The magnesium ion, Mg 2+ , is essential for all life as a cofactor for ATP, polyphosphates such a... more The magnesium ion, Mg 2+ , is essential for all life as a cofactor for ATP, polyphosphates such as DNA and RNA, and metabolic enzymes, but whether it plays a role in intracellular signaling similar to Ca 2+ is unknown. In this study, we identify mutations in the magnesium transporter gene, MAGT1, in a novel X-linked human immunodeficiency characterized by CD4 lymphopenia, severe chronic viral infections, and defective T lymphocyte activation. We demonstrate that a rapid transient Mg 2+ influx is induced by antigen receptor stimulation in T cells or growth factor Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use:
Although Mg 2+ has a well-recognized role as an essential cofactor for all ATP-binding enzymes, i... more Although Mg 2+ has a well-recognized role as an essential cofactor for all ATP-binding enzymes, its role as a signaling ion like Ca 2+ has been controversial. A requirement for Mg 2+ for optimal T lymphocyte stimulation was demonstrated more than 30 years ago, but the mechanism of its synergistic effect with Ca 2+ in T cell activation remains elusive. Here, we summarize our recent discovery of a signaling role for Mg 2+ in the T cell antigen receptor (TCR) signaling pathway from the study of a novel primary immunodeficiency now named X-linked immunodeficiency with Mg 2+ defect, EBV infection and neoplasia (XMEN). XMEN patients were found to have a deficiency in Magnesium Transporter 1 (MAGT1), a Mg 2+-specific transporter, which leads to the absence of a TCR-stimulated Mg 2+ flux and attenuation of T cell activation. We further showed that this Mg 2+ flux is required proximally for the temporal orchestration of phospholipase C-γ1 (PLCγ1) activation. Thus, our study not only provides a second messenger role for Mg 2+ to explain its synergism with calcium in T cell signaling, it also identifies a potential extracellular therapeutic target for T cell-specific immunomodulation.
Epstein-Barr virus (EBV) is an oncogenic gammaherpesvirus that infects and persists in 95% of adu... more Epstein-Barr virus (EBV) is an oncogenic gammaherpesvirus that infects and persists in 95% of adults worldwide and has the potential to cause fatal disease, especially lymphoma, in immunocompromised hosts. Primary immunodeficiencies (PIDs) that predispose to EBV-associated malignancies have provided novel insights into the molecular mechanisms of immune defense against EBV. We have recently characterized a novel PID now named "Xlinked immunodeficiency with magnesium defect, EBV infection, and neoplasia" (XMEN) disease characterized by lossof-function mutations in the gene encoding magnesium transporter 1 (MAGT1), chronic high-level EBV with increased EBV-infected B cells, and heightened susceptibility to EBV-associated lymphomas. The genetic etiology of XMEN disease has revealed an unexpected quantitative role for intracellular free magnesium in immune functions and has led to novel diagnostic and therapeutic strategies. Here, we review the clinical presentation, genetic mutation spectrum, molecular mechanisms of pathogenesis, and diagnostic and therapeutic considerations for this previously unrecognized disease. (Blood.
