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Michele Pagano - One of the best experts on this subject based on the ideXlab platform.
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mechanisms and function of substrate recruitment by f box Proteins
Nature Reviews Molecular Cell Biology, 2013Co-Authors: Jeffrey R Skaar, Julia K Pagan, Michele PaganoAbstract:S phase kinase-associated protein 1 (SKP1)-cullin 1 (CUL1)-F-Box protein (SCF) ubiquitin ligase complexes use a family of F-Box Proteins as substrate adaptors to mediate the degradation of a large number of regulatory Proteins involved in diverse processes. The dysregulation of SCF complexes and their substrates contributes to multiple pathologies. In the 14 years since the identification and annotation of the F-Box protein family, the continued identification and characterization of novel substrates has greatly expanded our knowledge of the regulation of substrate targeting and the roles of F-Box Proteins in biological processes. Here, we focus on the evolution of our understanding of substrate recruitment by F-Box Proteins, the dysregulation of substrate recruitment in disease and potential avenues for F-Box protein-directed disease therapies.
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deregulated proteolysis by the f box Proteins skp2 and beta trcp tipping the scales of cancer
Nature Reviews Cancer, 2008Co-Authors: David Frescas, Michele PaganoAbstract:Cell cycle progression is regulated by phosphorylation and protein degradation, which is mediated by ubiquitin ligases. This Review explores the relevance of two ubiquitin ligase specificity factors (F-Box Proteins) that are emerging as important players in tumour development. The maintenance and preservation of distinct phases during the cell cycle is a highly complex and coordinated process. It is regulated by phosphorylation — through the activity of cyclin-dependent kinases (CDKs) — and protein degradation, which occurs through ubiquitin ligases such as SCF (SKP1–CUL1–F-Box protein) complexes and APC/C (anaphase-promoting complex/cyclosome). Here, we explore the functionality and biology of the F-Box Proteins, SKP2 (S-phase kinase-associated protein 2) and β-TrCP (β-transducin repeat-containing protein), which are emerging as important players in cancer biogenesis owing to the deregulated proteolysis of their substrates.
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deregulated proteolysis by the f box Proteins skp2 and beta trcp tipping the scales of cancer
Nature Reviews Cancer, 2008Co-Authors: David Frescas, Michele PaganoAbstract:The maintenance and preservation of distinct phases during the cell cycle is a highly complex and coordinated process. It is regulated by phosphorylation--through the activity of cyclin-dependent kinases (CDKs)--and protein degradation, which occurs through ubiquitin ligases such as SCF (SKP1-CUL1-F-Box protein) complexes and APC/C (anaphase-promoting complex/cyclosome). Here, we explore the functionality and biology of the F-Box Proteins, SKP2 (S-phase kinase-associated protein 2) and beta-TrCP (beta-transducin repeat-containing protein), which are emerging as important players in cancer biogenesis owing to the deregulated proteolysis of their substrates.
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insights into scf ubiquitin ligases from the structure of the skp1 skp2 complex
Nature, 2000Co-Authors: Brenda A Schulman, Philip D Jeffrey, Michele Pagano, Andrea C Carrano, Zachary Bowen, E Kinnucan, Michael S Finnin, Nikola P PavletichAbstract:F-Box Proteins are members of a large family that regulates the cell cycle, the immune response, signalling cascades and developmental programmes by targeting Proteins, such as cyclins, cyclin-dependent kinase inhibitors, IκBα and β-catenin, for ubiquitination (reviewed in refs 1,2,3). F-Box Proteins are the substrate-recognition components of SCF (Skp1–Cullin–F-Box protein) ubiquitin-protein ligases4,5. They bind the SCF constant catalytic core by means of the F-Box motif interacting with Skp1, and they bind substrates through their variable protein–protein interaction domains6. The large number of F-Box Proteins is thought to allow ubiquitination of numerous, diverse substrates6. Most organisms have several Skp1 family members, but the function of these Skp1 homologues and the rules of recognition between different F-Box and Skp1 Proteins remain unknown. Here we describe the crystal structure of the human F-Box protein Skp2 bound to Skp1. Skp1 recruits the F-Box protein through a bipartite interface involving both the F-Box and the substrate-recognition domain. The structure raises the possibility that different Skp1 family members evolved to function with different subsets of F-Box Proteins, and suggests that the F-Box protein may not only recruit substrate, but may also position it optimally for the ubiquitination reaction.
