Levels of Zinc Transporters mRNA Depending on Zinc Status and HIV-1 Tat Induced Inflammation in Muscle (Rhabdomyosarcoma) and Monocyte (THP-1) Cell Lines

Abstract

Monocytes and muscles demonstrate functionally contrasting behavior under conditions of zinc deficiency with relation to zinc storage system (muscle retain zinc in contrast to monocytes). We aimed to understand the effects of zinc status and HIV-1 Tat mediated inflammation on expression of zinc transporters in these types of cells. Expression of zinc transporters [ZnTs, ZIPs, and metallothionein (MT)] was quantified by qRT-PCR in RD, THP-1 cells separately and in co-cultured THP-1–RD cells. ZnT1 protein expression levels were confirmed by Western blot. Significant increase of MT and ZnT1 mRNA in response to zinc supplementation and decrease during zinc deficiency indicates significance of the genes encoding transporters in maintaining zinc homeostasis in these tissues. In the RD cells ZIP10 exhibited inverse relation to zinc status whereas no correlation was found in the THP-1 cells. Tat-induced inflammation resulted in the significant elevation of MT, IL6, ZIP7, ZIP8, ZIP9 transcripts in the co-cultured RD cells, whereas THP-1 cells demonstrated increased IL-1β levels and reduced levels of ZIP7 and ZIP14. Zinc status and HIV-1Tat induced inflammation appear to influence differential expression of MT, ZnTs, and ZIPs in the muscle and monocyte cells.

This is a preview of subscription content, access via your institution.

Fig. 1.
Fig. 2.
Fig. 3.
Fig. 4.
Fig. 5.
Fig. 6.
Fig. 7.

Abbreviations

DMEM:

Dulbecco’s modified eagle medium

FBS:

fetal bovine serum

gp120:

glycoprotein120

HIV:

Human Immunodeficiency Virus

IL:

interleukin

MT:

metallothionein

MTF:

metal transcription factor

MHC:

major histocompatibility complex

Tat:

trans-activator of transcription (Tat)

TNF:

tumour necrosis factor-α

RD:

rhabdomyosarcoma cells

TPEN:

N,N,N′,N′TetraKis (2-pyridylmethyl) ethylene diamine

ZIPs:

Zrt- and Irt-like protein

ZnTs:

zinc transporters

References

  1. 1

    Kambe, T., Tsuji, T., Hashimoto, A., and Itsumura, N. (2015) The physiological, biochemical, and molecular roles of zinc transporters in zinc homeostasis and metabolism, Physiol. Rev., 95, 749-784.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  2. 2

    Jackson, M. (1989) Physiology of Zinc: General Aspects, Zinc in Human Biology, pp. 1-14.

  3. 3

    Beck, F. W., Kaplan, J., Fine, N., Handschu, W., and Prasad, A. S. (1997) Decreased expression of CD73 (ecto-5′-nucleotidase) in the CD8+ subset is associated with zinc deficiency in human patients, J. Lab. Clin. Med., 130, 147-156.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  4. 4

    Prasad, A. S. (2008) Zinc in human health: effect of zinc on immune cells, Mol. Med., 14, 353-357.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  5. 5

    Giacconi, R., Malavolta, M., Costarelli, L., Busco, F., Galeazzi, R., et al. (2012) Comparison of intracellular zinc signals in nonadherent lymphocytes from young-adult and elderly donors: role of zinc transporters (Zip family) and proinflammatory cytokines, J. Nutr. Biochem., 23, 1256-1263.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  6. 6

    Mao, X., Kim, B. E., Wang, F., Eide, D. J., and Petris, M. J. (2007) A histidine-rich cluster mediates the ubiquitination and degradation of the human zinc transporter, hZIP4, and protects against zinc cytotoxicity, J. Biol. Chem., 282, 6992-7000.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  7. 7

    Weaver, B. P., and Andrews, G. K. (2012) Regulation of zinc-responsive Slc39a5 (Zip5) translation is mediated by conserved elements in the 3′-untranslated region, Biometals, 25, 319-335.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  8. 8

