Autophagy Inducing Capacities of a
Commercial Preparation Based on of Bacillus thuringiensis Cry 1A-2A Toxins in Human Macrophages:
Implication Against Mycobacterial Infections
Ruiz DH Andy1, Juárez Esmeralda1, González Yolanda1, Favela-Hernández Juan Manuel2 and Guerrero
1Instituto Nacional de Enfermedades Respiratorias (INER), Departamento de Microbiología, México
2Universidad Juárez del Estado de Durango, Facultad de Química, México
3Universidad Autónoma de Zacatecas, Unidad Académica de Ciencias Biológicas, Lab de Imunobiologia, Campus II, Zacatecas, Zac, México
*Corresponding Author: Guerrero G Gloria, Universidad Autónoma de Zacatecas, Unidad Académica de Ciencias Biológicas, Lab de Imunobiologia, Campus II,
Zacatecas, Zac, México.
November 07, 2022; Published: November 18, 2022
Autophagy is a highly conserved degradative and recycling program to maintain homeostasis. In particular, it plays an important role in the innate immune response against intracellular pathogens. Several studies have shown that BCG and/or M. tuberculosis block autophagosome formation, inhibiting thus, activation of the autophagy machinery, and survival of mycobacteria. Human macrophages preparation from Peripheral blood mononuclear cells (buffy coats of the healthy donor) (blood bank of INER. MX). Monocytes were further isolated by CD14 positive selection and differentiated into monocyte-derived macrophages. Infection of macrophages with M. bovis BCG vaccine (ATCC, Manassas, VA, US) at MOI 1:5. Stimulation with Cry1A-Cr2A (5 mg/ml Rapamycin (250 mg/ml) and/or Wortmannin (100 nM). Autophagy detection and fluorescence microscopy were made in the uninfected well and the infected-stimulated cells were stained with rabbit anti-LC3B coupled to Alexa Fluor 488. The percentage of cells with more than 5 LC3+ puncta (autophagosomes) was calculated as well as the percentage of bacteria co-localizing with LC3. The percentage of mycobacteria BCG co-localizing with light chain 3 (LC3-II) in human macrophages is greater in infected and stimulated human macrophages with the commercial preparation based on Cry1A-Cry2A Bt proteins versus rapamycin and/or Wortmannin. The commercial preparation of Cry1A-Cry2A in combination with the BCG vaccine represents a potential alternative to enhance the autophagy-mediated elimination of intracellular pathogens such as M. tuberculosis.
Keywords: Bacillus thuringiensis; Cry1A Toxins; Autophagy; Macrophages; Microtubule Light Chain 3 (LC3); Tuberculosis
- “Guidelines for the treatment of drug-susceptible tuberculosis and patient care”. 2017 update. WHO (2017).
- “The top 10 causes of death”. WHO (2019).
- Zumla A and Maeurer M. “Host-Directed therapies for tackling multi-drug resistant tuberculosis: learning from the Pasteur-Bechamp debates”. Clinical Infectious Diseases 61 (2015): 1432-1438.
- Maher D and Raviglione M. “Global epidemiology of Tuberculosis”. Clinics in Chest Medicine 26 (2005): 167-182.
- Young DB., et al. “Confronting the scientific obstacle to global control of Tuberculosis”. Journal of Clinical Investigation 118 (2008): 1255-1260.
- Trunz BB., et al. “Effect of BCG vaccination on childhood tuberculous meningitis and miliary tuberculosis worldwide: a meta-analysis and assessment of cost-effectivenes”. Lancet 367 (2006): 1173-1180.
- da Costa A C., et al. “Recombinant BCG: innovations on an old vaccine. Scope of BCG strains and strategies to improve long-lasting memory”. Frontiers in Immunology 5 (2014): 152.
- Wilkie M., et al. “Functional in-vitro evaluation of the non-specific effects of BCG vaccination in a randomised controlled clinical study”. Scientific Report 12 (2022): 7808.
- Abebe F. “Is Interferon-gamma the right marker for Bacille Calmette-Güerin-induced immune protection? The missing link in our understanding of Tuberculosis”. Clinical and Experimental Immunology 169 (2012): 213-219.
- Koul A., et al. “Interplay between mycobacteria and host signaling pathways”. Nature Reviews Microbiology 2 (2004): 189-202.
- Pulendran B and Ahmed R. “Translating innate immunity into immunological memory: implications for vaccine development”. Cell 124 (2006): 849-863.
- Mohr I and Sonenberg N. “Host translation of the nexus of infection and immunity”. Cell Host Microbe 12 (2012): 470-483.
- Kolloli A and Subbian S. “Host-directed therapeutic strategies for tuberculosis”. Frontiers in Medicine (Lausanne). 4 (2017): 171.
