Skip to main content
PMC home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1989 Aug;86(15):5974–5978. doi: 10.1073/pnas.86.15.5974

Tumor necrosis factor alpha activates human immunodeficiency virus type 1 through induction of nuclear factor binding to the NF-kappa B sites in the long terminal repeat.

E J Duh 1, W J Maury 1, T M Folks 1, A S Fauci 1, A B Rabson 1
PMCID: PMC297754  PMID: 2762307

Abstract

Expression of human immunodeficiency virus type 1 (HIV-1) can be activated in a chronically infected T-cell line (ACH2 cells) by a cytokine, human tumor necrosis factor alpha (TNF-alpha). TNF-alpha treatment of ACH2 cells resulted in an increase in steady-state levels of HIV RNA and HIV transcription. Gel mobility shift assays demonstrated that the transcriptional activation of the HIV long terminal repeat (LTR) by TNF-alpha was associated with the induction of a nuclear factor(s) binding to the NF-kappa B sites in the LTR. Deletion of the NF-kappa B sites from the LTR eliminated activation by TNF-alpha in T cells transfected with plasmids in which the HIV LTR directed the expression of the bacterial chloramphenicol acetyltransferase gene. Thus, TNF-alpha appears to activate HIV RNA and virus production by ACH2 cells through the induction of transcription-activating factors that bind to the NF-kappa B sequences in the HIV LTR.

Full text

PDF
5974

Images in this article

5975Image
on p.5975
5975Image
on p.5975
5976Image
on p.5976
5976Image
on p.5976
5976Image
on p.5976

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Aggarwal B. B., Eessalu T. E., Hass P. E. Characterization of receptors for human tumour necrosis factor and their regulation by gamma-interferon. Nature. 1985 Dec 19;318(6047):665–667. doi: 10.1038/318665a0. [DOI] [PubMed] [Google Scholar]
  2. Alwine J. C., Kemp D. J., Stark G. R. Method for detection of specific RNAs in agarose gels by transfer to diazobenzyloxymethyl-paper and hybridization with DNA probes. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5350–5354. doi: 10.1073/pnas.74.12.5350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Baeuerle P. A., Baltimore D. I kappa B: a specific inhibitor of the NF-kappa B transcription factor. Science. 1988 Oct 28;242(4878):540–546. doi: 10.1126/science.3140380. [DOI] [PubMed] [Google Scholar]
  4. Baldwin A. S., Jr, Sharp P. A. Two transcription factors, NF-kappa B and H2TF1, interact with a single regulatory sequence in the class I major histocompatibility complex promoter. Proc Natl Acad Sci U S A. 1988 Feb;85(3):723–727. doi: 10.1073/pnas.85.3.723. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Ballard D. W., Böhnlein E., Lowenthal J. W., Wano Y., Franza B. R., Greene W. C. HTLV-I tax induces cellular proteins that activate the kappa B element in the IL-2 receptor alpha gene. Science. 1988 Sep 23;241(4873):1652–1655. doi: 10.1126/science.241.4873.1652. [DOI] [PubMed] [Google Scholar]
  6. Beutler B., Cerami A. Cachectin: more than a tumor necrosis factor. N Engl J Med. 1987 Feb 12;316(7):379–385. doi: 10.1056/NEJM198702123160705. [DOI] [PubMed] [Google Scholar]
  7. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  8. Chomczynski P., Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem. 1987 Apr;162(1):156–159. doi: 10.1006/abio.1987.9999. [DOI] [PubMed] [Google Scholar]
