All Title Author
Keywords Abstract

PLOS ONE  2013 

STAT1 Regulates Human Glutaminase 1 Promoter Activity through Multiple Binding Sites in HIV-1 Infected Macrophages

DOI: 10.1371/journal.pone.0076581

Full-Text   Cite this paper   Add to My Lib


Mononuclear phagocytes (MP, macrophages and microglia), the main targets of HIV-1 infection in the brain, play a pathogenic role in HIV-associated neurocognitive disorders (HAND) through the production and release of various soluble neurotoxic factors including glutamate. We have previously reported that glutaminase (GLS), the glutamate-generating enzyme, is upregulated in HIV-1 infected MP and in the brain tissues of HIV dementia individuals, and that HIV-1 or interferon-α (IFN-α) regulates human glutaminase 1 (GLS1) promoter through signal transducer and activator of transcription 1 (STAT1) phosphorylation in macrophages. However, there are multiple putative STAT1 binding sites in human GLS1 promoter, the exact molecular mechanism of how HIV-1 or IFN-α regulates human GLS1 promoter remains unclear. To further study the function of the putative STAT1 binding sites, we mutated the sequence of each binding site to ACTAGTCTC and found that six mutants (mut 1,3,4,5,7,8) had significantly higher promoter activity and two mutants (mut 2 and mut 6) completely lost the promoter activity compared with the wild type. To determine whether sites 2 and 6 could interfere with other inhibitory sites, particularly the nearby inhibitory sites 3 and 5, we made double mutants dmut 2/3 and dmut 5/6, and found that both the double mutants had significantly higher activity than the wild type, indicating that sites 3 and 5 are critical inhibitory elements, while sites 2 and 6 are excitatory elements. ChIP assay verified that STAT1 could bind with sites 2/3 and 5/6 within human GLS1 promoter in IFN-α stimulated or HIV-1-infected monocyte-derived macrophages. Interestingly, we found that rat Gls1 promoter was regulated through a similar way as human GLS1 promoter. Together, our data identified critical elements that regulate GLS1 promoter activity.