    Mocchegiani, E., Giacconi, R., Cipriano, C., and Malavolta, M. (2009) NK and NKT cells in aging and longevity: role of zinc and metallothioneins, J. Clin. Immunol., 29, 416-425.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  9. 9

    Nishida, K., Hasegawa, A., Nakae, S., Oboki, K., Saito, H., et al. (2009) Zinc transporter Znt5/Slc30a5 is required for the mast cell-mediated delayed-type allergic reaction but not the immediate-type reaction, J. Exp. Med., 206, 1351-1364.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  10. 10

    Liu, M.-J., Bao, S., Gálvez-Peralta, M., Pyle, C. J., Rudawsky, A. C., et al. (2013) The zinc transporter SLC39A8 is a negative feedback regulator of NF-κB through zinc-mediated inhibition of IKK, Cell Rep., 3, 386.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  11. 11

    Taniguchi, M., Fukunaka, A., Hagihara, M., Watanabe, K., Kamino, S., Kambe, T., Enomoto, S., and Hiromura, M. (2013) Essential role of the zinc transporter ZIP9/SLC39A9 in regulating the activations of Akt and Erk in B-cell receptor signaling pathway in DT40 cells, PLoS One, 8, e58022.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  12. 12

    Haraguchi, Y., Sakurai, H., Hussain, S., Anner, B. M., and Hoshino, H. (1999) Inhibition of HIV-1 infection by zinc group metal compounds, Antiviral Res., 43, 123-133.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  13. 13

    Mocchegiani, E., and Muzzioli, M. (2000) Therapeutic application of zinc in human immunodeficiency virus against opportunistic infections, J. Nutr., 130, 1424S-1431S.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  14. 14

    Bobat, R., Coovadia, H., Stephen, C., Naidoo, K. L., McKerrow, N., Black, R. E., and Moss, W. J. (2005) Safety and efficacy of zinc supplementation for children with HIV-1 infection in South Africa: a randomised double-blind placebo-controlled trial, Lancet, 366, 1862-1867.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  15. 15

    Baum, M. K., Lai, S., Sales, S., Page, J. B., and Campa, A. (2010) Randomized, controlled clinical trial of zinc supplementation to prevent immunological failure in HIV-infected adults, Clin. Infect. Dis., 50, 1653-1660.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  16. 16

    Tang, A. M., Graham, N. M., Kirby, A. J., McCall, L. D., Willett, W. C., and Saah, A. J. (1993) dietary micronutrient intake and risk of progression to Acquired Immunodeficiency Syndrome (AIDS) in Human Immunodeficiency Virus Type 1 (HlV-1)-infected homosexual men, Am. J. Epidemiol., 138, 937-951.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  17. 17

    Lee, S. P., and Han, M. K. (1996) Zinc stimulates Mg2+-dependent 3′-processing activity of human immunodeficiency virus type 1 integrase in vitro, Biochemistry, 35, 3837-3844.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  18. 18

    Rice, W. G., Schaeffer, C. A., Harten, B., Villinger, F., South, T. L., et al. (1993) Inhibition of HIV-1 infectivity by zinc-ejecting aromatic C-nitroso compounds, Nature, 361, 473-475.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  19. 19

    Reid, W., Sadowska, M., Denaro, F., Rao, S., Foulke, J., et al. (2001) An HIV-1 transgenic rat that develops HIV-related pathology and immunologic dysfunction, Proc. Natl. Acad. Sci. USA, 98, 9271-9276.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  20. 20

    Reid, W., Abdelwahab, S., Sadowska, M., Huso, D., Neal, A., et al. (2004) HIV-1 transgenic rats develop T cell abnormalities, Virology, 321, 111-119.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  21. 21

    Purvis, S. F., Jacobberger, J. W., Sramkoski, R. M., Patki, A. H., and Lederman, M. M. (1995) HIV type 1 Tat protein induces apoptosis and death in Jurkat cells, AIDS Res. Hum. Retroviruses, 11, 443-450.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  22. 22

