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Mahmut Safak - One of the best experts on this subject based on the ideXlab platform.
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human neurotropic polyomavirus jc virus Agnoprotein targets mitochondrion and modulates its functions
Virology, 2021Co-Authors: Reshu Saxena, Rafal Kaminski, Sami Saribas, Pooja Jadiya, Dhanendra Tomar, John W Elrod, Mahmut SafakAbstract:JC virus encodes an important regulatory protein, known as Agnoprotein (Agno). We have recently reported Agno's first protein-interactome with its cellular partners revealing that it targets various cellular networks and organelles, including mitochondria. Here, we report further characterization of the functional consequences of its mitochondrial targeting and demonstrated its co-localization with the mitochondrial networks and with the mitochondrial outer membrane. The mitochondrial targeting sequence (MTS) of Agno and its dimerization domain together play major roles in this targeting. Data also showed alterations in various mitochondrial functions in Agno-positive cells; including a significant reduction in mitochondrial membrane potential, respiration rates and ATP production. In contrast, a substantial increase in ROS production and Ca2+ uptake by the mitochondria were also observed. Finally, findings also revealed a significant decrease in viral replication when Agno MTS was deleted, highlighting a role for MTS in the function of Agno during the viral life cycle.
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a comprehensive proteomics analysis of jc virus Agnoprotein interacting proteins Agnoprotein primarily targets the host proteins with coiled coil motifs
Virology, 2020Co-Authors: Sami A Saribas, Prasun K Datta, Mahmut SafakAbstract:Abstract JC virus (JCV) Agnoprotein (Agno) plays critical roles in successful completion of the viral replication cycle. Understanding its regulatory roles requires a complete map of JCV-host protein interactions. Here, we report the first Agno interactome with host cellular targets utilizing “Two-Strep-Tag” affinity purification system coupled with mass spectroscopy (AP/MS). Proteomics data revealed that Agno primarily targets 501 cellular proteins, most of which contain “coiled-coil” motifs. Agno-host interactions occur in several cellular networks including those involved in protein synthesis and degradation; and cellular transport; and in organelles, including mitochondria, nucleus and ER-Golgi network. Among the Agno interactions, Rab11B, Importin and Crm-1 were first validated biochemically and further characterization was done for Crm-1, using a HIV-1 Rev-M10-like Agno mutant (L33D + E34L), revealing the critical roles of L33 and E34 residues in Crm-1 interaction. This comprehensive proteomics data provides new foundations to unravel the critical regulatory roles of Agno during the JCV life cycle.
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Expression of novel proteins by polyomaviruses and recent advances in the structural and functional features of Agnoprotein of JC virus, BK virus, and simian virus 40
Journal of Cellular Physiology, 2018Co-Authors: A. S. Saribas, Pascale Coric, Serge Bouaziz, Mahmut SafakAbstract:Polyomavirus family consists of a highly diverse group of small DNA viruses. The founding family member (MPyV) was first discovered in the newborn mouse in the late 1950s, which induces solid tumors in a wide variety of tissue types that are the epithelial and mesenchymal origin. Later, other family members were also isolated from a number of mammalian, avian and fish species. Some of these viruses significantly contributed to our current understanding of the fundamentals of modern biology such as transcription, replication, splicing, RNA editing, and cell transformation. After the discovery of first two human polyomaviruses (JC virus [JCV] and BK virus [BKV]) in the early 1970s, there has been a rapid expansion in the number of human polyomaviruses in recent years due to the availability of the new technologies and brought the present number to 14. Some of the human polyomaviruses cause considerably serious human diseases, including progressive multifocal leukoencephalopathy, polyomavirus-associated nephropathy, Merkel cell carcinoma, and trichodysplasia spinulosa. Emerging evidence suggests that the expression of the polyomavirus genome is more complex than previously thought. In addition to encoding universally expressed regulatory and structural proteins (LT-Ag, Sm t-Ag, VP1, VP2, and VP3), some polyomaviruses express additional virus-specific regulatory proteins and microRNAs. This review summarizes the recent advances in polyomavirus genome expression with respect to the new viral proteins and microRNAs other than the universally expressed ones. In addition, a special emphasis is devoted to the recent structural and functional discoveries in the field of polyomavirus Agnoprotein which is expressed only by JCV, BKV, and simian virus 40 genomes.