The magnesium transporter 1 (MAGT1) is a critical regulator of basal intracellular free Mg 2+ lev... more The magnesium transporter 1 (MAGT1) is a critical regulator of basal intracellular free Mg 2+ levels. Individuals with genetic deficiencies in MAGT1 have high levels of Epstein-Barr virus (EBV) and a predisposition to lymphoma. We show that decreased intracellular free Mg 2+ causes defective expression of the natural killer activating receptor NKG2D in natural killer (NK) and CD8 + T cells and impairs cytolytic responses against EBV. Remarkably, magnesium supplementation in MAGT1-deficient patients restores intracellular free Mg 2+ and NKG2D while concurrently reducing EBV-infected cells in vivo, demonstrating a link between NKG2D cytolytic activity and EBV antiviral immunity in humans. Moreover, these findings reveal a specific molecular function of free basal intracellular Mg 2+ in eukaryotic cells. Divalent metal cations, including Mg 2+ , play important roles in cellular processes. In eukaryotic cells, 95% of intracellular Mg 2+ ([Mg 2+ ] i) is bound (1). The remaining unbound, free [Mg 2+ ] i (~0.5-1 mM) is tightly regulated, yet its specific molecular functions are unknown (1-3). Magnesium transporter 1 (MAGT1) selectively conducts Mg 2+ across the plasma membrane (4, 5). We previously characterized a novel primary immunodeficiency,
Death receptor signaling is initiated by the assembly of the deathinducing signaling complex, whi... more Death receptor signaling is initiated by the assembly of the deathinducing signaling complex, which culminates in the activation of the initiator caspase, either caspase-8 or caspase-10. A family of viral and cellular proteins, known as FLIP, plays an essential role in the regulation of death receptor signaling. Viral FLIP (v-FLIP) and short cellular FLIP (c-FLIP S) inhibit apoptosis by interfering with death receptor signaling. The structure and mechanisms of v-FLIP and c-FLIP S remain largely unknown. Here we report a high resolution crystal structure of MC159, a v-FLIP derived from the molluscum contagiosum virus, which is a member of the human poxvirus family. Unexpectedly, the two tandem death effector domains (DEDs) of MC159 rigidly associate with each other through a hydrophobic interface. Structure-based sequence analysis suggests that this interface is conserved in the tandem DEDs from other v-FLIP, c-FLIP S , and caspase-8 and-10. Strikingly, the overall packing arrangement between the two DEDs of MC159 resembles that between the caspase recruitment domains of Apaf-1 and caspase-9. In addition, each DED of MC159 contains a highly conserved binding motif on the surface, to which loss-of-function mutations in MC159 map. These observations, in conjunction with published evidence, reveal significant insights into the function of v-FLIP and suggest a mechanism by which v-FLIP and c-FLIP S inhibit death receptor signaling.
The magnesium ion, Mg2+, is essential for all life as a cofactor for ATP, polyphosphates such as ... more The magnesium ion, Mg2+, is essential for all life as a cofactor for ATP, polyphosphates such as DNA and RNA, and metabolic enzymes, but whether it plays a role in intracellular signaling similar to Ca2+ is unknown. In this study, we identify mutations in the magnesium transporter gene, MAGT1, in a novel X-linked human immunodeficiency characterized by CD4 lymphopenia, severe chronic viral infections, and defective T lymphocyte activation. We demonstrate that a rapid transient Mg2+ influx is induced by antigen receptor stimulation in T cells or growth factor Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use:http://www.nature.com/authors/editorial_policies/license.html#terms Correspondence and requests for materials should be addressed to M.J.L. ([email protected]).†These authors contributed equally Author Contributions F-Y.L. characterized the MAGT1 mutations a...
Author(s): Li, Feng-Yen | Advisor(s): Lenardo, Michael J | Abstract: The etiologies of human prim... more Author(s): Li, Feng-Yen | Advisor(s): Lenardo, Michael J | Abstract: The etiologies of human primary immunodeficiencies (PIDs) often yield novel insights about the immune system. This thesis revealed an unexpected regulatory role for magnesium in the immune system from the characterization of a novel PID now named X-linked immunodeficiency with magnesium defect, Epstein Barr Virus (EBV) infection and neoplasia (XMEN) disease. This story stemmed from the characterization of two young brothers of a non-consanguineous family exhibiting recurrent viral infections and decreased thymic output of CD4+ T cells. The mother of these two boys was found to have completely skewed lyonization in her T cells, suggesting that she carried a defective X-linked gene that confers decreased fitness relative to the wildtype allele. Using X-chromosome exon capture next-generation sequencing, a 10 base pair deletion ablating a splicing junction of Magnesium Transporter 1 (MAGT1) was found by identifying ge...