- Deretic V. “Autophagy in inflammation, infection, and immunometabolism”. Immunity 54 (2021): 437-453.
- Benbrook DM and Long A. “Integration of autophagy, proteasome degradation, unfolded protein response and apoptosis”. Experimental Oncology 34 (2012): 286-297.
- Kimmey JM and Stallings CL. “Bacterial pathogens versus autophagy: implications for therapeutic interventions”. Trends in Molecular Medicine 22 (2016): 1060-1076.
- Klionsky DJ and Emr SD. “Autophagy as a regulated pathway of cellular degradation”. Science 290 (2000): 1717-1721.
- Pareja ME and Colombo MI. “Autophagic clearance of bacterial pathogens: molecular recognition of intracellular microorganisms”. Frontiers in Cellular and Infection Microbiology 3 (2013): 54.
- Bah A and Vergne I. “Macrophage autophagy and bacterial infections”. Frontiers in Immunology 8 (2017): 1483.
- Levine B and Kroemer G. “Biological Functions of Autophagy Genes: A Disease Perspective”. Cell 176 (2019): 11-42.
- Castillo EF., et al. “Autophagy protects against active tuberculosis by suppressing bacterial burden and inﬂammation”. Proceedings of the National Academy of Sciences of the United States of America 109 (2012): E3168-76.
- Goletti D., et al. “Autophagy in Mycobacterium tuberculosis infection: a passe partout to ﬂush the intruder out?” Cytokine Growth Factor Review 24 (2013): 335-343.
- Romagnoli A., et al. “ESX-1 dependent impairment of autophagic ﬂux by Mycobacterium tuberculosis in human dendritic cells”. Autophagy 8 (2012): 1357-1370.
- Kaufmann SHE., et al. “Host- directed therapies for bacterial and viral infections”. Nature Reviews Drug Discovery 17 (2018): 35-56.
- Gutierrez MG., et al. “Autophagy is a defense mechanism inhibiting BCG and Mycobacterium tuberculosis survival in infected macrophages”. Cell 119 (2004): 753-766.
- Palk S., et al. “Autophagy: A new strategy for host-directed therapy of tuberculosis”. Virulence 10 (2019): 448-459.
- Pellegrini JM., et al. “Shedding Light on Autophagy During Human Tuberculosis. A Long Way to Go”. Frontiers in Cellular and Infection Microbiology 11 (2022): 820095.
- Pellegrini JM., et al. “Neutrophil autophagy during human active tuberculosis is modulated by SLAMF1”. Autophagy 17 (2021): 2629-2638.
- Khan A., et al. “An autophagy-inducing and TLR-2 activating BCG vaccine induces a robust protection against Tuberculosis in mice”. NPJ Vaccines 4 (2019): 34.
- Gupta A., et al. “Targeted pulmonary delivery of inducers of host macrophage autophagy as a potential host-directed chemotherapy of tuberculosis”. Advanced Drug Delivery Reviews 1 (2016): 10-20.
- Rovetta AI., et al. “IFN--mediated immune responses enhance autophagy against Mycobacterium tuberculosis antigens in patients with active tuberculosis”. Autophagy 10 (2014): 2109-2121.
- Krakauer T. “Inflammasomes, Autophagy, and Cell Death: The Trinity of Innate Host Defense against Intracellular Bacteria”. Nutrition and Inflammation (2019).
- Kuma A., et al. “Autophagy-monitoring and autophagy-deficiente mice”. Autophagy 12 (2017): 1619-1628.
- Lerena MC., et al. “Mycobacterium marinum induces a marked LC3 recruitment to its containing phagosome that depends on a functional ESX-1 secretion system”. Cell Microbiology 13 (2011): 814-835.
- Mahrpour M., et al. “Autophagy in Health and Disease. 1. Regulation and significance of autophagy; an overview”. American Journal of Physiology-Cell Physiology 298 (2010): C776-C785.
- Mizushima N., et al. “Autophagy fights disease through cellular self-digestion”. Nature 451 (2008): 1069-1075.
- Delgado MA and Deretic V. “Toll-like receptors in control of immunological autophagy”. Cell Death and Differentiation 16 (2009): 976-983.
- Yuting M., et al. “Autophagy and cellular immune responses”. Immunity 39 (2013): 211-227.
- Vincent O., et al. “The WIPI Gene Family and Neurodegenerative Diseases: Insights From Yeast and Dictyostelium Models”. Frontiers in Cell and Developmental Biology 9 (2021): 737071.
- Kumar S., et al. “Mammalian Atg8-family proteins are upstream regulators of the lysosomal system by controlling MTOR and TFEB”. Autophagy 16 (2020): 2305-2306.
- Ragus M., et al. “Architecture. Architecture of the Atg17 complex as a Scaffold for Autophagosome Biogenesis”. Cell 151 (2012): 1501-1512.