  9. Clouse K. A., Powell D., Washington I., Poli G., Strebel K., Farrar W., Barstad P., Kovacs J., Fauci A. S., Folks T. M. Monokine regulation of human immunodeficiency virus-1 expression in a chronically infected human T cell clone. J Immunol. 1989 Jan 15;142(2):431–438. [PubMed] [Google Scholar]
  10. Collins T., Lapierre L. A., Fiers W., Strominger J. L., Pober J. S. Recombinant human tumor necrosis factor increases mRNA levels and surface expression of HLA-A,B antigens in vascular endothelial cells and dermal fibroblasts in vitro. Proc Natl Acad Sci U S A. 1986 Jan;83(2):446–450. doi: 10.1073/pnas.83.2.446. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Davis M. G., Kenney S. C., Kamine J., Pagano J. S., Huang E. S. Immediate-early gene region of human cytomegalovirus trans-activates the promoter of human immunodeficiency virus. Proc Natl Acad Sci U S A. 1987 Dec;84(23):8642–8646. doi: 10.1073/pnas.84.23.8642. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Dignam J. D., Martin P. L., Shastry B. S., Roeder R. G. Eukaryotic gene transcription with purified components. Methods Enzymol. 1983;101:582–598. doi: 10.1016/0076-6879(83)01039-3. [DOI] [PubMed] [Google Scholar]
  13. Dinter H., Chiu R., Imagawa M., Karin M., Jones K. A. In vitro activation of the HIV-1 enhancer in extracts from cells treated with a phorbol ester tumor promoter. EMBO J. 1987 Dec 20;6(13):4067–4071. doi: 10.1002/j.1460-2075.1987.tb02752.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Dorsett D. L., Keshet I., Winocour E. Quantitation of a simian virus 40 nonhomologous recombination pathway. J Virol. 1983 Oct;48(1):218–228. doi: 10.1128/jvi.48.1.218-228.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Enerbäck S., Semb H., Tavernier J., Bjursell G., Olivecrona T. Tissue-specific regulation of guinea pig lipoprotein lipase; effects of nutritional state and of tumor necrosis factor on mRNA levels in adipose tissue, heart and liver. Gene. 1988 Apr 15;64(1):97–106. doi: 10.1016/0378-1119(88)90484-2. [DOI] [PubMed] [Google Scholar]
  16. Fauci A. S. The human immunodeficiency virus: infectivity and mechanisms of pathogenesis. Science. 1988 Feb 5;239(4840):617–622. doi: 10.1126/science.3277274. [DOI] [PubMed] [Google Scholar]
  17. Favaloro J., Treisman R., Kamen R. Transcription maps of polyoma virus-specific RNA: analysis by two-dimensional nuclease S1 gel mapping. Methods Enzymol. 1980;65(1):718–749. doi: 10.1016/s0076-6879(80)65070-8. [DOI] [PubMed] [Google Scholar]
  18. Folks T. M., Clouse K. A., Justement J., Rabson A., Duh E., Kehrl J. H., Fauci A. S. Tumor necrosis factor alpha induces expression of human immunodeficiency virus in a chronically infected T-cell clone. Proc Natl Acad Sci U S A. 1989 Apr;86(7):2365–2368. doi: 10.1073/pnas.86.7.2365. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Folks T. M., Justement J., Kinter A., Dinarello C. A., Fauci A. S. Cytokine-induced expression of HIV-1 in a chronically infected promonocyte cell line. Science. 1987 Nov 6;238(4828):800–802. doi: 10.1126/science.3313729. [DOI] [PubMed] [Google Scholar]