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the f box protein family
Genome Biology, 2000Co-Authors: Edward T Kipreos, Michele PaganoAbstract:The F-Box is a protein motif of approximately 50 amino acids that functions as a site of protein-protein interaction. F-Box Proteins were first characterized as components of SCF ubiquitin-ligase complexes (named after their main components, Skp I, Cullin, and an F-Box protein), in which they bind substrates for ubiquitin-mediated proteolysis. The F-Box motif links the F-Box protein to other components of the SCF complex by binding the core SCF component Skp I. F-Box Proteins have more recently been discovered to function in non-SCF protein complexes in a variety of cellular functions. There are 11 F-Box Proteins in budding yeast, 326 predicted in Caenorhabditis elegans, 22 in Drosophila, and at least 38 in humans. F-Box Proteins often include additional carboxy-terminal motifs capable of protein-protein interaction; the most common secondary motifs in yeast and human F-Box Proteins are WD repeats and leucine-rich repeats, both of which have been found to bind phosphorylated substrates to the SCF complex. The majority of F-Box Proteins have other associated motifs, and the functions of most of these Proteins have not yet been defined.
Mike Tyers - One of the best experts on this subject based on the ideXlab platform.
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inhibition of scf ubiquitin ligases by engineered ubiquitin variants that target the cul1 binding site on the skp1 f box interface
Proceedings of the National Academy of Sciences of the United States of America, 2016Co-Authors: Maryna Gorelik, Igor Kurinov, Stephen Orlicky, Maria A Sartori, Mike Tyers, Xiaojing Tang, Jack Greenblatt, Edyta Marcon, Jason MoffatAbstract:Skp1–Cul1–F-Box (SCF) E3 ligases play key roles in multiple cellular processes through ubiquitination and subsequent degradation of substrate Proteins. Although Skp1 and Cul1 are invariant components of all SCF complexes, the 69 different human F-Box Proteins are variable substrate binding modules that determine specificity. SCF E3 ligases are activated in many cancers and inhibitors could have therapeutic potential. Here, we used phage display to develop specific ubiquitin-based inhibitors against two F-Box Proteins, Fbw7 and Fbw11. Unexpectedly, the ubiquitin variants bind at the interface of Skp1 and F-Box Proteins and inhibit ligase activity by preventing Cul1 binding to the same surface. Using structure-based design and phage display, we modified the initial inhibitors to generate broad-spectrum inhibitors that targeted many SCF ligases, or conversely, a highly specific inhibitor that discriminated between even the close homologs Fbw11 and Fbw1. We propose that most F-Box Proteins can be targeted by this approach for basic research and for potential cancer therapies.
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The F-Box: a new motif for ubiquitin dependent proteolysis in cell cycle regulation and signal transduction.
Progress in biophysics and molecular biology, 1999Co-Authors: Karen L. Craig, Mike TyersAbstract:Abstract The ubiquitin system of intracellular protein degradation controls the abundance of many critical regulatory Proteins. Specificity in the ubiquitin system is determined largely at the level of substrate recognition, a step that is mediated by E3 ubiquitin ligases. Analysis of the mechanisms of phosphorylation directed proteolysis in cell cycle regulation has uncovered a new class of E3 ubiquitin ligases called SCF complexes, which are composed of the subunits Skp1, Rbx1, Cdc53 and any one of a large number of different F-Box Proteins. The substrate specificity of SCF complexes is determined by the interchangeable F-Box protein subunit, which recruits a specific set of substrates for ubiquitination to the core complex composed of Skp1, Rbx1, Cdc53 and the E2 enzyme Cdc34. F-Box Proteins have a bipartite structure — the shared F-Box motif links F-Box Proteins to Skp1 and the core complex, whereas divergent protein–protein interaction motifs selectively bind their cognate substrates. To date all known SCF substrates are recognised in a strictly phosphorylation dependent manner, thus linking intracellular signalling networks to the ubiquitin system. The plethora of different F-Box Proteins in databases suggests that many pathways will be governed by SCF-dependent proteolysis. Indeed, genetic analysis has uncovered roles for F-Box Proteins in a variety of signalling pathways, ranging from nutrient sensing in yeast to conserved developmental pathways in plants and animals. Moreover, structural analysis has revealed ancestral relationships between SCF complexes and two other E3 ubiquitin ligases, suggesting that the combinatorial use of substrate specific adaptor Proteins has evolved to allow the regulation of many cellular processes. Here, we review the known signalling pathways that are regulated by SCF complexes and highlight current issues in phosphorylation dependent protein degradation.