- Alers S., et al. “ATG13. Just a companion, or an executor of the autophagic program. ATG13”. Autophagy 6 (2014): 944-956.
- Soto-Burgos J., et al. “Dynamics of Autophagosome Formation”. Plant Physiology 126 (2018): 219-229.
- Stanley RE., et al. “The beginning of the end: How scaffolds nucleate autophagosome biogenesis”. Trends in Cell Biology 24 (2014): 1-19.
- Zavodszky E., et al. “Biology and trafficking of ATG9 and ATG16l two proteins that regulate autophagosome formation”. FEBS Letter 587 (2013): 1988-1996.
- Harris J., et al. “Th1-Th2 polarisation and autophagy in the control of intracellular mycobacteria by macrophages”. Veterinary Immunology and Immunopathology 128 (2009): 37-43.
- Songane M., et al. “The role of autophagy in host defence against Mycobacterium tuberculosis infection”. Tuberculosis (Edinb) 92 (2012): 388-396.
- Qin C., et al. “The molecular regulation of autophagy in antimicrobial immunity”. Journal of Molecular Cell Biology 14 (2022): mjac015.
- Siregar TAP., et al. “The autophagy-resistant Mycobacterium tuberculosis Beijing strain upregulates KatG to evade starvation-induced autophagic restriction”. Pathogens and Disease 80 (2022): ftac004.
- Jurado JO., et al. “IL-17 and IFN-gamma expression in lymphocytes from patients with active tuberculosis correlates with the severity of the disease”. Journal of Leukocyte Biology 91 (2012): 991-1002.
- Laopanupong T., et al. “Lysosome repositioning as an autophagy escape mechanism by Mycobacterium tuberculosis Beijing strain”. Scientific Report 11 (2021): 4342.
- Intemann CD., et al. “Autophagy gene variant IRGM -261T contributes to protection from tuberculosis caused by Mycobacterium tuberculosis but not by africanum strains”. PLoS Pathogen 5 (2009): e1000577.
- Thellung S., et al. “Autophagy activator drugs: A new Opportunity in Neuroprotection from Misfolded Protein Toxicity”. International Journal of Molecular Sciences 20 (2019): 901-934.
- Favela-Hernández JM., et al. “The potential of a commercial product based on Bacillus thuringiensis Cry1A-Cry2A as a immunogen and adjuvant”. Madridge Journal of Immunology 2 (2018): 58-64.
- Verdin-Teran SL., et al. “Immunization with Cry1Ac from Bacillus thuringiensis Increases Intestinal IgG Response and Induces the Expression of FcRn in the Intestinal Epithelium of Adult Mice”. Journal of Immunology 70 (2009): 596-607.
- Rubio-Infante N., et al. “The Macrophage Activation Induced by Bacillus thuringiensis Cry1Ac Protoxin Involves ERK1/2 and p38 Pathways and the Interaction with Cell-Surface-HSP70”. Journal of Cellular Biochemistry 119 (2018): 580-598.
- Mehta P., et al. “Noncanonical autophagy: one small step for LC3, one giant leap for immunity”. Current Opinion in Immunology 26 (2014): 69-75.
- Ma J., et al. “Dectin-1-triggered recruitment of light chain 3 protein to phagosomes facilitates major histocompatibility complex class II presentation of fungal-derived antigens”. Journal of Biological Chemistry 287 (2012): 34149-34156.
- Sprenkeler EG., et al. “LC3-associated phagocytosis: a crucial mechanism for antifungal host defence against Aspergillus fumigatus”. Cell Microbiology 18 (2016): 1208-1216.
- Martinez J., et al. “Molecular characterizationof LC3-associated phagocytosis reveals distinctroles for Rubicon, NOX2 and autophagy proteins”. Nature Cell Biology 17 (2015): 893-906.
- Gubas A and Dikic I. “A guide to the regulation of selective autophagy receptors”. FEBS Journal 289 (2022): 75-89.
- Park HJ., et al. “IL-10 inhibits the starvation induced autophagy in macrophages via class I phosphatidylinositol 3-kinase (PI3K) pathway”. Molecular Immunology 48 (2011): 720-727.
- Feng CG., et al. “Interferon inducible immunity-related GTPase Irgm1 regulates IFN gamma-dependent host defense, lymphocyte survival and autophagy”. Autophagy 5 (2009): 232-234.
- Kim BH., et al. “A family of IFN-gamma-inducible 65-kD GTPases protects against bacterial infection”. Science 332 (2011): 717-721.
- Sanjuan MA., et al. “Toll-like receptor signalling in macrophages links the autophagy pathway to phagocytosis”. Nature 450 (2007): 1253-1257.
- Sanjuan MA., et al. “Toll-like receptor signaling in the lysosomal pathways”. Immunology Review 227 (2009): 203-220.