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structure based release analysis of the jc virus Agnoprotein regions a role for the hydrophilic surface of the major alpha helix domain in release
Journal of Cellular Physiology, 2018Co-Authors: Sami A Saribas, Martyn K. White, Mahmut SafakAbstract:Agnoprotein (Agno) is an important regulatory protein of JC virus (JCV), BK virus (BKV) and simian virus 40 (SV40) and these viruses are unable to replicate efficiently in the absence of this protein. Recent 3D-NMR structural data revealed that Agno contains two alpha-helices (a minor and a major) while the rest of the protein adopts an unstructured conformation (Coric et al., 2017, J Cell Biochem). Previously, release of the JCV Agno from the Agno-positive cells was reported. Here, we have further mapped the regions of Agno responsible for its release by a structure-based systematic mutagenesis approach. Results revealed that amino acid residues (Lys22, Lys23, Phe31, Glu34, and Asp38) located either on or adjacent to the hydrophilic surface of the major alpha-helix domain of Agno play critical roles in release. Additionally, Agno was shown to strongly interact with unidentified components of the cell surface when cells are treated with Agno, suggesting additional novel roles for Agno during the viral infection cycle.
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Nuclear Magnetic Resonance Structure of the Human Polyoma JC Virus Agnoprotein.
Journal of Cellular Biochemistry, 2017Co-Authors: Pascale Coric, A. S. Saribas, Wayne E. Childers, Magid Abou-gharbia, Martyn K. White, Mahmut Safak, Jon H. Condra, Serge BouazizAbstract:Agnoprotein is an important regulatory protein of the human polyoma JC virus (JCV) and plays critical roles during the viral replication cycle. It forms highly stable dimers and oligomers through its Leu/Ile/Phe-rich domain, which is important for the stability and function of the protein. We recently resolved the partial 3D structure of this protein by NMR using a synthetic peptide encompassing amino acids Thr17 to Gln52, where the Leu/Ile/Phe- rich region was found to adopt a major alpha-helix conformation spanning amino acids 23-39. Here, we report the resolution of the 3D structure of full-length JCV Agnoprotein by NMR, which not only confirmed the existence of the previously reported major α-helix domain at the same position but also revealed the presence of an additional minor α-helix region spanning amino acid residues Leu6 to lys13. The remaining regions of the protein adopt an intrinsically unstructured conformation. J. Cell. Biochem. 118: 3268-3280, 2017. © 2017 Wiley Periodicals, Inc.
Martyn K. White - One of the best experts on this subject based on the ideXlab platform.
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structure based release analysis of the jc virus Agnoprotein regions a role for the hydrophilic surface of the major alpha helix domain in release
Journal of Cellular Physiology, 2018Co-Authors: Sami A Saribas, Martyn K. White, Mahmut SafakAbstract:Agnoprotein (Agno) is an important regulatory protein of JC virus (JCV), BK virus (BKV) and simian virus 40 (SV40) and these viruses are unable to replicate efficiently in the absence of this protein. Recent 3D-NMR structural data revealed that Agno contains two alpha-helices (a minor and a major) while the rest of the protein adopts an unstructured conformation (Coric et al., 2017, J Cell Biochem). Previously, release of the JCV Agno from the Agno-positive cells was reported. Here, we have further mapped the regions of Agno responsible for its release by a structure-based systematic mutagenesis approach. Results revealed that amino acid residues (Lys22, Lys23, Phe31, Glu34, and Asp38) located either on or adjacent to the hydrophilic surface of the major alpha-helix domain of Agno play critical roles in release. Additionally, Agno was shown to strongly interact with unidentified components of the cell surface when cells are treated with Agno, suggesting additional novel roles for Agno during the viral infection cycle.
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Nuclear Magnetic Resonance Structure of the Human Polyoma JC Virus Agnoprotein.
Journal of Cellular Biochemistry, 2017Co-Authors: Pascale Coric, A. S. Saribas, Wayne E. Childers, Magid Abou-gharbia, Martyn K. White, Mahmut Safak, Jon H. Condra, Serge BouazizAbstract:Agnoprotein is an important regulatory protein of the human polyoma JC virus (JCV) and plays critical roles during the viral replication cycle. It forms highly stable dimers and oligomers through its Leu/Ile/Phe-rich domain, which is important for the stability and function of the protein. We recently resolved the partial 3D structure of this protein by NMR using a synthetic peptide encompassing amino acids Thr17 to Gln52, where the Leu/Ile/Phe- rich region was found to adopt a major alpha-helix conformation spanning amino acids 23-39. Here, we report the resolution of the 3D structure of full-length JCV Agnoprotein by NMR, which not only confirmed the existence of the previously reported major α-helix domain at the same position but also revealed the presence of an additional minor α-helix region spanning amino acid residues Leu6 to lys13. The remaining regions of the protein adopt an intrinsically unstructured conformation. J. Cell. Biochem. 118: 3268-3280, 2017. © 2017 Wiley Periodicals, Inc.
