title: Identification of Signaling Pathways by Which CD40 Stimulates Autophagy and Antimicrobial Activity against Toxoplasma gondii in Macrophages.
abstract: CD40 is an important stimulator of autophagy and autophagic killing of Toxoplasma gondii in host cells. In contrast to autophagy induced by nutrient deprivation or pattern recognition receptors, less is known about the effects of cell-mediated immunity on Beclin 1 and ULK1, key regulators of autophagy. Here we studied the molecular mechanisms by which CD40 stimulates autophagy in macrophages. CD40 ligation caused biphasic Jun N-terminal protein kinase (JNK) phosphorylation. The second phase of JNK phosphorylation was dependent on autocrine production of tumor necrosis factor alpha (TNF-α). TNF-α and JNK signaling were required for the CD40-induced increase in autophagy. JNK signaling downstream of CD40 caused Ser-87 phosphorylation of Bcl-2 and dissociation between Bcl-2 and Beclin 1, an event known to stimulate the autophagic function of Beclin 1. However, TNF-α alone was unable to stimulate autophagy. CD40 also stimulated autophagy via a pathway that included calcium/calmodulin-dependent kinase kinase β (CaMKKβ), AMP-activated protein kinase (AMPK), and ULK1. CD40 caused AMPK phosphorylation at its activating site, Thr-172. This effect was mediated by CaMKKβ and was not impaired by neutralization of TNF-α. CD40 triggered AMPK-dependent Ser-555 phosphorylation of ULK1. CaMKKβ, AMPK, and ULK1 were required for CD40-induced increase in autophagy. CD40-mediated autophagic killing of Toxoplasma gondii is known to require TNF-α. Knockdown of JNK, CaMKKβ, AMPK, or ULK1 prevented T. gondii killing in CD40-activated macrophages. The second phase of JNK phosphorylation-Bcl-2 phosphorylation-Bcl-2-Beclin 1 dissociation and AMPK phosphorylation-ULK1 phosphorylation occurred simultaneously at ∼4 h post-CD40 stimulation. Thus, CaMKKβ and TNF-α are upstream molecules by which CD40 acts on ULK1 and Beclin 1 to stimulate autophagy and killing of T. gondii.
authors.affiliation: Department of Pathology, Case Western Reserve University, Cleveland, Ohio, USA.
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Liu
authors.affiliation: Division of Infectious Diseases and HIV Medicine, Department of Medicine, Case Western Reserve University, Cleveland, Ohio, USA.
authors.forename: Yalitza
authors.initials: Y
authors.lastname: Lopez Corcino
authors.affiliation: Division of Infectious Diseases and HIV Medicine, Department of Medicine, Case Western Reserve University, Cleveland, Ohio, USA.
authors.forename: Jose-Andres C
authors.initials: JA
authors.lastname: Portillo
authors.affiliation: Division of Infectious Diseases and HIV Medicine, Department of Medicine, Case Western Reserve University, Cleveland, Ohio, USA.
authors.forename: Yanling
authors.initials: Y
authors.lastname: Miao
authors.affiliation: Department of Pathology, Case Western Reserve University, Cleveland, Ohio, USA Division of Infectious Diseases and HIV Medicine, Department of Medicine, Case Western Reserve University, Cleveland, Ohio, USA Department of Ophthalmology and Visual Sciences, Case Western Reserve University, Cleveland, Ohio, USA carlos.subauste@case.edu.
authors.forename: Carlos S
authors.initials: CS
authors.lastname: Subauste
mesh_terms: D055372:AMP-Activated Protein Kinases / Q000378:metabolism; D000818:Animals; D000890:Anti-Infective Agents / Q000494:pharmacology*; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism; D001343:Autophagy / Q000276:immunology*; D000071186:Beclin-1 / Q000378:metabolism; D019013:CD40 Antigens / Q000276:immunology*; D006801:Humans; D007111:Immunity, Cellular / Q000276:immunology; D008264:Macrophages / Q000276:immunology* / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D010766:Phosphorylation / Q000276:immunology; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D015398:Signal Transduction / Q000276:immunology*; D014122:Toxoplasma / Q000276:immunology*; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
chemical_list: D000890:Anti-Infective Agents; D051017:Apoptosis Regulatory Proteins; D000071186:Beclin-1; D019013:CD40 Antigens; D014409:Tumor Necrosis Factor-alpha; D017346:Protein Serine-Threonine Kinases; D055372:AMP-Activated Protein Kinases