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combinatorial control in ubiquitin dependent proteolysis don t skp the f box hypothesis
Trends in Genetics, 1998Co-Authors: Mike Tyers, Elizabeth E Patton, Andrew WillemsAbstract:Abstract The ubiquitin-dependent proteolytic pathway targets many key regulatory Proteins for rapid intracellular degradation. Specificity in protein ubiquitination derives from E3 ubiquitin protein ligases, which recognize substrate Proteins. Recently, analysis of the E3s that regulate cell division has revealed common themes in structure and function. One particularly versatile class of E3s, referred to as Skp1p–Cdc53p–F-Box protein (SCF) complexes, utilizes substrate-specific adaptor subunits called F-Box Proteins to recruit various substrates to a core ubiquitination complex. A vast array of F-Box Proteins have been revealed by genome sequencing projects, and the early returns from genetic analysis in several organisms promise that F-Box Proteins will participate in the regulation of many processes, including cell division, transcription, signal transduction and development.
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f box Proteins are receptors that recruit phosphorylated substrates to the scf ubiquitin ligase complex
Cell, 1997Co-Authors: Dorota Skowyra, Mike Tyers, Stephen J Elledge, Karen L. Craig, Wade J HarperAbstract:We have reconstituted the ubiquitination pathway for the Cdk inhibitor Sic1 using recombinant Proteins. Skp1, Cdc53, and the F-Box protein Cdc4 form a complex, SCFCdc4, which functions as a Sic1 ubiquitin-ligase (E3) in combination with the ubiquitin conjugating enzyme (E2) Cdc34 and E1. Cdc4 assembled with Skp1 functions as the receptor that selectively binds phosphorylated Sic1. Grr1, an F-Box protein involved in Cln destruction, forms complexes with Skp1 and Cdc53 and binds phosphorylated Cln1 and Cln2, but not Sic1. Because the constituents of the SCF complex are members of protein families, SCFCdc4 is likely to serve as the prototype for a large class of E3s formed by combinatorial interactions of related family members. SCF complexes couple protein kinase signaling pathways to the control of protein abundance.
Gary Kleiger - One of the best experts on this subject based on the ideXlab platform.
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ubiquitin ligation to f box protein targets by scf rbr e3 e3 super assembly
Nature, 2021Co-Authors: Daniel Hornghetko, David T Krist, Rajan J Prabu, Kheewoong Baek, Monique P C Mulder, Maren Klugel, Daniel C Scott, Huib Ovaa, Gary KleigerAbstract:E3 ligases are typically classified by hallmark domains such as RING and RBR, which are thought to specify unique catalytic mechanisms of ubiquitin transfer to recruited substrates1,2. However, rather than functioning individually, many neddylated cullin–RING E3 ligases (CRLs) and RBR-type E3 ligases in the ARIH family—which together account for nearly half of all ubiquitin ligases in humans—form E3–E3 super-assemblies3–7. Here, by studying CRLs in the SKP1–CUL1–F-Box (SCF) family, we show how neddylated SCF ligases and ARIH1 (an RBR-type E3 ligase) co-evolved to ubiquitylate diverse substrates presented on various F-Box Proteins. We developed activity-based chemical probes that enabled cryo-electron microscopy visualization of steps in E3–E3 ubiquitylation, initiating with ubiquitin linked to the E2 enzyme UBE2L3, then transferred to the catalytic cysteine of ARIH1, and culminating in ubiquitin linkage to a substrate bound to the SCF E3 ligase. The E3–E3 mechanism places the ubiquitin-linked active site of ARIH1 adjacent to substrates bound to F-Box Proteins (for example, substrates with folded structures or limited length) that are incompatible with previously described conventional RING E3-only mechanisms. The versatile E3–E3 super-assembly may therefore underlie widespread ubiquitylation. Cryo-electron microscopy of neddylated SCF-family ligases interacting with the RBR-type E3 ligase ARIH1 reveals the steps through which E3–E3 super-assemblies ubiquitylate a diverse set of substrates presented on F-Box Proteins.