    Bettaccini, A. A., Baj, A., Accolla, R. S., Basolo, F., and Toniolo, A. Q. (2005) Proliferative activity of extracellular HIV-1 Tat protein in human epithelial cells: expression profile of pathogenetically relevant genes, BMC Microbiol., 5, 20.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  23. 23

    Frankel, A. D., Bredt, D. S., and Pabo, C. O. (1988) Tat protein from human immunodeficiency virus forms a metal-linked dimer, Science, 240, 70-73.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  24. 24

    Canani, R. B., Ruotolo, S., Buccigrossi, V., Passariello, A., Porcaro, F., Siani, M. C., and Guarino, A. (2007) Zinc fights diarrhoea in HIV-1-infected children: in-vitro evidence to link clinical data and pathophysiological mechanism, AIDS, 21, 108-110.

    PubMed  Article  PubMed Central  Google Scholar 

  25. 25

    Li, J. C., Yim, H. C., and Lau, A. S. (2010) Role of HIV-1 Tat in AIDS pathogenesis: its effects on cytokine dysregulation and contributions to the pathogenesis of opportunistic infection, AIDS, 24, 1609-1623.

    PubMed  Article  PubMed Central  Google Scholar 

  26. 26

    Matsui, M., Warburton, R. J., Cogswell, P. C., Baldwin, A. S., Jr., and Frelinger, J. A. (1996) Effects of HIV-1 Tat on expression of HLA class I molecules, J. Acquir. Immune Defic. Syndr. Hum. Retrovirol., 11, 233-240.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  27. 27

    Gherardi, R. K. (1994) Skeletal muscle involvement in HIV-infected patients, Neuropathol. Appl. Neurobiol., 20, 232-237.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  28. 28

    Illa, I., Nath, A., and Dalakas, M. (1991) Immunocytochemical and virological characteristics of HIV-associated inflammatory myopathies: similarities with seronegative polymyositis, Ann. Neurol., 29, 474-481.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  29. 29

    Belec, L., Meillet, D., Hernvann, A., Gresenguet, G., and Gherardi, R. (1994) Differential elevation of circulating interleukin-1 beta, tumor necrosis factor alpha, and interleukin-6 in AIDS associated cachectic states, Clin. Diagn. Lab. Immunol., 1, 117-120.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  30. 30

    Llovera, M., Garcia-Martinez, C., Agell, N., Lopez-Soriano, F. J., Authier, F. J., Gherardi, R. K., and Argiles, J. M. (1998) Ubiquitin and proteasome gene expression is increased in skeletal muscle of slim AIDS patients, Int. J. Mol. Med., 2, 69-73.

    CAS  PubMed  PubMed Central  Google Scholar 

  31. 31

    Gonzalez-Cadavid, N. F., Taylor, W. E., Yarasheski, K., Sinha-Hikim, I., Ma, K., et al. (1998) Organization of the human myostatin gene and expression in healthy men and HIV-infected men with muscle wasting, Proc. Natl. Acad. Sci. USA, 95, 14938-14943.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  32. 32

    Tibaduiza, E. C., and Bobilya, D. J. (1996) Zinc transport across an endothelium includes vesicular cotransport with albumin, J. Cell. Physiol., 167, 539-547.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  33. 33

    Sreenivasulu, K., Raghu, P., and Nair, K. M. (2010) Polyphenol-rich beverages enhance zinc uptake and metallothionein expression in Caco-2 cells, J. Food Sci., 75, H123-H128.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  34. 34

    Coyle, P., Zalewski, P. D., Philcox, J. C., Forbes, I. J., Ward, A. D., et al. (1994) Measurement of zinc in hepatocytes by using a fluorescent probe, zinquin: relationship to metallothionein and intracellular zinc, Biochem. J., 303, 781-786.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  35. 35

    Chen, L., Frister, A., Wang, S., Ludwig, A., Behr, H., et al. (2009) Interaction of vascular smooth muscle cells and monocytes by soluble factors synergistically enhances interleukin-6 and MCP-1 production, Am. J. Physiol. Heart Circ. Physiol., 296, H987-H996.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  36. 36