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emerging from the unknown structural and functional features of Agnoprotein of polyomaviruses
Journal of Cellular Physiology, 2016Co-Authors: Sami A Saribas, Pascale Coric, Wayne E. Childers, Martyn K. White, Jon H. Condra, Magid Abougharbia, Anahit Hamazaspyan, William E Davis, Rachael Axman, Serge BouazizAbstract:Agnoprotein is an important regulatory protein of polyomaviruses, including JCV, BKV, and SV40. In the absence of its expression, these viruses are unable to sustain their productive life cycle. It is a highly basic phosphoprotein that localizes mostly to the perinuclear area of infected cells, although a small amount of the protein is also found in nucleus. Much has been learned about the structure and function of this important regulatory protein in recent years. It forms highly stable dimers/oligomers in vitro and in vivo through its Leu/Ile/Phe-rich domain. Structural NMR studies revealed that this domain adopts an alpha-helix conformation and plays a critical role in the stability of the protein. It associates with cellular proteins, including YB-1, p53, Ku70, FEZ1, HP1α, PP2A, AP-3, PCNA, and α-SNAP; and viral proteins, including small t antigen, large T antigen, HIV-1 Tat, and JCV VP1; and significantly contributes the viral transcription and replication. This review summarizes the recent advances in the structural and functional properties of this important regulatory protein. J. Cell. Physiol. 231: 2115-2127, 2016. © 2016 Wiley Periodicals, Inc.
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correction for coric et al nuclear magnetic resonance structure revealed that the human polyomavirus jc virus Agnoprotein contains an α helix encompassing the leu ile phe rich domain
Journal of Virology, 2014Co-Authors: Pascale Coric, A. S. Saribas, Wayne E. Childers, Serge Bouaziz, Martyn K. White, Magid Abougharbia, Mahmut SafakAbstract:Volume 88, no. 12, p. [6556–6575][1], 2014. Page 6556, abstract, line 5: “Glu55” should read “Gln52.” Page 6556, importance, line 4: “Q54” should read “Q52.” Page 6557, column 2, second full paragraph, line 3: “Glu54” should read “Gln52.” The authors regret these errors
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Nuclear Magnetic Resonance Structure Revealed that the Human Polyomavirus JC Virus Agnoprotein Contains an α-Helix Encompassing the Leu/Ile/Phe-Rich Domain
Journal of Virology, 2014Co-Authors: Pascale Coric, A. S. Saribas, Wayne E. Childers, Serge Bouaziz, Magid Abou-gharbia, Martyn K. White, Mahmut SafakAbstract:ABSTRACT Agnoprotein is a small multifunctional regulatory protein required for sustaining the productive replication of JC virus (JCV). It is a mostly cytoplasmic protein localizing in the perinuclear area and forms highly stable dimers/oligomers through a Leu/Ile/Phe-rich domain. There have been no three-dimensional structural data available for Agnoprotein due to difficulties associated with the dynamic conversion from monomers to oligomers. Here, we report the first nuclear magnetic resonance (NMR) structure of a synthetic Agnoprotein peptide spanning amino acids Thr17 to Glu55 where Lys23 to Phe39 encompassing the Leu/Ile/Phe-rich domain forms an amphipathic α-helix. On the basis of these structural data, a number of Ala substitution mutations were made to investigate the role of the α-helix in the structure and function of Agnoprotein. Single L29A and L36A mutations exhibited a significant negative effect on both protein stability and viral replication, whereas the L32A mutation did not. In addition, the L29A mutant displayed a highly nuclear localization pattern, in contrast to the pattern for the wild type (WT). Interestingly, a triple mutant, the L29A+L32A+L36A mutant, yielded no detectable Agnoprotein expression, and the replication of this JCV mutant was significantly reduced, suggesting that Leu29 and Leu36 are located at the dimer interface, contributing to the structure and stability of Agnoprotein. Two other single mutations, L33A and E34A, did not perturb Agnoprotein stability as drastically as that observed with the L29A and L36A mutations, but they negatively affected viral replication, suggesting that the role of these residues is functional rather than structural. Thus, the Agnoprotein dimerization domain can be targeted for the development of novel drugs active against JCV infection. IMPORTANCE Agnoprotein is a small regulatory protein of JC virus (JCV) and is required for the successful completion of the viral replication cycle. It forms highly stable dimers and oligomers through its hydrophobic (Leu/Ile/Phe-rich) domain, which has been shown to play essential roles in the stability and function of the protein. In this work, the Leu/Ile/Phe-rich domain has been further characterized by NMR studies using an Agnoprotein peptide spanning amino acids T17 to Q54. Those studies revealed that the dimerization domain of the protein forms an amphipathic α-helix. Subsequent NMR structure-based mutational analysis of the region highlighted the critical importance of certain amino acids within the α-helix for the stability and function of Agnoprotein. In conclusion, this study provides a solid foundation for developing effective therapeutic approaches against the dimerization domain of the protein to inhibit its critical roles in JCV infection.
Kamel Khalili - One of the best experts on this subject based on the ideXlab platform.