Magnesium research : official organ of the International Society for the Development of Research on Magnesium, 2011
Although Mg(2+) has a well-recognized role as an essential cofactor for all ATP-binding enzymes, ... more Although Mg(2+) has a well-recognized role as an essential cofactor for all ATP-binding enzymes, its role as a signaling ion, like Ca(2+), has been controversial. A requirement for Mg(2+)for optimal T lymphocyte stimulation was demonstrated more than 30 years ago, but the mechanism of its synergistic effect with Ca(2+)in T cell activation remains elusive. Here, we summarize our recent discovery of a signaling role for Mg(2+)in the T cell antigen receptor (TCR) signaling pathway from the study of a novel primary immunodeficiency, now named X-linked immunodeficiency with Mg(2+)defect, EBV infection and neoplasia (XMEN). XMEN patients were found to have a deficiency in magnesium transporter 1 (MAGT1), an Mg(2+)-specific transporter, which leads to the absence of a TCR-stimulated Mg(2+)flux and an attenuation of T cell activation. We further showed that this Mg(2+)flux is required proximally for the temporal orchestration of phospholipase C-γ1 (PLCγ1) activation. Thus, our study not onl...
Histone N-acetyltransferases (HATs) are a group of enzymes which acetylate specific lysine residu... more Histone N-acetyltransferases (HATs) are a group of enzymes which acetylate specific lysine residues in the N-terminal tails of nucleosomal histones to promote transcriptional activation. Recent structural and enzymatic work on the GCN5/PCAF HAT family has elucidated the structure of their catalytic domain and mechanism of histone acetylation. However, the substrate specificity of these enzymes has not been quantitatively investigated. Utilizing a novel microplate fluorescent HAT assay which detects the enzymatic production of coenzyme A (CoA), we have compared the activities of the HAT domains of human PCAF and its GCN5 homologue from yeast and Tetrahymena and found that they have similar kinetic parameters. PCAF was further assayed with a series of different length histone H3 peptide substrates, which revealed that the determinants for substrate recognition lie within a 19-residue sequence. Finally, we evaluated the acetylation of three putative PCAF substrates, histones H3 and H4 and the transcription factor p53, and have determined that histone H3 is significantly preferred over the histone H4 and p53 substrates. Taken together, the fluorescent acetyltransferase assay presented here should be widely applicable to other HAT enzymes, and the results obtained with PCAF demonstrate a strong substrate preference for the N-terminal residues of histone H3.
Epstein-Barr virus (EBV) is an oncogenic gammaherpesvirus that infects and persists in 95% of adu... more Epstein-Barr virus (EBV) is an oncogenic gammaherpesvirus that infects and persists in 95% of adults worldwide and has the potential to cause fatal disease, especially lymphoma, in immunocompromised hosts. Primary immunodeficiencies (PIDs) that predispose to EBV-associated malignancies have provided novel insights into the molecular mechanisms of immune defense against EBV. We have recently characterized a novel PID now named “X-linked immunodeficiency with magnesium defect, EBV infection, and neoplasia” (XMEN) disease characterized by loss-of-function mutations in the gene encoding magnesium transporter 1 (MAGT1), chronic high-level EBV with increased EBV-infected B cells, and heightened susceptibility to EBV-associated lymphomas. The genetic etiology of XMEN disease has revealed an unexpected quantitative role for intracellular free magnesium in immune functions and has led to novel diagnostic and therapeutic strategies. Here, we review the clinical presentation, genetic mutation ...
In this study we have investigated the impact of differentiation of neuronal cells on their sensi... more In this study we have investigated the impact of differentiation of neuronal cells on their sensitivity to microbial toxins. We used the human neural crest-derived tumor cell line Paju, which can be induced to differentiation in vitro by treatment with phorbol 12-myristate 13-acetate. Addition of the highly toxic potassium ionophores cereulide (4.5 and 9.0 ng/ml) or valinomycin (20 ng/ml), to cultures of undifferentiated Paju cells caused collapse of the mitochondrial membrane potential - measured with the fluorescent probe 5,5',6,6'-tetrachloro-1,1',3,3'-tetrabenzimidazole carbocyanine iodide (JC-1) followed by detachment of the cells and their apoptotic death. After induced differentiation of the Paju cells, their mitochondria retained the membrane potential upon exposure to the toxins and the cells displayed increased resistance to apoptosis as compared with undifferentiated cells. This effect may be caused by an elevated expression of the anti-apoptotic protein B...