[1]  Cui M, Huang Y, Zhao Y, Zheng J (2009) New insights for FOXO and cell-fate decision in HIV infection and HIV associated neurocognitive disorder. Adv Exp Med Biol 665: 143-159. doi:10.1007/978-1-4419-1599-3_11. PubMed: 20429422.
[2]  Ferrando SJ (2000) Diagnosis and treatment of HIV-associated neurocognitive disorders. New Dir Ment Health Serv: 25-35. PubMed: 11031798.
[3]  Lindl KA, Marks DR, Kolson DL, Jordan-Sciutto KL (2010) HIV-associated neurocognitive disorder: pathogenesis and therapeutic opportunities. J Neuroimmune Pharmacol 5: 294-309. doi:10.1007/s11481-010-9205-z. PubMed: 20396973.
[4]  Kraft-Terry SD, Buch SJ, Fox HS, Gendelman HE (2009) A coat of many colors: neuroimmune crosstalk in human immunodeficiency virus infection. Neuron 64: 133-145. doi:10.1016/j.neuron.2009.09.042. PubMed: 19840555.
[5]  Jiang ZG, Piggee C, Heyes MP, Murphy C, Quearry B et al. (2001) Glutamate is a mediator of neurotoxicity in secretions of activated HIV-1-infected macrophages. J Neuroimmunol 117: 97-107. doi:10.1016/S0165-5728(01)00315-0. PubMed: 11431009.
[6]  Lipton SA (1994) Neuronal injury associated with HIV-1 and potential treatment with calcium-channel and NMDA antagonists. Dev Neurosci 16: 145-151. doi:10.1159/000112101. PubMed: 7705221.
[7]  Zhao J, Lopez AL, Erichsen D, Herek S, Cotter RL et al. (2004) Mitochondrial glutaminase enhances extracellular glutamate production in HIV-1-infected macrophages: linkage to HIV-1 associated dementia. J Neurochem 88: 169-180. PubMed: 14675161.
[8]  Rothman SM (1985) The neurotoxicity of excitatory amino acids is produced by passive chloride influx. J Neurosci 5: 1483-1489. PubMed: 3925091.
[9]  Saransaari P, Oja SS (1998) Release of endogenous glutamate, aspartate, GABA, and taurine from hippocampal slices from adult and developing mice under cell-damaging conditions. Neurochem Res 23: 563-570. doi:10.1023/A:1022494921018. PubMed: 9566593.
[10]  Huang Y, Zhao L, Jia B, Wu L, Li Y et al. (2011) Glutaminase dysregulation in HIV-1-infected human microglia mediates neurotoxicity: relevant to HIV-1-associated neurocognitive disorders. J Neurosci 31: 15195-15204. doi:10.1523/JNEUROSCI.2051-11.2011. PubMed: 22016553.
[11]  Erdmann N, Zhao J, Lopez AL, Herek S, Curthoys N et al. (2007) Glutamate production by HIV-1 infected human macrophage is blocked by the inhibition of glutaminase. J Neurochem 102: 539-549. doi:10.1111/j.1471-4159.2007.04594.x. PubMed: 17596215.
[12]  Mock B, Kozak C, Seldin MF, Ruff N, D’Hoostelaere L et al. (1989) A glutaminase (gis) gene maps to mouse chromosome 1, rat chromosome 9, and human chromosome 2. Genomics 5: 291-297. doi:10.1016/0888-7543(89)90060-8. PubMed: 2571577.
[13]  Erdmann N, Tian C, Huang Y, Zhao J, Herek S et al. (2009) In vitro glutaminase regulation and mechanisms of glutamate generation in HIV-1-infected macrophage. J Neurochem 109: 551-561. doi:10.1111/j.1471-4159.2009.05989.x. PubMed: 19222703.
[14]  Zhao L, Huang Y, Tian C, Taylor L, Curthoys N et al. (2012) Interferon-alpha regulates glutaminase 1 promoter through STAT1 phosphorylation: relevance to HIV-1 associated neurocognitive disorders. PLOS ONE 7: e32995. doi:10.1371/journal.pone.0032995. PubMed: 22479354.
[15]  Erickson JW, Cerione RA (2010) Glutaminase: a hot spot for regulation of cancer cell metabolism? Oncotarget 1: 734-740. PubMed: 21234284.
[16]  Hu W, Zhang C, Wu R, Sun Y, Levine A et al. (2010) Glutaminase 2, a novel p53 target gene regulating energy metabolism and antioxidant function. Proc Natl Acad Sci U S A 107: 7455-7460. doi:10.1073/pnas.1001006107. PubMed: 20378837.
[17]  Márquez J, de la Oliva AR, Matés JM, Segura JA, Alonso FJ (2006) Glutaminase: a multifaceted protein not only involved in generating glutamate. Neurochem Int 48: 465-471. doi:10.1016/j.neuint.2005.10.015. PubMed: 16516349.
[18]  Andratsch M, Feifel E, Taylor L, O’Hayre M, Schramek H et al. (2007) TGF-beta signaling and its effect on glutaminase expression in LLC-PK1-FBPase+ cells. Am J Physiol Renal Physiol 293: F846-F853. doi:10.1152/ajprenal.00139.2007. PubMed: 17596530.
[19]  Taylor L, Liu X, Newsome W, Shapiro RA, Srinivasan M et al. (2001) Isolation and characterization of the promoter region of the rat kidney-type glutaminase gene. Biochim Biophys Acta 1518: 132-136. doi:10.1016/S0167-4781(01)00183-X. PubMed: 11267668.
