The CD40-Autophagy Pathway Is Needed for Host Protection Despite IFN-Γ-Dependent Immunity and CD40 Induces Autophagy via Control of P21 Levels

Portillo, Jose‐Andres C.; Okenka, Genevieve; Reed, E.; Subauste, Angela; Grol, Jennifer Van; Gentil, Katrin; Komatsu, Masaaki; Tanaka, Keiji; Landreth, Gary E.; Levine, Beth; Subauste, Carlos S.

doi:10.1371/journal.pone.0014472

Cited by 71 publications

(117 citation statements)

References 64 publications

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“…Although it remains controversial whether autophagy is involved in the IFN-g-mediated anti-T. gondii cellular response (17,35), the recruitment of the immunityrelated GTPases to the parasite is regulated independently of at least Atg9a and Atg14, which are key players in autophagosome formation in autophagy. Thus, our current study expands the notion that autophagy proteins differentially participate in mouse IFN-ginduced anti-T. gondii activity, which originally was proposed by the previous pioneer findings about the essential, but autophagosomeindependent, function of Atg5 and the dispensable role of another essential autophagy regulator, Beclin1, in IFN-g-induced antiparasite activity (17,36).…”

Section: Discussionsupporting

confidence: 80%

Role of Mouse and Human Autophagy Proteins in IFN-γ–Induced Cell-Autonomous Responses against Toxoplasma gondii

Ohshima

Lee

Sasai

et al. 2014

The Journal of Immunology

111

127

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IFN-γ mediates cellular innate immunity against an intracellular parasite, Toxoplasma gondii, by inducing immunity-related GTPases such as p47 IFN-γ–regulated GTPases (IRGs) and p65 guanylate-binding proteins (GBPs), which also participate in antibacterial responses via autophagy. An essential autophagy protein, Atg5, was previously shown to play a critical role in anti–T. gondii cell-autonomous immunity. However, the involvement of other autophagy proteins remains unknown. In this study, we show that essential autophagy proteins differentially participate in anti–T. gondii cellular immunity by recruiting IFN-γ–inducible GTPases. IFN-γ–induced suppression of T. gondii proliferation and recruitment of an IRG Irgb6 and GBPs are profoundly impaired in Atg7- or Atg16L1-deficient cells. In contrast, cells lacking other essential autophagy proteins, Atg9a and Atg14, are capable of mediating the anti–T. gondii response and recruiting Irgb6 and GBPs to the parasites. Although IFN-γ also stimulates anti–T. gondii cellular immunity in humans, whether this response requires GBPs and human autophagy proteins remains to be seen. To analyze the role of human ATG16L1 and GBPs in IFN-γ–mediated anti–T. gondii responses, human cells lacking ATG16L1 or GBPs were generated by the Cas9/CRISPR genome-editing technique. Although both ATG16L1 and GBPs are dispensable for IFN-γ–induced inhibition of T. gondii proliferation in the human cells, human ATG16L1 is also required for the recruitment of GBPs. Taken together, human ATG16L1 and mouse autophagy components Atg7 and Atg16L1, but not Atg9a and Atg14, participate in the IFN-γ–induced recruitment of the immunity-related GTPases to the intracellular pathogen.

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Section: Discussionsupporting

confidence: 80%

Role of Mouse and Human Autophagy Proteins in IFN-γ–Induced Cell-Autonomous Responses against Toxoplasma gondii

Ohshima

Lee

Sasai

et al. 2014

The Journal of Immunology

111

127

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“…As opposed to p27, the role of another CDK inhibitor p21 is more equivocal. Although p21 has been shown to impair C(2)-ceramide-and CD40-induced autophagic activity, enforced expression of cytoplasmic p21 induces autophagy (Fujiwara et al, 2008;Portillo et al, 2010;. These findings indicate that, similar to p53, the regulatory effect of p21 on autophagy may depend on its subcellular localization.…”