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ubiquitin ligation to f box protein targets by scf rbr e3 e3 super assembly
Nature, 2021Co-Authors: Daniel Hornghetko, David T Krist, Rajan J Prabu, Kheewoong Baek, Monique P C Mulder, Maren Klugel, Daniel C Scott, Huib Ovaa, Gary KleigerAbstract:E3 ligases are typically classified by hallmark domains such as RING and RBR, which are thought to specify unique catalytic mechanisms of ubiquitin transfer to recruited substrates1,2. However, rather than functioning individually, many neddylated cullin-RING E3 ligases (CRLs) and RBR-type E3 ligases in the ARIH family-which together account for nearly half of all ubiquitin ligases in humans-form E3-E3 super-assemblies3-7. Here, by studying CRLs in the SKP1-CUL1-F-Box (SCF) family, we show how neddylated SCF ligases and ARIH1 (an RBR-type E3 ligase) co-evolved to ubiquitylate diverse substrates presented on various F-Box Proteins. We developed activity-based chemical probes that enabled cryo-electron microscopy visualization of steps in E3-E3 ubiquitylation, initiating with ubiquitin linked to the E2 enzyme UBE2L3, then transferred to the catalytic cysteine of ARIH1, and culminating in ubiquitin linkage to a substrate bound to the SCF E3 ligase. The E3-E3 mechanism places the ubiquitin-linked active site of ARIH1 adjacent to substrates bound to F-Box Proteins (for example, substrates with folded structures or limited length) that are incompatible with previously described conventional RING E3-only mechanisms. The versatile E3-E3 super-assembly may therefore underlie widespread ubiquitylation.
David T Krist - One of the best experts on this subject based on the ideXlab platform.
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ubiquitin ligation to f box protein targets by scf rbr e3 e3 super assembly
Nature, 2021Co-Authors: Daniel Hornghetko, David T Krist, Rajan J Prabu, Kheewoong Baek, Monique P C Mulder, Maren Klugel, Daniel C Scott, Huib Ovaa, Gary KleigerAbstract:E3 ligases are typically classified by hallmark domains such as RING and RBR, which are thought to specify unique catalytic mechanisms of ubiquitin transfer to recruited substrates1,2. However, rather than functioning individually, many neddylated cullin–RING E3 ligases (CRLs) and RBR-type E3 ligases in the ARIH family—which together account for nearly half of all ubiquitin ligases in humans—form E3–E3 super-assemblies3–7. Here, by studying CRLs in the SKP1–CUL1–F-Box (SCF) family, we show how neddylated SCF ligases and ARIH1 (an RBR-type E3 ligase) co-evolved to ubiquitylate diverse substrates presented on various F-Box Proteins. We developed activity-based chemical probes that enabled cryo-electron microscopy visualization of steps in E3–E3 ubiquitylation, initiating with ubiquitin linked to the E2 enzyme UBE2L3, then transferred to the catalytic cysteine of ARIH1, and culminating in ubiquitin linkage to a substrate bound to the SCF E3 ligase. The E3–E3 mechanism places the ubiquitin-linked active site of ARIH1 adjacent to substrates bound to F-Box Proteins (for example, substrates with folded structures or limited length) that are incompatible with previously described conventional RING E3-only mechanisms. The versatile E3–E3 super-assembly may therefore underlie widespread ubiquitylation. Cryo-electron microscopy of neddylated SCF-family ligases interacting with the RBR-type E3 ligase ARIH1 reveals the steps through which E3–E3 super-assemblies ubiquitylate a diverse set of substrates presented on F-Box Proteins.
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ubiquitin ligation to f box protein targets by scf rbr e3 e3 super assembly
Nature, 2021Co-Authors: Daniel Hornghetko, David T Krist, Rajan J Prabu, Kheewoong Baek, Monique P C Mulder, Maren Klugel, Daniel C Scott, Huib Ovaa, Gary KleigerAbstract:E3 ligases are typically classified by hallmark domains such as RING and RBR, which are thought to specify unique catalytic mechanisms of ubiquitin transfer to recruited substrates1,2. However, rather than functioning individually, many neddylated cullin-RING E3 ligases (CRLs) and RBR-type E3 ligases in the ARIH family-which together account for nearly half of all ubiquitin ligases in humans-form E3-E3 super-assemblies3-7. Here, by studying CRLs in the SKP1-CUL1-F-Box (SCF) family, we show how neddylated SCF ligases and ARIH1 (an RBR-type E3 ligase) co-evolved to ubiquitylate diverse substrates presented on various F-Box Proteins. We developed activity-based chemical probes that enabled cryo-electron microscopy visualization of steps in E3-E3 ubiquitylation, initiating with ubiquitin linked to the E2 enzyme UBE2L3, then transferred to the catalytic cysteine of ARIH1, and culminating in ubiquitin linkage to a substrate bound to the SCF E3 ligase. The E3-E3 mechanism places the ubiquitin-linked active site of ARIH1 adjacent to substrates bound to F-Box Proteins (for example, substrates with folded structures or limited length) that are incompatible with previously described conventional RING E3-only mechanisms. The versatile E3-E3 super-assembly may therefore underlie widespread ubiquitylation.