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the Agnoprotein of polyomavirus jc is released by infected cells evidence for its cellular uptake by uninfected neighboring cells
Virology, 2014Co-Authors: Onder Otlu, Kamel Khalili, Francesca Isabella De Simone, Yolandalopez Otalora, Ilker Kudret SariyerAbstract:Poliomavirus JC replicates in glial cells in the brain, and causes the fatal demyelinating disease, progressive multifocal leukoencephalopathy (PML). PML is usually seen in patients with underlying immunocompromised conditions, notably among AIDS patients and those on chronic immunosuppressive regimens. The late leader sequence of JC virus contains an open reading frame encoding a small regulatory protein called Agnoprotein. Agnoprotein contributes to progressive viral infection by playing significant roles in viral replication cycle. Here, we demonstrate that Agnoprotein can be detected in cell-free fractions of glial cultures infected with JCV, transfected with expression plasmids or transduced with an adenovirus expression system. We also provide evidence that extracellular Agnoprotein can be taken up by uninfected neighboring cells. These studies have revealed a novel phenomenon of Agnoprotein during the viral life cycle with a potential of developing diagnostic and therapeutic interventions.
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jcv Agnoprotein induced reduction in cxcl5 lix secretion by oligodendrocytes is associated with activation of apoptotic signaling in neurons
Journal of Cellular Physiology, 2012Co-Authors: Nana Merabova, Barbara Krynska, Rafal M Kaminski, Shohreh Amini, Kamel Khalili, Armine DarbinyanAbstract:An indispensable role for oligodendrocytes in the protection of axon function and promotion of neuronal survival is strongly supported by the finding of progressive neuron/axon degeneration in human neurological diseases that affect oligodendrocytes. Imaging and pathological studies of the CNS have shown the presence of neuroaxonal injury in progressive multifocal leukoencephalopathy (PML), a demyelinating disease of the CNS, resulting from destruction of oligodendrocytes upon productive replication of the pathogenic neurotropic polyomavirus JC. Here, we examined the extracellular factors involved in communication between oligodendrocytes and neurons. Culturing cortical neurons with conditioned medium (CM) from rat CG4 oligodendrocytic cells that express the JCV Agnoprotein showed that CXCL5/LIX, which is a chemokine closely related to the human CXCL5/ENA78 and CXCL6/GCP-2 chemokines, is essential for neuronal cell survival. We found that in CM from Agnoprotein-producing CG-4 cells level of CXC5/LIX is decreased compared to control cells. We also demonstrated that a reduced expression of CXCL5/LIX by CG4 GFP-Agno cells triggered a cascade of signaling events in cortical neurons. Analysis of mitogen-activated protein kinases (MAPK) and glycogen synthase kinase (GSK3) pathways showed that they are involved in mechanisms of neuronal apoptosis in response to the depletion of CXCL5/LIX signaling. These data suggest that Agnoprotein-induced dysregulation of chemokine production by oligodendrocytes may contribute to neuronal/axonal injury in the pathogenesis of PML lesions.
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jcv Agnoprotein induced reduction in cxcl5 lix secretion by oligodendrocytes is associated with activation of apoptotic signaling in neurons
Journal of Cellular Physiology, 2012Co-Authors: Nana Merabova, Barbara Krynska, Rafal M Kaminski, Shohreh Amini, Kamel Khalili, Armine DarbinyanAbstract:An indispensable role for oligodendrocytes in the protection of axon function and promotion of neuronal survival is strongly supported by the finding of progressive neuron/axon degeneration in human neurological diseases that affect oligodendrocytes. Imaging and pathological studies of the CNS have shown the presence of neuroaxonal injury in progressive multifocal leukoencephalopathy (PML), a demyelinating disease of the CNS, resulting from destruction of oligodendrocytes upon productive replication of the pathogenic neurotropic polyomavirus JC. Here, we examined the extracellular factors involved in communication between oligodendrocytes and neurons. Culturing cortical neurons with conditioned medium (CM) from rat CG4 oligodendrocytic cells that express the JCV Agnoprotein showed that CXCL5/LIX, which is a chemokine closely related to the human CXCL5/ENA78 and CXCL6/GCP-2 chemokines, is essential for neuronal cell survival. We found that in CM from Agnoprotein-producing CG-4 cells level of CXC5/LIX is decreased compared to control cells. We also demonstrated that a reduced expression of CXCL5/LIX by CG4 GFP-Agno cells triggered a cascade of signaling events in cortical neurons. Analysis of mitogen-activated protein kinases (MAPK) and glycogen synthase kinase (GSK3) pathways showed that they are involved in mechanisms of neuronal apoptosis in response to the depletion of CXCL5/LIX signaling. These data suggest that Agnoprotein-induced dysregulation of chemokine production by oligodendrocytes may contribute to neuronal/axonal injury in the pathogenesis of PML lesions.