  20. Folks T., Benn S., Rabson A., Theodore T., Hoggan M. D., Martin M., Lightfoote M., Sell K. Characterization of a continuous T-cell line susceptible to the cytopathic effects of the acquired immunodeficiency syndrome (AIDS)-associated retrovirus. Proc Natl Acad Sci U S A. 1985 Jul;82(13):4539–4543. doi: 10.1073/pnas.82.13.4539. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Folks T., Powell D. M., Lightfoote M. M., Benn S., Martin M. A., Fauci A. S. Induction of HTLV-III/LAV from a nonvirus-producing T-cell line: implications for latency. Science. 1986 Feb 7;231(4738):600–602. doi: 10.1126/science.3003906. [DOI] [PubMed] [Google Scholar]
  22. Franza B. R., Jr, Josephs S. F., Gilman M. Z., Ryan W., Clarkson B. Characterization of cellular proteins recognizing the HIV enhancer using a microscale DNA-affinity precipitation assay. 1987 Nov 26-Dec 2Nature. 330(6146):391–395. doi: 10.1038/330391a0. [DOI] [PubMed] [Google Scholar]
  23. Gendelman H. E., Phelps W., Feigenbaum L., Ostrove J. M., Adachi A., Howley P. M., Khoury G., Ginsberg H. S., Martin M. A. Trans-activation of the human immunodeficiency virus long terminal repeat sequence by DNA viruses. Proc Natl Acad Sci U S A. 1986 Dec;83(24):9759–9763. doi: 10.1073/pnas.83.24.9759. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Gimble J. M., Duh E., Ostrove J. M., Gendelman H. E., Max E. E., Rabson A. B. Activation of the human immunodeficiency virus long terminal repeat by herpes simplex virus type 1 is associated with induction of a nuclear factor that binds to the NF-kappa B/core enhancer sequence. J Virol. 1988 Nov;62(11):4104–4112. doi: 10.1128/jvi.62.11.4104-4112.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Gorman C. M., Moffat L. F., Howard B. H. Recombinant genomes which express chloramphenicol acetyltransferase in mammalian cells. Mol Cell Biol. 1982 Sep;2(9):1044–1051. doi: 10.1128/mcb.2.9.1044. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Harada S., Koyanagi Y., Nakashima H., Kobayashi N., Yamamoto N. Tumor promoter, TPA, enhances replication of HTLV-III/LAV. Virology. 1986 Oct 30;154(2):249–258. doi: 10.1016/0042-6822(86)90451-4. [DOI] [PubMed] [Google Scholar]
  27. Horvat R. T., Wood C., Balachandran N. Transactivation of human immunodeficiency virus promoter by human herpesvirus 6. J Virol. 1989 Feb;63(2):970–973. doi: 10.1128/jvi.63.2.970-973.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Jeang K. T., Shank P. R., Rabson A. B., Kumar A. Synthesis of functional human immunodeficiency virus tat protein in baculovirus as determined by a cell-cell fusion assay. J Virol. 1988 Oct;62(10):3874–3878. doi: 10.1128/jvi.62.10.3874-3878.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Jones K. A., Kadonaga J. T., Luciw P. A., Tjian R. Activation of the AIDS retrovirus promoter by the cellular transcription factor, Sp1. Science. 1986 May 9;232(4751):755–759. doi: 10.1126/science.3008338. [DOI] [PubMed] [Google Scholar]
  30. Kaufman J. D., Valandra G., Roderiquez G., Bushar G., Giri C., Norcross M. A. Phorbol ester enhances human immunodeficiency virus-promoted gene expression and acts on a repeated 10-base-pair functional enhancer element. Mol Cell Biol. 1987 Oct;7(10):3759–3766. doi: 10.1128/mcb.7.10.3759. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Kawakami K., Scheidereit C., Roeder R. G. Identification and purification of a human immunoglobulin-enhancer-binding protein (NF-kappa B) that activates transcription from a human immunodeficiency virus type 1 promoter in vitro. Proc Natl Acad Sci U S A. 1988 Jul;85(13):4700–4704. doi: 10.1073/pnas.85.13.4700. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Kehrl J. H., Alvarez-Mon M., Delsing G. A., Fauci A. S. Lymphotoxin is an important T cell-derived growth factor for human B cells. Science. 1987 Nov 20;238(4830):1144–1146. doi: 10.1126/science.3500512. [DOI] [PubMed] [Google Scholar]