Section: V12mentioning

confidence: 95%

The autophagic paradox in cancer therapy

et al. 2011

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Autophagy, hallmarked by the formation of doublemembrane bound organelles known as autophagosomes, is a lysosome-dependent pathway for protein degradation. The role of autophagy in carcinogenesis is context dependent. As a tumor-suppressing mechanism in earlystage carcinogenesis, autophagy inhibits inflammation and promotes genomic stability. Moreover, disruption of autophagy-related genes accelerates tumorigenesis in animals. However, autophagy may also act as a prosurvival mechanism to protect cancer cells from various forms of cellular stress. In cancer therapy, adaptive autophagy in cancer cells sustains tumor growth and survival in face of the toxicity of cancer therapy. To this end, inhibition of autophagy may sensitize cancer cells to chemotherapeutic agents and ionizing radiation. Nevertheless, in certain circumstances, autophagy mediates the therapeutic effects of some anticancer agents. Data from recent studies are beginning to unveil the apparently paradoxical nature of autophagy as a cellfate decision machinery. Taken together, modulation of autophagy is a novel approach for enhancing the efficacy of existing cancer therapy, but its Janus-faced nature may complicate the clinical development of autophagy modulators as anticancer therapeutics.

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“…At the indicated time points, monolayers were fixed and stained with Diff-Quick (Dade Diagnostics, Aguada, Puerto Rico). The percentages of infected macrophages and the numbers of parasites per 100 cells in triplicate monolayers were determined by light microscopy by counting at least 200 cells per monolayer (19,21).…”

Section: Methodsmentioning

confidence: 99%

“…The interaction of CD40 with CD154 (CD40 ligand) increases the formation of autophagosomes and autolysosomes (increased autophagy flux) and increases the conversion of the autophagy protein LC3 I to LC3 II, events that require Atg5, Atg7, and Beclin 1 (19)(20)(21)(22)(23)(24). CD40 triggers autophagy-mediated killing of Toxoplasma gondii (19-21, 23, 24) and probably of Mycobacterium tuberculosis (25).…”

mentioning

confidence: 99%

“…CD40 ligation in mammalian cells results in the encasement of T. gondii by an LC3-positive (LC3 ϩ ) structure, followed by Rab7-mediated vacuole-lysosome fusion and parasite killing dependent on Atg5, Atg7, Beclin 1, PI3KC3, protein kinase double-stranded RNA-dependent (PKR), and lysosomal enzymes (19-21, 23, 24). These events are relevant to protection against toxoplasmosis since CD40 Ϫ/Ϫ , Becn1 ϩ/Ϫ , and PKR Ϫ/Ϫ mice or mice with an Atg7 deficiency targeted to macrophages/microglia are susceptible to cerebral and ocular toxoplasmosis (20,21).…”

mentioning

confidence: 99%

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Identification of Signaling Pathways by Which CD40 Stimulates Autophagy and Antimicrobial Activity against Toxoplasma gondii in Macrophages

et al. 2016

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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-␣). TNF-␣ 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-␣ alone was unable to stimulate autophagy. CD40 also stimulated autophagy via a pathway that included calcium/calmodulin-dependent kinase kinase ␤ (CaMKK␤), AMP-activated protein kinase (AMPK), and ULK1. CD40 caused AMPK phosphorylation at its activating site, Thr-172. This effect was mediated by CaMKK␤ and was not impaired by neutralization of TNF-␣. CD40 triggered AMPK-dependent Ser-555 phosphorylation of ULK1. CaMKK␤, AMPK, and ULK1 were required for CD40-induced increase in autophagy. CD40-mediated autophagic killing of Toxoplasma gondii is known to require TNF-␣. Knockdown of JNK, CaMKK␤, 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 ϳ4 h post-CD40 stimulation. Thus, CaMKK␤ and TNF-␣ are upstream molecules by which CD40 acts on ULK1 and Beclin 1 to stimulate autophagy and killing of T. gondii.

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The CD40-Autophagy Pathway Is Needed for Host Protection Despite IFN-Γ-Dependent Immunity and CD40 Induces Autophagy via Control of P21 Levels

Cited by 71 publications

References 64 publications

Role of Mouse and Human Autophagy Proteins in IFN-γ–Induced Cell-Autonomous Responses against Toxoplasma gondii

Role of Mouse and Human Autophagy Proteins in IFN-γ–Induced Cell-Autonomous Responses against Toxoplasma gondii

The autophagic paradox in cancer therapy

Identification of Signaling Pathways by Which CD40 Stimulates Autophagy and Antimicrobial Activity against Toxoplasma gondii in Macrophages

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