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Detection of Human Polyomavirus Proteins, T-Antigen and Agnoprotein, in Human Tumor Tissue Arrays
Journal of medical virology, 2010Co-Authors: Luís J. Del Valle, Kamel KhaliliAbstract:Expression of the human polyomavirus JCV genome in several experimental animals induces a variety of neural origin tumors. The viral proteins, T-antigen and Agnoprotein, contribute to the oncogenesis of JCV by associating with several tumor suppressor proteins and dysregulating signaling pathways, which results in uncontrolled cell proliferation. In addition, T-antigen and Agnoprotein have been associated with DNA damage and interfering with DNA repair mechanisms. In this study, we have utilized commercially available tissue arrays of human tumors of various origins and demonstrated the expression of both T-antigen and Agnoprotein in some, but not all, tumors of neural and non-neural origin. Most notably, more than 40% of human glioblastomas and greater than 30% of colon adenocarcinomas express viral proteins. The detection of viral transforming proteins, T-antigen and Agnoprotein in the absence of viral capsid proteins suggests a role for JCV in the development and/or progression of human tumors. These results invite further large-scale investigation on the role of polyomaviruses, particularly JCV in the pathogenesis of human cancer.
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dephosphorylation of jc virus Agnoprotein by protein phosphatase 2a inhibition by small t antigen
Virology, 2008Co-Authors: Ilker Kudret Sariyer, Kamel Khalili, Mahmut SafakAbstract:Previous studies have demonstrated that the JC virus (JCV) late regulatory protein Agnoprotein is phosphorylated by the serine/threonine-specific protein kinase-C (PKC) and mutants of this protein at the PKC phosphorylation sites exhibit defects in the viral replication cycle. We have now investigated whether Agnoprotein phosphorylation is regulated by PP2A, a serine/threonine-specific protein phosphatase and whether JCV small t antigen (Sm t-Ag) is involved in this regulation. Protein–protein interaction studies demonstrated that PP2A associates with Agnoprotein and dephosphorylates it at PKC-specific sites. Sm t-Ag was also found to interact with PP2A and this interaction inhibited the dephosphorylation of Agnoprotein by PP2A. The interaction domains of Sm t-Ag and Agnoprotein with PP2A were mapped, as were the interaction domains of Sm t-Ag with Agnoprotein. The middle portion of Sm t-Ag (aa 82–124) was found to be critical for the interaction with both Agnoprotein and PP2A and the N-terminal region of Agnoprotein for interaction with Sm t-Ag. To further understand the role of Sm t-Ag in JCV regulation, a stop codon was introduced at Ser90 immediately after splice donor site of the JCV early gene and the functional consequences of this mutation were investigated. The ability of this mutant virus to replicate was substantially reduced compared to WT. Next, the functional significance of PP2A in JCV replication was examined by siRNA targeting. Downregulation of PP2A caused a significant reduction in the level of JCV replication. Moreover, the impact of Sm t-Ag on Agnoprotein phosphorylation was investigated by creating a double mutant of JCV, where Sm t-Ag stop codon mutant was combined with an Agnoprotein triple phosphorylation mutant (Ser7, Ser11 and Thr21 to Ala). Results showed that double mutant behaves much like the triple phosphorylation mutant of Agnoprotein during viral replication cycle, which suggests that Agnoprotein might be an important target of Sm t-Ag with respect to the regulation of its phosphorylation. Collectively, these results suggest that there is an interplay between Agnoprotein, Sm t-Ag and PP2A with respect to the regulation of JCV life cycle and this could be important for the progression of the JCV-induced disease, PML.
Serge Bouaziz - One of the best experts on this subject based on the ideXlab platform.
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Expression of novel proteins by polyomaviruses and recent advances in the structural and functional features of Agnoprotein of JC virus, BK virus, and simian virus 40
Journal of Cellular Physiology, 2018Co-Authors: A. S. Saribas, Pascale Coric, Serge Bouaziz, Mahmut SafakAbstract:Polyomavirus family consists of a highly diverse group of small DNA viruses. The founding family member (MPyV) was first discovered in the newborn mouse in the late 1950s, which induces solid tumors in a wide variety of tissue types that are the epithelial and mesenchymal origin. Later, other family members were also isolated from a number of mammalian, avian and fish species. Some of these viruses significantly contributed to our current understanding of the fundamentals of modern biology such as transcription, replication, splicing, RNA editing, and cell transformation. After the discovery of first two human polyomaviruses (JC virus [JCV] and BK virus [BKV]) in the early 1970s, there has been a rapid expansion in the number of human polyomaviruses in recent years due to the availability of the new technologies and brought the present number to 14. Some of the human polyomaviruses cause considerably serious human diseases, including progressive multifocal leukoencephalopathy, polyomavirus-associated nephropathy, Merkel cell carcinoma, and trichodysplasia spinulosa. Emerging evidence suggests that the expression of the polyomavirus genome is more complex than previously thought. In addition to encoding universally expressed regulatory and structural proteins (LT-Ag, Sm t-Ag, VP1, VP2, and VP3), some polyomaviruses express additional virus-specific regulatory proteins and microRNAs. This review summarizes the recent advances in polyomavirus genome expression with respect to the new viral proteins and microRNAs other than the universally expressed ones. In addition, a special emphasis is devoted to the recent structural and functional discoveries in the field of polyomavirus Agnoprotein which is expressed only by JCV, BKV, and simian virus 40 genomes.