Hui Zhang - One of the best experts on this subject based on the ideXlab platform.
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electrostatic potentials of the s locus f box Proteins contribute to the pollen s specificity in self incompatibility in petunia hybrida
Plant Journal, 2017Co-Authors: Yue Zhang, Hui Zhang, Yanzhai Song, Yongbiao Xue, Jiangbo Fan, Dongfen ZhangAbstract:Summary Self-incompatibility (SI) is a self/non-self discrimination system widely found in angiosperms and, in many species, is controlled by a single polymorphic S-locus. In the Solanaceae, Rosaceae and Plantaginaceae, the S-locus encodes a single S-RNase and a cluster of S-locus F-Box (SLF) Proteins to control the pistil and pollen expression of SI, respectively. Previous studies have shown that their cytosolic interactions determine their recognition specificity, but the physical force between their interactions remains unclear. In this study, we show that the electrostatic potentials of SLF contribute to the pollen S specificity through a physical mechanism of “like charges repel and unlike charges attract” between SLFs and S-RNases in Petunia hybrida. Strikingly, alteration of a single C-terminal amino acid of SLF reversed its surface electrostatic potentials and subsequently the pollen S specificity. Collectively, our results reveal that the electrostatic potentials act as a major physical force between cytosolic SLFs and S-RNases, providing a mechanistic insight into the self/non-self discrimination between cytosolic Proteins in angiosperms. This article is protected by copyright. All rights reserved.
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cand1 binds to unneddylated cul1 and regulates the formation of scf ubiquitin e3 ligase complex
Molecular Cell, 2002Co-Authors: Jianyu Zheng, Jennifer M Harrell, Sophia Ryzhikov, Eunhee Shim, Karin Lykkeandersen, Ryuji Kobayashi, Xiaoming Yang, Hui ZhangAbstract:Abstract The SCF ubiquitin E3 ligase regulates ubiquitin-dependent proteolysis of many regulatory Proteins such as p27 Kip1 , IκB, and β-catenin. We report the isolation of a CUL1 binding protein, p120 CAND1 . We found the majority of CUL1 is in a complex with CAND1 and ROC1 independent of SKP1 and F box protein SKP2. Both in vivo and in vitro, CAND1 prevents the binding of SKP1 and SKP2 to CUL1 while dissociation of CAND1 from CUL1 promotes the reverse reaction. Neddylation of CUL1 or the presence of SKP1 and ATP causes CAND1 dissociation. Our data suggest that CAND1 regulates the formation of the SCF complex, and its dissociation from CUL1 is coupled with the incorporation of F box Proteins into the SCF complex, causing their destabilization.
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cand1 binds to unneddylated cul1 and regulates the formation of scf ubiquitin e3 ligase complex
Molecular Cell, 2002Co-Authors: Jianyu Zheng, Jennifer M Harrell, Sophia Ryzhikov, Eunhee Shim, Karin Lykkeandersen, Ryuji Kobayashi, Xiaoming Yang, Ning Wei, Hong Sun, Hui ZhangAbstract:The SCF ubiquitin E3 ligase regulates ubiquitin-dependent proteolysis of many regulatory Proteins such as p27(Kip1), IkappaB, and beta-catenin. We report the isolation of a CUL1 binding protein, p120(CAND1). We found the majority of CUL1 is in a complex with CAND1 and ROC1 independent of SKP1 and F box protein SKP2. Both in vivo and in vitro, CAND1 prevents the binding of SKP1 and SKP2 to CUL1 while dissociation of CAND1 from CUL1 promotes the reverse reaction. Neddylation of CUL1 or the presence of SKP1 and ATP causes CAND1 dissociation. Our data suggest that CAND1 regulates the formation of the SCF complex, and its dissociation from CUL1 is coupled with the incorporation of F box Proteins into the SCF complex, causing their destabilization.
