Induction of autophagy through CLEC4E in combination with TLR4: an innovative strategy to restrict the survival of Mycobacterium tuberculosis

Autophagy. 2020 Jun;16(6):1021-1043. doi: 10.1080/15548627.2019.1658436. Epub 2019 Sep 8.


Host-directed therapies are gaining considerable impetus because of the emergence of drug-resistant strains of pathogens due to antibiotic therapy. Therefore, there is an urgent need to exploit alternative and novel strategies directed at host molecules to successfully restrict infections. The C-type lectin receptor CLEC4E and Toll-like receptor TLR4 expressed by host cells are among the first line of defense in encountering pathogens. Therefore, we exploited signaling of macrophages through CLEC4E in association with TLR4 agonists (C4.T4) to control the growth of Mycobacterium tuberculosis (Mtb). We observed significant improvement in host immunity and reduced bacterial load in the lungs of Mtb-infected mice and guinea pigs treated with C4.T4 agonists. Further, intracellular killing of Mtb was achieved with a 10-fold lower dose of isoniazid or rifampicin in conjunction with C4.T4 than the drugs alone. C4.T4 activated MYD88, PtdIns3K, STAT1 and RELA/NFKB, increased lysosome biogenesis, decreased Il10 and Il4 gene expression and enhanced macroautophagy/autophagy. Macrophages from autophagy-deficient (atg5 knockout or Becn1 knockdown) mice showed elevated survival of Mtb. The present findings also unveiled the novel role of CLEC4E in inducing autophagy through MYD88, which is required for control of Mtb growth. This study suggests a unique immunotherapeutic approach involving CLEC4E in conjunction with TLR4 to restrict the survival of Mtb through autophagy.

Abbreviations: 3MA: 3 methyladenine; AO: acridine orange; Atg5: autophagy related 5; AVOs: acidic vesicular organelles; BECN1: beclin 1, autophagy related; BMDMs: bone marrow derived macrophages; bw: body weight; C4.T4: agonists of CLEC4E (C4/TDB) and TLR4 (T4/ultra-pure-LPS); CFU: colony forming unit; CLEC4E/Mincle: C-type lectin domain family 4, member e; CLR: c-type lectin receptor; INH: isoniazid; LAMP1: lysosomal-associated membrane protein 1; MφC4.T4: Mtb-infected C4.T4 stimulated macrophages; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; MDC: monodansylcadaverine; MTOR: mechanistic target of rapamycin kinase; MYD88: myeloid differentiation primary response 88; NFKB: nuclear factor of kappa light polypeptide gene enhance in B cells; NLR: NOD (nucleotide-binding oligomerization domain)-like receptors; PFA: paraformaldehyde; PPD: purified protein derivative; PtdIns3K: class III phosphatidylinositol 3-kinase; RELA: v-rel reticuloendotheliosis viral oncogene homolog A (avian); RIF: rifampicin; RLR: retinoic acid-inducible gene-I-like receptors; TDB: trehalose-6,6´-dibehenate; TLR4: toll-like receptor 4; Ultra-pure-LPS: ultra-pure lipopolysaccharide-EK; V-ATPase: vacuolar-type H+ ATPase.

Keywords: Becn1 knockdown; Mycobacterium tuberculosis; atg5 knockout; Anti-TB drugs; CLEC4E/Mincle; TLR4; autophagy; host-directed immunotherapy; macrophages; tuberculosis.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Autophagy / drug effects
  • Autophagy / genetics*
  • Autophagy-Related Protein 5 / genetics
  • Autophagy-Related Protein 5 / metabolism
  • Beclin-1 / genetics
  • Beclin-1 / metabolism
  • CD4-Positive T-Lymphocytes / drug effects
  • CD4-Positive T-Lymphocytes / immunology
  • Guinea Pigs
  • Host Microbial Interactions
  • Interleukin-10 / genetics
  • Interleukin-10 / metabolism
  • Interleukin-4 / genetics
  • Interleukin-4 / metabolism
  • Lectins, C-Type / metabolism*
  • Lung / drug effects*
  • Lung / immunology
  • Lung / metabolism
  • Lung / microbiology
  • Lysosomes / metabolism
  • Macrophages / drug effects
  • Macrophages / metabolism*
  • Macrophages / ultrastructure
  • Membrane Proteins / metabolism*
  • Mice
  • Mice, Inbred C57BL
  • Microscopy, Electron, Scanning
  • Mycobacterium tuberculosis / growth & development*
  • Mycobacterium tuberculosis / immunology
  • Myeloid Differentiation Factor 88 / metabolism
  • Phosphatidylinositol 3-Kinases / metabolism
  • Phosphorylation
  • Rifampin / pharmacology
  • Signal Transduction / drug effects
  • Signal Transduction / genetics
  • Signal Transduction / immunology
  • Th1 Cells / drug effects
  • Th1 Cells / immunology
  • Th17 Cells / drug effects
  • Th17 Cells / immunology
  • Toll-Like Receptor 4 / agonists
  • Toll-Like Receptor 4 / metabolism*
  • Transcription Factor RelA / metabolism


  • Autophagy-Related Protein 5
  • Beclin-1
  • Becn1 protein, mouse
  • Clecsf8 protein, mouse
  • Lectins, C-Type
  • Membrane Proteins
  • Myeloid Differentiation Factor 88
  • Rela protein, mouse
  • Tlr4 protein, mouse
  • Toll-Like Receptor 4
  • Transcription Factor RelA
  • Interleukin-10
  • Interleukin-4
  • Rifampin

Grants and funding

We thank the Council of Scientific & Industrial Research (CSIR) for financial support (Project Number: OLP 088).