, 186 (2013); 341 Science et al. Benjamin Chaigne-Delalande Chronic EBV Infection Through NKG2D R... more , 186 (2013); 341 Science et al. Benjamin Chaigne-Delalande Chronic EBV Infection Through NKG2D Regulates Cytotoxic Functions of NK and CD8 T Cells in 2+ Mg This copy is for your personal, non-commercial use only. clicking here. colleagues, clients, or customers by , you can order high-quality copies for your If you wish to distribute this article to others here. following the guidelines can be obtained by Permission to republish or repurpose articles or portions of articles ): December 3, 2013 www.sciencemag.org (this information is current as of The following resources related to this article are available online at http://www.sciencemag.org/content/341/6142/186.full.html version of this article at: including high-resolution figures, can be found in the online Updated information and services, http://www.sciencemag.org/content/suppl/2013/07/10/341.6142.186.DC1.html can be found at: Supporting Online Material http://www.sciencemag.org/content/341/6142/186.full.html#related found at: can be related to this article A list of selected additional articles on the Science Web sites http://www.sciencemag.org/content/341/6142/186.full.html#ref-list-1 , 12 of which can be accessed free: cites 44 articles This article http://www.sciencemag.org/cgi/collection/immunology Immunology subject collections: This article appears in the following
The magnesium transporter 1 (MAGT1) is a critical regulator of basal intracellular free magnesium... more The magnesium transporter 1 (MAGT1) is a critical regulator of basal intracellular free magnesium (Mg(2+)) concentrations. Individuals with genetic deficiencies in MAGT1 have high levels of Epstein-Barr virus (EBV) and a predisposition to lymphoma. We show that decreased intracellular free Mg(2+) causes defective expression of the natural killer activating receptor NKG2D in natural killer (NK) and CD8(+) T cells and impairs cytolytic responses against EBV. Notably, magnesium supplementation in MAGT1-deficient patients restores intracellular free Mg(2+) and NKG2D while concurrently reducing EBV-infected cells in vivo, demonstrating a link between NKG2D cytolytic activity and EBV antiviral immunity in humans. Moreover, these findings reveal a specific molecular function of free basal intracellular Mg(2+) in eukaryotic cells.
Death receptor signaling is initiated by the assembly of the deathinducing signaling complex, whi... more Death receptor signaling is initiated by the assembly of the deathinducing signaling complex, which culminates in the activation of the initiator caspase, either caspase-8 or caspase-10. A family of viral and cellular proteins, known as FLIP, plays an essential role in the regulation of death receptor signaling. Viral FLIP (v-FLIP) and short cellular FLIP (c-FLIP S) inhibit apoptosis by interfering with death receptor signaling. The structure and mechanisms of v-FLIP and c-FLIP S remain largely unknown. Here we report a high resolution crystal structure of MC159, a v-FLIP derived from the molluscum contagiosum virus, which is a member of the human poxvirus family. Unexpectedly, the two tandem death effector domains (DEDs) of MC159 rigidly associate with each other through a hydrophobic interface. Structure-based sequence analysis suggests that this interface is conserved in the tandem DEDs from other v-FLIP, c-FLIP S , and caspase-8 and-10. Strikingly, the overall packing arrangement between the two DEDs of MC159 resembles that between the caspase recruitment domains of Apaf-1 and caspase-9. In addition, each DED of MC159 contains a highly conserved binding motif on the surface, to which loss-of-function mutations in MC159 map. These observations, in conjunction with published evidence, reveal significant insights into the function of v-FLIP and suggest a mechanism by which v-FLIP and c-FLIP S inhibit death receptor signaling.