  33. Lenardo M. J., Kuang A., Gifford A., Baltimore D. NF-kappa B protein purification from bovine spleen: nucleotide stimulation and binding site specificity. Proc Natl Acad Sci U S A. 1988 Dec;85(23):8825–8829. doi: 10.1073/pnas.85.23.8825. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Leung K., Nabel G. J. HTLV-1 transactivator induces interleukin-2 receptor expression through an NF-kappa B-like factor. Nature. 1988 Jun 23;333(6175):776–778. doi: 10.1038/333776a0. [DOI] [PubMed] [Google Scholar]
  35. Lin J. X., Vilcek J. Tumor necrosis factor and interleukin-1 cause a rapid and transient stimulation of c-fos and c-myc mRNA levels in human fibroblasts. J Biol Chem. 1987 Sep 5;262(25):11908–11911. [PubMed] [Google Scholar]
  36. Lowenthal J. W., Ballard D. W., Böhnlein E., Greene W. C. Tumor necrosis factor alpha induces proteins that bind specifically to kappa B-like enhancer elements and regulate interleukin 2 receptor alpha-chain gene expression in primary human T lymphocytes. Proc Natl Acad Sci U S A. 1989 Apr;86(7):2331–2335. doi: 10.1073/pnas.86.7.2331. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Lusso P., Ensoli B., Markham P. D., Ablashi D. V., Salahuddin S. Z., Tschachler E., Wong-Staal F., Gallo R. C. Productive dual infection of human CD4+ T lymphocytes by HIV-1 and HHV-6. Nature. 1989 Jan 26;337(6205):370–373. doi: 10.1038/337370a0. [DOI] [PubMed] [Google Scholar]
  38. Lähdevirta J., Maury C. P., Teppo A. M., Repo H. Elevated levels of circulating cachectin/tumor necrosis factor in patients with acquired immunodeficiency syndrome. Am J Med. 1988 Sep;85(3):289–291. doi: 10.1016/0002-9343(88)90576-1. [DOI] [PubMed] [Google Scholar]
  39. Maio J. J., Brown F. L. Regulation of expression driven by human immunodeficiency virus type 1 and human T-cell leukemia virus type I long terminal repeats in pluripotential human embryonic cells. J Virol. 1988 Apr;62(4):1398–1407. doi: 10.1128/jvi.62.4.1398-1407.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Matsushima K., Morishita K., Yoshimura T., Lavu S., Kobayashi Y., Lew W., Appella E., Kung H. F., Leonard E. J., Oppenheim J. J. Molecular cloning of a human monocyte-derived neutrophil chemotactic factor (MDNCF) and the induction of MDNCF mRNA by interleukin 1 and tumor necrosis factor. J Exp Med. 1988 Jun 1;167(6):1883–1893. doi: 10.1084/jem.167.6.1883. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Matsuyama T., Hamamoto Y., Kobayashi S., Kurimoto M., Minowada J., Kobayashi N., Yamamoto N. Enhancement of human immunodeficiency virus production by natural lymphotoxin. Med Microbiol Immunol. 1988;177(4):181–187. doi: 10.1007/BF00211217. [DOI] [PubMed] [Google Scholar]
  42. McDougal J. S., Mawle A., Cort S. P., Nicholson J. K., Cross G. D., Scheppler-Campbell J. A., Hicks D., Sligh J. Cellular tropism of the human retrovirus HTLV-III/LAV. I. Role of T cell activation and expression of the T4 antigen. J Immunol. 1985 Nov;135(5):3151–3162. [PubMed] [Google Scholar]