[20]  Pérez-Gómez C, Matés JM, Gómez-Fabre PM, del Castillo-Olivares A, Alonso FJ et al. (2003) Genomic organization and transcriptional analysis of the human l-glutaminase gene. Biochem J 370: 771-784. doi:10.1042/BJ20021445. PubMed: 12444921.
[21]  Yadav A, Collman RG (2009) CNS inflammation and macrophage/microglial biology associated with HIV-1 infection. J Neuroimmune Pharmacol 4: 430-447. doi:10.1007/s11481-009-9174-2. PubMed: 19768553.
[22]  Abramovich C, Shulman LM, Ratovitski E, Harroch S, Tovey M et al. (1994) Differential tyrosine phosphorylation of the IFNAR chain of the type I interferon receptor and of an associated surface protein in response to IFN-alpha and IFN-beta. EMBO J 13: 5871-5877. PubMed: 7813427.
[23]  Levy DE, Darnell JE Jr. (2002) Stats: transcriptional control and biological impact. Nat Rev Mol Cell Biol 3: 651-662. doi:10.1038/nrg893. PubMed: 12209125.
[24]  Shuai K, Schindler C, Prezioso VR, Darnell JE Jr. (1992) Activation of transcription by IFN-gamma: tyrosine phosphorylation of a 91-kD DNA binding protein. Science 258: 1808-1812. doi:10.1126/science.1281555. PubMed: 1281555.
[25]  Huang Y, Walstrom A, Zhang L, Zhao Y, Cui M et al. (2009) Type I interferons and interferon regulatory factors regulate TNF-related apoptosis-inducing ligand (TRAIL) in HIV-1-infected macrophages. PLOS ONE 4: e5397. doi:10.1371/journal.pone.0005397. PubMed: 19404407.
[26]  Phipps T (2008) Regulation of transcription and gene expression in eukaryotes. Nature. Education 1.
[27]  Müller M, Briscoe J, Laxton C, Guschin D, Ziemiecki A et al. (1993) The protein tyrosine kinase JAK1 complements defects in interferon-alpha/beta and -gamma signal transduction. Nature 366: 129-135. doi:10.1038/366129a0. PubMed: 8232552.
[28]  Chatterjee-Kishore M, Wright KL, Ting JP, Stark GR (2000) How Stat1 mediates constitutive gene expression: a complex of unphosphorylated Stat1 and IRF1 supports transcription of the LMP2 gene. EMBO J 19: 4111-4122. doi:10.1093/emboj/19.15.4111. PubMed: 10921891.
[29]  Cheon H, Stark GR (2009) Unphosphorylated STAT1 prolongs the expression of interferon-induced immune regulatory genes. Proc Natl Acad Sci U S A 106: 9373-9378. doi:10.1073/pnas.0903487106. PubMed: 19478064.
[30]  Dekker J (2008) Gene regulation in the third dimension. Science 319: 1793-1794. doi:10.1126/science.1152850. PubMed: 18369139.
[31]  Yuasa K, Takeda S, Hijikata T (2012) A conserved regulatory element located far downstream of the GLS locus modulates GLS expression through chromatin loop formation during myogenesis. FEBS Lett 586: 3464-3470. doi:10.1016/j.febslet.2012.07.074. PubMed: 22979984.
[32]  Barber SA, Herbst DS, Bullock BT, Gama L, Clements JE (2004) Innate immune responses and control of acute simian immunodeficiency virus replication in the central nervous system. J Neurovirol 10 Suppl 1: 15-20. doi:10.1080/13550280490469590. PubMed: 14982734.
[33]  Kim DY, Hong GU, Ro JY (2011) Signal pathways in astrocytes activated by cross-talk between of astrocytes and mast cells through CD40-CD40L. J Neuroinflammation 8: 25. doi:10.1186/1742-2094-8-25. PubMed: 21410936.
[34]  Mizuno T, Zhang G, Takeuchi H, Kawanokuchi J, Wang J et al. (2008) Interferon-gamma directly induces neurotoxicity through a neuron specific, calcium-permeable complex of IFN-gamma receptor and AMPA GluR1 receptor. FASEB J 22: 1797-1806. doi:10.1096/fj.07-099499. PubMed: 18198214.
[35]  Ricciarelli R, d’Abramo C, Massone S, Marinari U, Pronzato M et al. (2004) Microarray analysis in Alzheimer’s disease and normal aging. IUBMB Life 56: 349-354. doi:10.1080/15216540412331286002. PubMed: 15370883.
[36]  Ghazawi I, Cutler SJ, Low P, Mellick AS, Ralph SJ (2005) Inhibitory effects associated with use of modified Photinus pyralis and Renilla reniformis luciferase vectors in dual reporter assays and implications for analysis of ISGs. J Interferon Cytokine Res 25: 92-102. doi:10.1089/jir.2005.25.92. PubMed: 15695930.
[37]  Gendelman HE, Orenstein JM, Martin MA, Ferrua C, Mitra R et al. (1988) Efficient isolation and propagation of human immunodeficiency virus on recombinant colony-stimulating factor 1-treated monocytes. J Exp Med 167: 1428-1441. doi:10.1084/jem.167.4.1428. PubMed: 3258626.
[38]  Constantino AA, Huang Y, Zhang H, Wood C, Zheng JC (2011) HIV-1 clade B and C isolates exhibit differential replication: relevance to macrophage-mediated neurotoxicity. Neurotox Res 20: 277-288. doi:10.1007/s12640-011-9241-3. PubMed: 21336667.


comments powered by Disqus