The magnesium ion, Mg 2+ , is essential for all life as a cofactor for ATP, polyphosphates such a... more The magnesium ion, Mg 2+ , is essential for all life as a cofactor for ATP, polyphosphates such as DNA and RNA, and metabolic enzymes, but whether it plays a role in intracellular signaling similar to Ca 2+ is unknown. In this study, we identify mutations in the magnesium transporter gene, MAGT1, in a novel X-linked human immunodeficiency characterized by CD4 lymphopenia, severe chronic viral infections, and defective T lymphocyte activation. We demonstrate that a rapid transient Mg 2+ influx is induced by antigen receptor stimulation in T cells or growth factor Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use:
Although Mg 2+ has a well-recognized role as an essential cofactor for all ATP-binding enzymes, i... more Although Mg 2+ has a well-recognized role as an essential cofactor for all ATP-binding enzymes, its role as a signaling ion like Ca 2+ has been controversial. A requirement for Mg 2+ for optimal T lymphocyte stimulation was demonstrated more than 30 years ago, but the mechanism of its synergistic effect with Ca 2+ in T cell activation remains elusive. Here, we summarize our recent discovery of a signaling role for Mg 2+ in the T cell antigen receptor (TCR) signaling pathway from the study of a novel primary immunodeficiency now named X-linked immunodeficiency with Mg 2+ defect, EBV infection and neoplasia (XMEN). XMEN patients were found to have a deficiency in Magnesium Transporter 1 (MAGT1), a Mg 2+-specific transporter, which leads to the absence of a TCR-stimulated Mg 2+ flux and attenuation of T cell activation. We further showed that this Mg 2+ flux is required proximally for the temporal orchestration of phospholipase C-γ1 (PLCγ1) activation. Thus, our study not only provides a second messenger role for Mg 2+ to explain its synergism with calcium in T cell signaling, it also identifies a potential extracellular therapeutic target for T cell-specific immunomodulation.
Epstein-Barr virus (EBV) is an oncogenic gammaherpesvirus that infects and persists in 95% of adu... more Epstein-Barr virus (EBV) is an oncogenic gammaherpesvirus that infects and persists in 95% of adults worldwide and has the potential to cause fatal disease, especially lymphoma, in immunocompromised hosts. Primary immunodeficiencies (PIDs) that predispose to EBV-associated malignancies have provided novel insights into the molecular mechanisms of immune defense against EBV. We have recently characterized a novel PID now named "Xlinked immunodeficiency with magnesium defect, EBV infection, and neoplasia" (XMEN) disease characterized by lossof-function mutations in the gene encoding magnesium transporter 1 (MAGT1), chronic high-level EBV with increased EBV-infected B cells, and heightened susceptibility to EBV-associated lymphomas. The genetic etiology of XMEN disease has revealed an unexpected quantitative role for intracellular free magnesium in immune functions and has led to novel diagnostic and therapeutic strategies. Here, we review the clinical presentation, genetic mutation spectrum, molecular mechanisms of pathogenesis, and diagnostic and therapeutic considerations for this previously unrecognized disease. (Blood.