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Nuclear Magnetic Resonance Structure of the Human Polyoma JC Virus Agnoprotein.
Journal of Cellular Biochemistry, 2017Co-Authors: Pascale Coric, A. S. Saribas, Wayne E. Childers, Magid Abou-gharbia, Martyn K. White, Mahmut Safak, Jon H. Condra, Serge BouazizAbstract:Agnoprotein is an important regulatory protein of the human polyoma JC virus (JCV) and plays critical roles during the viral replication cycle. It forms highly stable dimers and oligomers through its Leu/Ile/Phe-rich domain, which is important for the stability and function of the protein. We recently resolved the partial 3D structure of this protein by NMR using a synthetic peptide encompassing amino acids Thr17 to Gln52, where the Leu/Ile/Phe- rich region was found to adopt a major alpha-helix conformation spanning amino acids 23-39. Here, we report the resolution of the 3D structure of full-length JCV Agnoprotein by NMR, which not only confirmed the existence of the previously reported major α-helix domain at the same position but also revealed the presence of an additional minor α-helix region spanning amino acid residues Leu6 to lys13. The remaining regions of the protein adopt an intrinsically unstructured conformation. J. Cell. Biochem. 118: 3268-3280, 2017. © 2017 Wiley Periodicals, Inc.
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emerging from the unknown structural and functional features of Agnoprotein of polyomaviruses
Journal of Cellular Physiology, 2016Co-Authors: Sami A Saribas, Pascale Coric, Wayne E. Childers, Martyn K. White, Jon H. Condra, Magid Abougharbia, Anahit Hamazaspyan, William E Davis, Rachael Axman, Serge BouazizAbstract:Agnoprotein is an important regulatory protein of polyomaviruses, including JCV, BKV, and SV40. In the absence of its expression, these viruses are unable to sustain their productive life cycle. It is a highly basic phosphoprotein that localizes mostly to the perinuclear area of infected cells, although a small amount of the protein is also found in nucleus. Much has been learned about the structure and function of this important regulatory protein in recent years. It forms highly stable dimers/oligomers in vitro and in vivo through its Leu/Ile/Phe-rich domain. Structural NMR studies revealed that this domain adopts an alpha-helix conformation and plays a critical role in the stability of the protein. It associates with cellular proteins, including YB-1, p53, Ku70, FEZ1, HP1α, PP2A, AP-3, PCNA, and α-SNAP; and viral proteins, including small t antigen, large T antigen, HIV-1 Tat, and JCV VP1; and significantly contributes the viral transcription and replication. This review summarizes the recent advances in the structural and functional properties of this important regulatory protein. J. Cell. Physiol. 231: 2115-2127, 2016. © 2016 Wiley Periodicals, Inc.
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correction for coric et al nuclear magnetic resonance structure revealed that the human polyomavirus jc virus Agnoprotein contains an α helix encompassing the leu ile phe rich domain
Journal of Virology, 2014Co-Authors: Pascale Coric, A. S. Saribas, Wayne E. Childers, Serge Bouaziz, Martyn K. White, Magid Abougharbia, Mahmut SafakAbstract:Volume 88, no. 12, p. [6556–6575][1], 2014. Page 6556, abstract, line 5: “Glu55” should read “Gln52.” Page 6556, importance, line 4: “Q54” should read “Q52.” Page 6557, column 2, second full paragraph, line 3: “Glu54” should read “Gln52.” The authors regret these errors
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Nuclear Magnetic Resonance Structure Revealed that the Human Polyomavirus JC Virus Agnoprotein Contains an α-Helix Encompassing the Leu/Ile/Phe-Rich Domain
Journal of Virology, 2014Co-Authors: Pascale Coric, A. S. Saribas, Wayne E. Childers, Serge Bouaziz, Magid Abou-gharbia, Martyn K. White, Mahmut SafakAbstract:ABSTRACT Agnoprotein is a small multifunctional regulatory protein required for sustaining the productive replication of JC virus (JCV). It is a mostly cytoplasmic protein localizing in the perinuclear area and forms highly stable dimers/oligomers through a Leu/Ile/Phe-rich domain. There have been no three-dimensional structural data available for Agnoprotein due to difficulties associated with the dynamic conversion from monomers to oligomers. Here, we report the first nuclear magnetic resonance (NMR) structure of a synthetic Agnoprotein peptide spanning amino acids Thr17 to Glu55 where Lys23 to Phe39 encompassing the Leu/Ile/Phe-rich domain forms an amphipathic α-helix. On the basis of these structural data, a number of Ala substitution mutations were made to investigate the role of the α-helix in the structure and function of Agnoprotein. Single L29A and L36A mutations exhibited a significant negative effect on both protein stability and viral replication, whereas the L32A mutation did not. In addition, the L29A mutant displayed a highly nuclear localization pattern, in contrast to the pattern for the wild type (WT). Interestingly, a triple mutant, the L29A+L32A+L36A mutant, yielded no detectable