Molecular Methods to Analyze the Effect of Proteins Expressed by Salmonella During Its Intracellular Stage

  • Carlos MedinaEmail author
  • Beatriz Mesa-Pereira
  • Eva M. Camacho
  • Amando Flores
  • Eduardo Santero
Part of the Methods in Molecular Biology book series (MIMB, volume 1734)


Salmonella is probably the intracellular pathogen most extensively studied. Once inside the cell, this bacterium produces different proteins involved in the infection process known as effectors that translocate through its own secretion systems to the eukaryotic cytosol exerting diverse effects on the cell. Additionally, Salmonella can be engineered to include a protein expression system that, upon the addition of an inducer molecule, can produce heterologous proteins at a specific time during the course of the infection. The effect of such proteins on the eukaryotic (i.e., tumoral) cells can be detected following distinct approaches, which converts Salmonella in an effective tool to produce proteins inside eukaryotic cells with different purposes, such as killing tumoral cells. Here, we present diverse technics currently used to produce proteins by Salmonella inside tumoral cells and analyze its cytotoxic effect.

Key words

Salmonella Protein expression Therapeutic proteins Antitumoral drugs Bacterial lysis Salicylate Drug delivery 



We are grateful to all members of the laboratory for their insights and helpful suggestions, and Guadalupe Martín Cabello for technical help. This work was supported by the Grant ‘Proyecto de Excelencia P07-CVI02518’ from the Andalusian government and by Spanish Ministry of Science and Innovation grants BIO2014-57545-R.


  1. 1.
    Pawelek JM, Low KB, Bermudes D (2003) Bacteria as tumour-targeting vectors. Lancet Oncol 4:548–556CrossRefPubMedGoogle Scholar
  2. 2.
    Malmgren RA, Flanigan CC (1955) Localization of the vegetative form of Clostridium tetani in mouse tumors following intravenous spore administration. Cancer Res 15:473–478PubMedGoogle Scholar
  3. 3.
    Pawelek JM, Low KB, Bermudes D (1997) Tumor-targeted Salmonella as a novel anticancer vector. Cancer Res 57:4537–4544PubMedGoogle Scholar
  4. 4.
    Kasinskas RW, Forbes NS (2007) Salmonella Typhimurium lacking ribose chemoreceptors localize in tumor quiescence and induce apoptosis. Cancer Res 67:3201–3209CrossRefPubMedGoogle Scholar
  5. 5.
    Zheng JH, Min JJ (2016) Targeted cancer therapy using engineered Salmonella Typhimurium. Chonnam Med J 52:173–184CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Wang CZ, Kazmierczak RA, Eisenstark A (2016) Strains, mechanism, and perspective: Salmonella-Based Cancer therapy. Int J Microbiol 2016:5678702CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Hoffman RM (2016) Future of bacterial therapy of cancer. Methods Mol Biol 1409:177–184CrossRefPubMedGoogle Scholar
  8. 8.
    Pinero-Lambea C, Ruano-Gallego D, Fernandez LA (2015) Engineered bacteria as therapeutic agents. Curr Opin Biotech 35:94–102CrossRefPubMedGoogle Scholar
  9. 9.
    Zhang M, Forbes NS (2015) Trg-deficient Salmonella colonize quiescent tumor regions by exclusively penetrating or proliferating. J Control Release 199:180–189CrossRefPubMedGoogle Scholar
  10. 10.
    Wong S, Slavcev RA (2015) Treating cancer with infection: a review on bacterial cancer therapy. Lett Appl Microbiol 61:107–112CrossRefPubMedGoogle Scholar
  11. 11.
    Kim JE, Phan TX, Nguyen VH et al (2015) Salmonella Typhimurium suppresses tumor growth via the pro-inflammatory cytokine interleukin-1beta. Theranostics 5:1328–1342CrossRefPubMedPubMedCentralGoogle Scholar
  12. 12.
    Chorobik P, Czaplicki D, Ossysek K et al (2013) Salmonella and cancer: from pathogens to therapeutics. Acta Biochim Pol 60:285–297PubMedGoogle Scholar
  13. 13.
    Forbes NS (2010) Engineering the perfect (bacterial) cancer therapy. Nat Rev Cancer 10:785–794CrossRefPubMedPubMedCentralGoogle Scholar
  14. 14.
    Williams KJ, Joyce G, Robertson BD (2010) Improved mycobacterial tetracycline inducible vectors. Plasmid 64:69–73CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Royo JL, Becker PD, Camacho EM et al (2007) In vivo gene regulation in Salmonella spp. by a salicylate-dependent control circuit. Nat Methods 4:937–942CrossRefPubMedGoogle Scholar
  16. 16.
    Cebolla A, Royo JL, De Lorenzo V et al (2002) Improvement of recombinant protein yield by a combination of transcriptional amplification and stabilization of gene expression. Appl Environ Microbiol 68:5034–5041CrossRefPubMedPubMedCentralGoogle Scholar
  17. 17.
    Medina C, Camacho EM, Flores A et al (2011) Improved expression systems for regulated expression in Salmonella infecting eukaryotic cells. PLoS One 6:e23055CrossRefPubMedPubMedCentralGoogle Scholar
  18. 18.
    Camacho EM, Mesa-Pereira B, Medina C et al (2016) Engineering Salmonella as intracellular factory for effective killing of tumour cells. Sci Rep 6:30591CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Beuzon CR, Meresse S, Unsworth KE et al (2000) Salmonella maintains the integrity of its intracellular vacuole through the action of SifA. EMBO J 19:3235–3249CrossRefPubMedPubMedCentralGoogle Scholar
  20. 20.
    Mesa-Pereira B, Medina C, Camacho EM et al (2013) Novel tools to analyze the function of Salmonella effectors show that SvpB ectopic expression induces cell cycle arrest in tumor cells. PLoS One 8:e78458CrossRefPubMedPubMedCentralGoogle Scholar
  21. 21.
    Kuriyama S, Masui K, Sakamoto T et al (1998) Bystander effect caused by cytosine deaminase gene and 5-fluorocytosine in vitro is substantially mediated by generated 5-fluorouracil. Anticancer Res 18:3399–3406PubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2018

Authors and Affiliations

  • Carlos Medina
    • 1
    • 2
    Email author
  • Beatriz Mesa-Pereira
    • 3
    • 4
  • Eva M. Camacho
    • 1
  • Amando Flores
    • 1
  • Eduardo Santero
    • 1
  1. 1.Departamento de Biología Molecular e Ingeniería Bioquímica, Centro Andaluz de Biología del DesarrolloCSIC/ Universidad Pablo de Olavide/ Junta de AndalucíaSevilleSpain
  2. 2.Departamento de Microbiología, Facultad de BiologíaUniversidad de SevillaSevilleSpain
  3. 3.Teagasc Food Research CentreTeagasc MooreparkCo. CorkIreland
  4. 4.APC Microbiome InstituteUniversity College CorkCorkIreland

Personalised recommendations