    Alluri, K., Nair, K. P., Kotturu, S. K., and Ghosh, S. (2020) Transcriptional regulation of zinc transporters in human osteogenic sarcoma (Saos-2) cells to zinc supplementation and zinc depletion, Biol. Trace Elem. Res., 194, 360-367.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  37. 37

    Davis, S. R., and Cousins, R. J. (2000) Metallothionein expression in animals: a physiological perspective on function, J. Nutr., 130, 1085-1088.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  38. 38

    Alluri, K., Nair, K. P., and Ghosh, S. (2019) Differential expression of zinc transporters in functionally contrasting tissues involved in zinc homeostasis, Nucleosides Nucleotides Nucleic Acids, 18, 1-5.

    Google Scholar 

  39. 39

    Cao, J., Bobo, J. A., Liuzzi, J. P., and Cousins, R. J. (2001) Effects of intracellular zinc depletion on metallothionein and ZIP2 transporter expression and apoptosis, J. Leukoc. Biol., 70, 559-566.

    CAS  PubMed  PubMed Central  Google Scholar 

  40. 40

    Andrews, G. K. (2000) Regulation of metallothionein gene expression by oxidative stress and metal ions, Biochem. Pharmacol., 59, 95-104.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  41. 41

    Langmade, S. J., Ravindra, R., Daniels, P. J., and Andrews, G. K. (2000) The transcription factor MTF-1 mediates metal regulation of the mouse ZnT1 gene, J. Biol. Chem., 275, 34803-34809.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  42. 42

    Cousins, R. J., Blanchard, R. K., Popp, M. P., Liu, L., Cao, J., et al. (2003) A global view of the selectivity of zinc deprivation and excess on genes expressed in human THP-1 mononuclear cells, Proc. Natl. Acad. Sci. USA, 100, 6952-6957.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  43. 43

    Overbeck, S., Uciechowski, P., Ackland, M. L., Ford, D., and Rink, L. (2008) Intracellular zinc homeostasis in leukocyte subsets is regulated by different expression of zinc exporters ZnT-1 to ZnT-9, J. Leuk. Biol., 83, 368-380.

    CAS  Article  Google Scholar 

  44. 44

    Liuzzi, J. P., Blanchard, R. K., and Cousins, R. J. (2001) Differential regulation of zinc transporter 1, 2, and 4 mRNA expression by dietary zinc in rats, J. Nut., 131, 46-52.

    CAS  Article  Google Scholar 

  45. 45

    Palmiter, R. D., and Findley, S. D. (1995) Cloning and functional characterization of a mammalian zinc transporter that confers resistance to zinc, EMBO J., 14, 639-649.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  46. 46

    Hara, T., Takeda, T. A., Takagishi, T., Fukue, K., Kambe, T., and Fukada, T. (2017) Physiological roles of zinc transporters: molecular and genetic importance in zinc homeostasis, J. Physiol. Sci., 67, 283-301.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  47. 47

    Lichten, L. A., Ryu, M.-S., Guo, L., Embury, J., and Cousins, R. J. (2011) MTF-1-mediated repression of the zinc transporter Zip10 is alleviated by zinc restriction, PLoS One, 6, e21526.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  48. 48

    Ryu, M.-S., Lichten, L. A., Liuzzi, J. P., and Cousins, R. J. (2008) Zinc transporters ZnT1 (Slc30a1), Zip8 (Slc39a8), and Zip10 (Slc39a10) in mouse red blood cells are differentially regulated during erythroid development and by dietary zinc deficiency, J. Nut., 138, 2076-2083.

    CAS  Article  Google Scholar 

  49. 49

    Hamon, R., Homan, C. C., Tran, H. B., Mukaro, V. R., Lester, S. E., et al. (2014) Zinc and zinc transporters in macrophages and their roles in efferocytosis in COPD, PLoS One, 9, e110056.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  50. 50

    Chen, P., Mayne, M., Power, C., and Nath, A. (1997) The Tat protein of HIV-1 induces Tumor Necrosis Factor-α production implications for HIV-1-associated neurological diseases, J. Biol. Chem., 272, 22385-22388.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  51. 51

    Nath, A., Conant, K., Chen, P., Scott, C., and Major, E. O. (1999) Transient exposure to HIV-1 Tat protein results in cytokine production in macrophages and astrocytes A hit and run phenomenon, J. Biol. Chem., 274, 17098-17102.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  52. 52

    Hojyo, S., and Fukada, T. J. (2016) Roles of zinc signaling in the immune system, Immunol Res., 2016, 6762343, https://doi.org/10.1155/2016/6762343.