Agnoprotein expression, and the replication of this JCV mutant was significantly reduced, suggesting that Leu29 and Leu36 are located at the dimer interface, contributing to the structure and stability of Agnoprotein. Two other single mutations, L33A and E34A, did not perturb Agnoprotein stability as drastically as that observed with the L29A and L36A mutations, but they negatively affected viral replication, suggesting that the role of these residues is functional rather than structural. Thus, the Agnoprotein dimerization domain can be targeted for the development of novel drugs active against JCV infection. IMPORTANCE Agnoprotein is a small regulatory protein of JC virus (JCV) and is required for the successful completion of the viral replication cycle. It forms highly stable dimers and oligomers through its hydrophobic (Leu/Ile/Phe-rich) domain, which has been shown to play essential roles in the stability and function of the protein. In this work, the Leu/Ile/Phe-rich domain has been further characterized by NMR studies using an Agnoprotein peptide spanning amino acids T17 to Q54. Those studies revealed that the dimerization domain of the protein forms an amphipathic α-helix. Subsequent NMR structure-based mutational analysis of the region highlighted the critical importance of certain amino acids within the α-helix for the stability and function of Agnoprotein. In conclusion, this study provides a solid foundation for developing effective therapeutic approaches against the dimerization domain of the protein to inhibit its critical roles in JCV infection.
Pascale Coric - One of the best experts on this subject based on the ideXlab platform.
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Expression of novel proteins by polyomaviruses and recent advances in the structural and functional features of Agnoprotein of JC virus, BK virus, and simian virus 40
Journal of Cellular Physiology, 2018Co-Authors: A. S. Saribas, Pascale Coric, Serge Bouaziz, Mahmut SafakAbstract:Polyomavirus family consists of a highly diverse group of small DNA viruses. The founding family member (MPyV) was first discovered in the newborn mouse in the late 1950s, which induces solid tumors in a wide variety of tissue types that are the epithelial and mesenchymal origin. Later, other family members were also isolated from a number of mammalian, avian and fish species. Some of these viruses significantly contributed to our current understanding of the fundamentals of modern biology such as transcription, replication, splicing, RNA editing, and cell transformation. After the discovery of first two human polyomaviruses (JC virus [JCV] and BK virus [BKV]) in the early 1970s, there has been a rapid expansion in the number of human polyomaviruses in recent years due to the availability of the new technologies and brought the present number to 14. Some of the human polyomaviruses cause considerably serious human diseases, including progressive multifocal leukoencephalopathy, polyomavirus-associated nephropathy, Merkel cell carcinoma, and trichodysplasia spinulosa. Emerging evidence suggests that the expression of the polyomavirus genome is more complex than previously thought. In addition to encoding universally expressed regulatory and structural proteins (LT-Ag, Sm t-Ag, VP1, VP2, and VP3), some polyomaviruses express additional virus-specific regulatory proteins and microRNAs. This review summarizes the recent advances in polyomavirus genome expression with respect to the new viral proteins and microRNAs other than the universally expressed ones. In addition, a special emphasis is devoted to the recent structural and functional discoveries in the field of polyomavirus Agnoprotein which is expressed only by JCV, BKV, and simian virus 40 genomes.
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Nuclear Magnetic Resonance Structure of the Human Polyoma JC Virus Agnoprotein.
Journal of Cellular Biochemistry, 2017Co-Authors: Pascale Coric, A. S. Saribas, Wayne E. Childers, Magid Abou-gharbia, Martyn K. White, Mahmut Safak, Jon H. Condra, Serge BouazizAbstract:Agnoprotein is an important regulatory protein of the human polyoma JC virus (JCV) and plays critical roles during the viral replication cycle. It forms highly stable dimers and oligomers through its Leu/Ile/Phe-rich domain, which is important for the stability and function of the protein. We recently resolved the partial 3D structure of this protein by NMR using a synthetic peptide encompassing amino acids Thr17 to Gln52, where the Leu/Ile/Phe- rich region was found to adopt a major alpha-helix conformation spanning amino acids 23-39. Here, we report the resolution of the 3D structure of full-length JCV Agnoprotein by NMR, which not only confirmed the existence of the previously reported major α-helix domain at the same position but also revealed the presence of an additional minor α-helix region spanning amino acid residues Leu6 to lys13. The remaining regions of the protein adopt an intrinsically unstructured conformation. J. Cell. Biochem. 118: 3268-3280, 2017. © 2017 Wiley Periodicals, Inc.