  43. Melbye M., Biggar R. J., Ebbesen P., Neuland C., Goedert J. J., Faber V., Lorenzen I., Skinhøj P., Gallo R. C., Blattner W. A. Long-term seropositivity for human T-lymphotropic virus type III in homosexual men without the acquired immunodeficiency syndrome: development of immunologic and clinical abnormalities. A longitudinal study. Ann Intern Med. 1986 Apr;104(4):496–500. doi: 10.7326/0003-4819-104-4-496. [DOI] [PubMed] [Google Scholar]
  44. Mosca J. D., Bednarik D. P., Raj N. B., Rosen C. A., Sodroski J. G., Haseltine W. A., Pitha P. M. Herpes simplex virus type-1 can reactivate transcription of latent human immunodeficiency virus. Nature. 1987 Jan 1;325(6099):67–70. doi: 10.1038/325067a0. [DOI] [PubMed] [Google Scholar]
  45. Nabel G., Baltimore D. An inducible transcription factor activates expression of human immunodeficiency virus in T cells. Nature. 1987 Apr 16;326(6114):711–713. doi: 10.1038/326711a0. [DOI] [PubMed] [Google Scholar]
  46. Osborn L., Kunkel S., Nabel G. J. Tumor necrosis factor alpha and interleukin 1 stimulate the human immunodeficiency virus enhancer by activation of the nuclear factor kappa B. Proc Natl Acad Sci U S A. 1989 Apr;86(7):2336–2340. doi: 10.1073/pnas.86.7.2336. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Pasco D. S., Boyum K. W., Merchant S. N., Chalberg S. C., Fagan J. B. Transcriptional and post-transcriptional regulation of the genes encoding cytochromes P-450c and P-450d in vivo and in primary hepatocyte cultures. J Biol Chem. 1988 Jun 25;263(18):8671–8676. [PubMed] [Google Scholar]
  48. Quinn J. P., Holbrook N., Levens D. Binding of a cellular protein to the gibbon ape leukemia virus enhancer. Mol Cell Biol. 1987 Aug;7(8):2735–2744. doi: 10.1128/mcb.7.8.2735. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Rabson A. B., Steele P. E., Garon C. F., Martin M. A. mRNA transcripts related to full-length endogenous retroviral DNA in human cells. Nature. 1983 Dec 8;306(5943):604–607. doi: 10.1038/306604a0. [DOI] [PubMed] [Google Scholar]
  50. Rando R. F., Pellett P. E., Luciw P. A., Bohan C. A., Srinivasan A. Transactivation of human immunodeficiency virus by herpesviruses. Oncogene. 1987 Mar;1(1):13–18. [PubMed] [Google Scholar]
  51. Ray A., Tatter S. B., May L. T., Sehgal P. B. Activation of the human "beta 2-interferon/hepatocyte-stimulating factor/interleukin 6" promoter by cytokines, viruses, and second messenger agonists. Proc Natl Acad Sci U S A. 1988 Sep;85(18):6701–6705. doi: 10.1073/pnas.85.18.6701. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Ruben S., Poteat H., Tan T. H., Kawakami K., Roeder R., Haseltine W., Rosen C. A. Cellular transcription factors and regulation of IL-2 receptor gene expression by HTLV-I tax gene product. Science. 1988 Jul 1;241(4861):89–92. doi: 10.1126/science.2838905. [DOI] [PubMed] [Google Scholar]
  53. Scheurich P., Thoma B., Ucer U., Pfizenmaier K. Immunoregulatory activity of recombinant human tumor necrosis factor (TNF)-alpha: induction of TNF receptors on human T cells and TNF-alpha-mediated enhancement of T cell responses. J Immunol. 1987 Mar 15;138(6):1786–1790. [PubMed] [Google Scholar]
  54. Schütze S., Scheurich P., Schlüter C., Ucer U., Pfizenmaier K., Krönke M. Tumor necrosis factor-induced changes of gene expression in U937 cells. Differentiation-dependent plasticity of the responsive state. J Immunol. 1988 May 1;140(9):3000–3005. [PubMed] [Google Scholar]