    CAS  Article  Google Scholar 

  53. 53

    Lang, C. J., Murgia, C., Leong, M., Tan, L.-W., Perozzi, G., et al. (2006) Anti-inflammatory effects of zinc and alterations in zinc transporter mRNA in mouse models of allergic inflammation, Am. J. Physiol. Lung Cell. Mol. Physiol., 292, L577-L584.

    PubMed  Article  CAS  PubMed Central  Google Scholar 

  54. 54

    Liuzzi, J. P., Lichten, L. A., Rivera, S., Blanchard, R. K., Aydemir, T. B., et al. (2005) Interleukin-6 regulates the zinc transporter Zip14 in liver and contributes to the hypozincemia of the acute-phase response, Proc. Natl. Acad. Sci. USA, 102, 6843-6848.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  55. 55

    Lichten, L. A., Liuzzi, J. P., and Cousins, R. J. (2009) Interleukin-1β contributes via nitric oxide to the upregulation and functional activity of the zinc transporter Zip14 (Slc39a14) in murine hepatocytes, Am. J. Physiol. Gastroint. Liver Physiol., 296, G860-G867.

    CAS  Article  Google Scholar 

  56. 56

    Kitamura, H., Morikawa, H., Kamon, H., Iguchi, M., Hojyo, S., et al. (2006) Toll-like receptor-mediated regulation of zinc homeostasis influences dendritic cell function, Nat. Immunol., 7, 971-977.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  57. 57

    Li, C. J., Friedman, D. J., Wang, C., Metelev, V., and Pardee, A. B. (1995) Induction of apoptosis in uninfected lymphocytes by HIV-1 Tat protein, Science, 268, 429-431.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  58. 58

    Chen, D., Wang, M., Zhou, S., and Zhou, Q. (2002) HIV-1 Tat targets microtubules to induce apoptosis, a process promoted by the pro-apoptotic Bcl-2 relative Bim, EMBO J., 21, 6801-6810.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  59. 59

    Joshi, P. C., and Guidot, D. M. (2011) HIV-1 transgene expression in rats induces differential expression of tumor necrosis factor alpha and zinc transporters in the liver and the lung, AIDS Res. Ther., 8, 36.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

Download references

Acknowledgements

We acknowledge CSIR-New Delhi, India, for financial assistance to Mr. Kiran Alluri for JRF and SRF (09/484/(0050)/2012-EMR-1). Authors gratefully acknowledge the support of Dr. B. Dinesh Kumar, Scientist G & Head-DTRC for extending the facility and chemicals/reagents from ICMR-Taskforce Project Ref. No. HIV/62/47/2016-ECD-II.

Funding

This research was financially supported by the Indian Council of Medical Research (intramural grants to KMN and grant to SG (5/9/1137/2014-NUT).

Author information

Affiliations

Authors

Corresponding authors

Correspondence to Krishna Madhavan Nair or Sudip Ghosh.

Ethics declarations

The authors declare no conflicts of interest in financial or any other sphere. This article does not contain any studies involving human participants or animals performed by any of the authors.

Electronic supplementary material

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Alluri, K., Yathapu, S.R., Babu Kondapalli, N. et al. Levels of Zinc Transporters mRNA Depending on Zinc Status and HIV-1 Tat Induced Inflammation in Muscle (Rhabdomyosarcoma) and Monocyte (THP-1) Cell Lines. Biochemistry Moscow 86, 168–178 (2021). https://doi.org/10.1134/S000629792102005X

Download citation

Keywords

  • zinc transporters
  • ZIPs
  • ZnTs
  • co-culture
  • HIV-1Tat
  • THP-1
  • RD