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emerging from the unknown structural and functional features of Agnoprotein of polyomaviruses
Journal of Cellular Physiology, 2016Co-Authors: Sami A Saribas, Pascale Coric, Wayne E. Childers, Martyn K. White, Jon H. Condra, Magid Abougharbia, Anahit Hamazaspyan, William E Davis, Rachael Axman, Serge BouazizAbstract:Agnoprotein is an important regulatory protein of polyomaviruses, including JCV, BKV, and SV40. In the absence of its expression, these viruses are unable to sustain their productive life cycle. It is a highly basic phosphoprotein that localizes mostly to the perinuclear area of infected cells, although a small amount of the protein is also found in nucleus. Much has been learned about the structure and function of this important regulatory protein in recent years. It forms highly stable dimers/oligomers in vitro and in vivo through its Leu/Ile/Phe-rich domain. Structural NMR studies revealed that this domain adopts an alpha-helix conformation and plays a critical role in the stability of the protein. It associates with cellular proteins, including YB-1, p53, Ku70, FEZ1, HP1α, PP2A, AP-3, PCNA, and α-SNAP; and viral proteins, including small t antigen, large T antigen, HIV-1 Tat, and JCV VP1; and significantly contributes the viral transcription and replication. This review summarizes the recent advances in the structural and functional properties of this important regulatory protein. J. Cell. Physiol. 231: 2115-2127, 2016. © 2016 Wiley Periodicals, Inc.
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correction for coric et al nuclear magnetic resonance structure revealed that the human polyomavirus jc virus Agnoprotein contains an α helix encompassing the leu ile phe rich domain
Journal of Virology, 2014Co-Authors: Pascale Coric, A. S. Saribas, Wayne E. Childers, Serge Bouaziz, Martyn K. White, Magid Abougharbia, Mahmut SafakAbstract:Volume 88, no. 12, p. [6556–6575][1], 2014. Page 6556, abstract, line 5: “Glu55” should read “Gln52.” Page 6556, importance, line 4: “Q54” should read “Q52.” Page 6557, column 2, second full paragraph, line 3: “Glu54” should read “Gln52.” The authors regret these errors
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Nuclear Magnetic Resonance Structure Revealed that the Human Polyomavirus JC Virus Agnoprotein Contains an α-Helix Encompassing the Leu/Ile/Phe-Rich Domain
Journal of Virology, 2014Co-Authors: Pascale Coric, A. S. Saribas, Wayne E. Childers, Serge Bouaziz, Magid Abou-gharbia, Martyn K. White, Mahmut SafakAbstract:ABSTRACT Agnoprotein is a small multifunctional regulatory protein required for sustaining the productive replication of JC virus (JCV). It is a mostly cytoplasmic protein localizing in the perinuclear area and forms highly stable dimers/oligomers through a Leu/Ile/Phe-rich domain. There have been no three-dimensional structural data available for Agnoprotein due to difficulties associated with the dynamic conversion from monomers to oligomers. Here, we report the first nuclear magnetic resonance (NMR) structure of a synthetic Agnoprotein peptide spanning amino acids Thr17 to Glu55 where Lys23 to Phe39 encompassing the Leu/Ile/Phe-rich domain forms an amphipathic α-helix. On the basis of these structural data, a number of Ala substitution mutations were made to investigate the role of the α-helix in the structure and function of Agnoprotein. Single L29A and L36A mutations exhibited a significant negative effect on both protein stability and viral replication, whereas the L32A mutation did not. In addition, the L29A mutant displayed a highly nuclear localization pattern, in contrast to the pattern for the wild type (WT). Interestingly, a triple mutant, the L29A+L32A+L36A mutant, yielded no detectable Agnoprotein expression, and the replication of this JCV mutant was significantly reduced, suggesting that Leu29 and Leu36 are located at the dimer interface, contributing to the structure and stability of Agnoprotein. Two other single mutations, L33A and E34A, did not perturb Agnoprotein stability as drastically as that observed with the L29A and L36A mutations, but they negatively affected viral replication, suggesting that the role of these residues is functional rather than structural. Thus, the Agnoprotein dimerization domain can be targeted for the development of novel drugs active against JCV infection. IMPORTANCE Agnoprotein is a small regulatory protein of JC virus (JCV) and is required for the successful completion of the viral replication cycle. It forms highly stable dimers and oligomers through its hydrophobic (Leu/Ile/Phe-rich) domain, which has been shown to play essential roles in the stability and function of the protein. In this work, the Leu/Ile/Phe-rich domain has been further characterized by NMR studies using an Agnoprotein peptide spanning amino acids T17 to Q54. Those studies revealed that the dimerization domain of the protein forms an amphipathic α-helix. Subsequent NMR structure-based mutational analysis of the region highlighted the critical importance of certain amino acids within the α-helix for the stability and function of Agnoprotein. In conclusion, this study provides a solid foundation for developing effective therapeutic approaches against the dimerization domain of the protein to inhibit its critical roles in JCV infection.