  55. Shaw J. P., Utz P. J., Durand D. B., Toole J. J., Emmel E. A., Crabtree G. R. Identification of a putative regulator of early T cell activation genes. Science. 1988 Jul 8;241(4862):202–205. doi: 10.1126/science.3260404. [DOI] [PubMed] [Google Scholar]
  56. Sherry B., Cerami A. Cachectin/tumor necrosis factor exerts endocrine, paracrine, and autocrine control of inflammatory responses. J Cell Biol. 1988 Oct;107(4):1269–1277. doi: 10.1083/jcb.107.4.1269. [DOI] [PMC free article] [PubMed] [Google Scholar]
  57. Siekevitz M., Josephs S. F., Dukovich M., Peffer N., Wong-Staal F., Greene W. C. Activation of the HIV-1 LTR by T cell mitogens and the trans-activator protein of HTLV-I. Science. 1987 Dec 11;238(4833):1575–1578. doi: 10.1126/science.2825351. [DOI] [PubMed] [Google Scholar]
  58. Solis-Herruzo J. A., Brenner D. A., Chojkier M. Tumor necrosis factor alpha inhibits collagen gene transcription and collagen synthesis in cultured human fibroblasts. J Biol Chem. 1988 Apr 25;263(12):5841–5845. [PubMed] [Google Scholar]
  59. Tong-Starkesen S. E., Luciw P. A., Peterlin B. M. Signaling through T lymphocyte surface proteins, TCR/CD3 and CD28, activates the HIV-1 long terminal repeat. J Immunol. 1989 Jan 15;142(2):702–707. [PubMed] [Google Scholar]
  60. Tong-Starksen S. E., Luciw P. A., Peterlin B. M. Human immunodeficiency virus long terminal repeat responds to T-cell activation signals. Proc Natl Acad Sci U S A. 1987 Oct;84(19):6845–6849. doi: 10.1073/pnas.84.19.6845. [DOI] [PMC free article] [PubMed] [Google Scholar]
  61. Torti F. M., Dieckmann B., Beutler B., Cerami A., Ringold G. M. A macrophage factor inhibits adipocyte gene expression: an in vitro model of cachexia. Science. 1985 Aug 30;229(4716):867–869. doi: 10.1126/science.3839597. [DOI] [PubMed] [Google Scholar]
  62. Torti S. V., Kwak E. L., Miller S. C., Miller L. L., Ringold G. M., Myambo K. B., Young A. P., Torti F. M. The molecular cloning and characterization of murine ferritin heavy chain, a tumor necrosis factor-inducible gene. J Biol Chem. 1988 Sep 5;263(25):12638–12644. [PubMed] [Google Scholar]
  63. Vilcek J., Palombella V. J., Henriksen-DeStefano D., Swenson C., Feinman R., Hirai M., Tsujimoto M. Fibroblast growth enhancing activity of tumor necrosis factor and its relationship to other polypeptide growth factors. J Exp Med. 1986 Mar 1;163(3):632–643. doi: 10.1084/jem.163.3.632. [DOI] [PMC free article] [PubMed] [Google Scholar]
  64. Wu F. K., Garcia J. A., Harrich D., Gaynor R. B. Purification of the human immunodeficiency virus type 1 enhancer and TAR binding proteins EBP-1 and UBP-1. EMBO J. 1988 Jul;7(7):2117–2130. doi: 10.1002/j.1460-2075.1988.tb03051.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  65. Zagury D., Bernard J., Leonard R., Cheynier R., Feldman M., Sarin P. S., Gallo R. C. Long-term cultures of HTLV-III--infected T cells: a model of cytopathology of T-cell depletion in AIDS. Science. 1986 Feb 21;231(4740):850–853. doi: 10.1126/science.2418502. [DOI] [PubMed] [Google Scholar]
  66. Zechner R., Newman T. C., Sherry B., Cerami A., Breslow J. L. Recombinant human cachectin/tumor necrosis factor but not interleukin-1 alpha downregulates lipoprotein lipase gene expression at the transcriptional level in mouse 3T3-L1 adipocytes. Mol Cell Biol. 1988 Jun;8(6):2394–2401. doi: 10.1128/mcb.8.6.2394. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES