2003;Liu et al, 2004;Mehle et al, 2004;Liu et al, 2005). Exogenous expression of APOBEC3G (A3G) and APOBEC3F (A3F) has also been found to reduce HBV core-associated DNA in transfected liver cell lines (Rosler et al, 2004;Turelli et al, 2004a,b;Noguchi et al, 2005;Suspene et al, 2005).…”

Zinc binding to the HCCH motif of HIV-1 virion infectivity factor induces a conformational change that mediates protein–protein interactions

“…The Vif N-terminal region contains several conserved Trp residues that are required for Apo3G recognition (22). HIV-1 Vif C114S and C133S mutants do not bind to Cul5 (9,10), and mutation of the conserved His and Cys residues of the HCCH motif in SIV AGM Vif inhibits Cul5 binding (12). Therefore, it appears that the Vif N-terminal region mediates Apo3G recognition (22), whereas the HCCH motif allows specific binding to Cul5 (12).…”

“…Vif promotes the ubiquitination and proteasomal degradation of Apo3G (5)(6)(7)(8). Interaction of Vif with a Cullin 5 (Cul5)-based E3 ligase is required for Apo3G degradation (9)(10)(11), and a recent study suggests that the conserved H-X 5 -C-X 17-18 -C-X 3-5 -H (HCCH) motif plays a vital role in the Vif-Cul5 interaction (12).…”

“…Several reports (9,10,12,(14)(15)(16)(17)(18)(19) have shown that mutations in the recently defined HCCH motif impair Vif function. In one report (19), Vif C114 (amino acid numbering used hereafter refers to the HIV-1 HXB2 Vif sequence) was proposed to constitute part of a cysteine protease active site involved in gp41 processing.…”

APOBEC3G: an intracellular centurion

“…The SLQ( Y/F)LA motif (amino acids 144-150) of Vif interacts directly with elongin C (Marin et al 2003;Yu et al 2003Yu et al , 2004cMehle et al 2004a,b). A cryptic zinc-coordination motif, His-Xaa 5 -Cys-Xaa 17-18 -Cys-Xaa 3-5 -His (HCCH, amino acids 108-139), connects Vif to cullin 5 (Luo et al 2005;Mehle et al 2006;Xiao et al 2006). Through these interactions, Vif recruits a ubiquitin ligase (E3) complex comprising elongin C, elongin B, cullin 5, Nedd8 and Rbx1 ( Yu et al 2003( Yu et al , 2004c) that mediates the polyubiquitylation and triggers degradation of A3G (Conticello et al 2003;Marin et al 2003;Sheehy et al 2003;Stopak et al 2003).…”