The magnesium transporter 1 (MAGT1) is a critical regulator of basal intracellular free Mg 2+ lev... more The magnesium transporter 1 (MAGT1) is a critical regulator of basal intracellular free Mg 2+ levels. Individuals with genetic deficiencies in MAGT1 have high levels of Epstein-Barr virus (EBV) and a predisposition to lymphoma. We show that decreased intracellular free Mg 2+ causes defective expression of the natural killer activating receptor NKG2D in natural killer (NK) and CD8 + T cells and impairs cytolytic responses against EBV. Remarkably, magnesium supplementation in MAGT1-deficient patients restores intracellular free Mg 2+ and NKG2D while concurrently reducing EBV-infected cells in vivo, demonstrating a link between NKG2D cytolytic activity and EBV antiviral immunity in humans. Moreover, these findings reveal a specific molecular function of free basal intracellular Mg 2+ in eukaryotic cells. Divalent metal cations, including Mg 2+ , play important roles in cellular processes. In eukaryotic cells, 95% of intracellular Mg 2+ ([Mg 2+ ] i) is bound (1). The remaining unbound, free [Mg 2+ ] i (~0.5-1 mM) is tightly regulated, yet its specific molecular functions are unknown (1-3). Magnesium transporter 1 (MAGT1) selectively conducts Mg 2+ across the plasma membrane (4, 5). We previously characterized a novel primary immunodeficiency,
Death receptor signaling is initiated by the assembly of the deathinducing signaling complex, whi... more Death receptor signaling is initiated by the assembly of the deathinducing signaling complex, which culminates in the activation of the initiator caspase, either caspase-8 or caspase-10. A family of viral and cellular proteins, known as FLIP, plays an essential role in the regulation of death receptor signaling. Viral FLIP (v-FLIP) and short cellular FLIP (c-FLIP S) inhibit apoptosis by interfering with death receptor signaling. The structure and mechanisms of v-FLIP and c-FLIP S remain largely unknown. Here we report a high resolution crystal structure of MC159, a v-FLIP derived from the molluscum contagiosum virus, which is a member of the human poxvirus family. Unexpectedly, the two tandem death effector domains (DEDs) of MC159 rigidly associate with each other through a hydrophobic interface. Structure-based sequence analysis suggests that this interface is conserved in the tandem DEDs from other v-FLIP, c-FLIP S , and caspase-8 and-10. Strikingly, the overall packing arrangement between the two DEDs of MC159 resembles that between the caspase recruitment domains of Apaf-1 and caspase-9. In addition, each DED of MC159 contains a highly conserved binding motif on the surface, to which loss-of-function mutations in MC159 map. These observations, in conjunction with published evidence, reveal significant insights into the function of v-FLIP and suggest a mechanism by which v-FLIP and c-FLIP S inhibit death receptor signaling.
The magnesium ion, Mg2+, is essential for all life as a cofactor for ATP, polyphosphates such as ... more The magnesium ion, Mg2+, is essential for all life as a cofactor for ATP, polyphosphates such as DNA and RNA, and metabolic enzymes, but whether it plays a role in intracellular signaling similar to Ca2+ is unknown. In this study, we identify mutations in the magnesium transporter gene, MAGT1, in a novel X-linked human immunodeficiency characterized by CD4 lymphopenia, severe chronic viral infections, and defective T lymphocyte activation. We demonstrate that a rapid transient Mg2+ influx is induced by antigen receptor stimulation in T cells or growth factor Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use:http://www.nature.com/authors/editorial_policies/license.html#terms Correspondence and requests for materials should be addressed to M.J.L. ([email protected]).†These authors contributed equally Author Contributions F-Y.L. characterized the MAGT1 mutations a...
Author(s): Li, Feng-Yen | Advisor(s): Lenardo, Michael J | Abstract: The etiologies of human prim... more Author(s): Li, Feng-Yen | Advisor(s): Lenardo, Michael J | Abstract: The etiologies of human primary immunodeficiencies (PIDs) often yield novel insights about the immune system. This thesis revealed an unexpected regulatory role for magnesium in the immune system from the characterization of a novel PID now named X-linked immunodeficiency with magnesium defect, Epstein Barr Virus (EBV) infection and neoplasia (XMEN) disease. This story stemmed from the characterization of two young brothers of a non-consanguineous family exhibiting recurrent viral infections and decreased thymic output of CD4+ T cells. The mother of these two boys was found to have completely skewed lyonization in her T cells, suggesting that she carried a defective X-linked gene that confers decreased fitness relative to the wildtype allele. Using X-chromosome exon capture next-generation sequencing, a 10 base pair deletion ablating a splicing junction of Magnesium Transporter 1 (MAGT1) was found by identifying ge...
Magnesium research : official organ of the International Society for the Development of Research on Magnesium, 2011
Although Mg(2+) has a well-recognized role as an essential cofactor for all ATP-binding enzymes, ... more Although Mg(2+) has a well-recognized role as an essential cofactor for all ATP-binding enzymes, its role as a signaling ion, like Ca(2+), has been controversial. A requirement for Mg(2+)for optimal T lymphocyte stimulation was demonstrated more than 30 years ago, but the mechanism of its synergistic effect with Ca(2+)in T cell activation remains elusive. Here, we summarize our recent discovery of a signaling role for Mg(2+)in the T cell antigen receptor (TCR) signaling pathway from the study of a novel primary immunodeficiency, now named X-linked immunodeficiency with Mg(2+)defect, EBV infection and neoplasia (XMEN). XMEN patients were found to have a deficiency in magnesium transporter 1 (MAGT1), an Mg(2+)-specific transporter, which leads to the absence of a TCR-stimulated Mg(2+)flux and an attenuation of T cell activation. We further showed that this Mg(2+)flux is required proximally for the temporal orchestration of phospholipase C-γ1 (PLCγ1) activation. Thus, our study not onl...
Histone N-acetyltransferases (HATs) are a group of enzymes which acetylate specific lysine residu... more Histone N-acetyltransferases (HATs) are a group of enzymes which acetylate specific lysine residues in the N-terminal tails of nucleosomal histones to promote transcriptional activation. Recent structural and enzymatic work on the GCN5/PCAF HAT family has elucidated the structure of their catalytic domain and mechanism of histone acetylation. However, the substrate specificity of these enzymes has not been quantitatively investigated. Utilizing a novel microplate fluorescent HAT assay which detects the enzymatic production of coenzyme A (CoA), we have compared the activities of the HAT domains of human PCAF and its GCN5 homologue from yeast and Tetrahymena and found that they have similar kinetic parameters. PCAF was further assayed with a series of different length histone H3 peptide substrates, which revealed that the determinants for substrate recognition lie within a 19-residue sequence. Finally, we evaluated the acetylation of three putative PCAF substrates, histones H3 and H4 and the transcription factor p53, and have determined that histone H3 is significantly preferred over the histone H4 and p53 substrates. Taken together, the fluorescent acetyltransferase assay presented here should be widely applicable to other HAT enzymes, and the results obtained with PCAF demonstrate a strong substrate preference for the N-terminal residues of histone H3.
Epstein-Barr virus (EBV) is an oncogenic gammaherpesvirus that infects and persists in 95% of adu... more Epstein-Barr virus (EBV) is an oncogenic gammaherpesvirus that infects and persists in 95% of adults worldwide and has the potential to cause fatal disease, especially lymphoma, in immunocompromised hosts. Primary immunodeficiencies (PIDs) that predispose to EBV-associated malignancies have provided novel insights into the molecular mechanisms of immune defense against EBV. We have recently characterized a novel PID now named “X-linked immunodeficiency with magnesium defect, EBV infection, and neoplasia” (XMEN) disease characterized by loss-of-function mutations in the gene encoding magnesium transporter 1 (MAGT1), chronic high-level EBV with increased EBV-infected B cells, and heightened susceptibility to EBV-associated lymphomas. The genetic etiology of XMEN disease has revealed an unexpected quantitative role for intracellular free magnesium in immune functions and has led to novel diagnostic and therapeutic strategies. Here, we review the clinical presentation, genetic mutation ...
In this study we have investigated the impact of differentiation of neuronal cells on their sensi... more In this study we have investigated the impact of differentiation of neuronal cells on their sensitivity to microbial toxins. We used the human neural crest-derived tumor cell line Paju, which can be induced to differentiation in vitro by treatment with phorbol 12-myristate 13-acetate. Addition of the highly toxic potassium ionophores cereulide (4.5 and 9.0 ng/ml) or valinomycin (20 ng/ml), to cultures of undifferentiated Paju cells caused collapse of the mitochondrial membrane potential - measured with the fluorescent probe 5,5',6,6'-tetrachloro-1,1',3,3'-tetrabenzimidazole carbocyanine iodide (JC-1) followed by detachment of the cells and their apoptotic death. After induced differentiation of the Paju cells, their mitochondria retained the membrane potential upon exposure to the toxins and the cells displayed increased resistance to apoptosis as compared with undifferentiated cells. This effect may be caused by an elevated expression of the anti-apoptotic protein B...
, 186 (2013); 341 Science et al. Benjamin Chaigne-Delalande Chronic EBV Infection Through NKG2D R... more , 186 (2013); 341 Science et al. Benjamin Chaigne-Delalande Chronic EBV Infection Through NKG2D Regulates Cytotoxic Functions of NK and CD8 T Cells in 2+ Mg This copy is for your personal, non-commercial use only. clicking here. colleagues, clients, or customers by , you can order high-quality copies for your If you wish to distribute this article to others here. following the guidelines can be obtained by Permission to republish or repurpose articles or portions of articles ): December 3, 2013 www.sciencemag.org (this information is current as of The following resources related to this article are available online at http://www.sciencemag.org/content/341/6142/186.full.html version of this article at: including high-resolution figures, can be found in the online Updated information and services, http://www.sciencemag.org/content/suppl/2013/07/10/341.6142.186.DC1.html can be found at: Supporting Online Material http://www.sciencemag.org/content/341/6142/186.full.html#related found at: can be related to this article A list of selected additional articles on the Science Web sites http://www.sciencemag.org/content/341/6142/186.full.html#ref-list-1 , 12 of which can be accessed free: cites 44 articles This article http://www.sciencemag.org/cgi/collection/immunology Immunology subject collections: This article appears in the following
The magnesium transporter 1 (MAGT1) is a critical regulator of basal intracellular free magnesium... more The magnesium transporter 1 (MAGT1) is a critical regulator of basal intracellular free magnesium (Mg(2+)) concentrations. Individuals with genetic deficiencies in MAGT1 have high levels of Epstein-Barr virus (EBV) and a predisposition to lymphoma. We show that decreased intracellular free Mg(2+) causes defective expression of the natural killer activating receptor NKG2D in natural killer (NK) and CD8(+) T cells and impairs cytolytic responses against EBV. Notably, magnesium supplementation in MAGT1-deficient patients restores intracellular free Mg(2+) and NKG2D while concurrently reducing EBV-infected cells in vivo, demonstrating a link between NKG2D cytolytic activity and EBV antiviral immunity in humans. Moreover, these findings reveal a specific molecular function of free basal intracellular Mg(2+) in eukaryotic cells.
Death receptor signaling is initiated by the assembly of the deathinducing signaling complex, whi... more Death receptor signaling is initiated by the assembly of the deathinducing signaling complex, which culminates in the activation of the initiator caspase, either caspase-8 or caspase-10. A family of viral and cellular proteins, known as FLIP, plays an essential role in the regulation of death receptor signaling. Viral FLIP (v-FLIP) and short cellular FLIP (c-FLIP S) inhibit apoptosis by interfering with death receptor signaling. The structure and mechanisms of v-FLIP and c-FLIP S remain largely unknown. Here we report a high resolution crystal structure of MC159, a v-FLIP derived from the molluscum contagiosum virus, which is a member of the human poxvirus family. Unexpectedly, the two tandem death effector domains (DEDs) of MC159 rigidly associate with each other through a hydrophobic interface. Structure-based sequence analysis suggests that this interface is conserved in the tandem DEDs from other v-FLIP, c-FLIP S , and caspase-8 and-10. Strikingly, the overall packing arrangement between the two DEDs of MC159 resembles that between the caspase recruitment domains of Apaf-1 and caspase-9. In addition, each DED of MC159 contains a highly conserved binding motif on the surface, to which loss-of-function mutations in MC159 map. These observations, in conjunction with published evidence, reveal significant insights into the function of v-FLIP and suggest a mechanism by which v-FLIP and c-FLIP S inhibit death receptor signaling.
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