Neddylation pathway regulates the proliferation and survival of macrophages

Li, Lihui; Liu, Bing-Hong; Dong, Tao; Lee, Hyuk Woo; Yu, Jinha; Zheng, Yijie; Gao, Haifeng; Zhang, Yan; Chu, Yiwei; Liu, Guangwei; Niu, Weidong; Zheng, Shangen; Jeong, Lak Shin; Jia, Lijun

doi:10.1016/j.bbrc.2013.02.028

Cited by 40 publications

(50 citation statements)

References 28 publications

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“…Previous studies have also revealed that MLN4924 markedly suppressed inflammatory responses mediated by innate immune cells, including macrophages and dendritic cells (19,20,34). The results of the present study suggest that the same suppressed inflammatory response is experienced in neutrophils.…”

Section: Discussionsupporting

confidence: 81%

“…Furthermore, prolonged and complete (1.0 µM for 24 or 48 h) inhibition of neddylation led to decreased macrophage viability (34). To detect whether MLN4924 inhibited LPS-induced inflammatory cytokine production in neutrophils as a result of neutrophil apoptosis, the viability of neutrophils was examined using ATPlite assays.…”

Section: Mln4924 Pretreatment Followed By Lps Stimulation For 4 H Revmentioning

confidence: 99%

See 1 more Smart Citation

MLN4924 suppresses lipopolysaccharide‑induced proinflammatory cytokine production in neutrophils in a dose‑dependent manner

Jin

Jing²,

Ye³

et al. 2018

Oncol Lett

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Abstract. Neddylation is a ubiquitination-like pathway. It has been reported that neddylation inhibition with the pharmacological agent MLN4924 potently uppresses lipopolysaccharide (LPS)-induced proinflammatory cytokine production, including tumor necrosis factor (TNF)-α and interleukin (IL)-6, by preventing the degradation of phosphorylated inhibitor of κB (p-IκB) in macrophages. However, whether neddylation serves a similar role in neutrophils remains unknown. In the present study MLN4924 treatment led to the accumulation of P-IκBα in neutrophils as well as the decreased production of TNF-α, IL-6 and IL-1β in response to LPS, in a dose-dependent manner. The viability of neutrophils was only marginally affected in the same conditions, without statistical significance. Furthermore, the nuclear factor (NF)-κB inhibitor JSH-23 mimicked the effects of MLN4924 in neutrophils, and the inhibitory effects of MLN4924 on LPS-induced proinflammatory cytokine production diminished in the presence of JSH-23. Thus, the results of the present study suggest that neddylation inhibition suppresses neutrophil function by suppressing the NF-κB signaling pathway. IntroductionNeutrophils serve crucial roles in the anti-bacterial and anti-fungal innate immune response and produce cytokines to initiate inflammatory responses; however, their inappropriate or excessive activation contributes to tissue damage during autoimmune and inflammatory diseases (1). Furthermore, the activity of neutrophils is associated with tumor progression, as demonstrated in colitis-associated cancer and glioblastoma (2,3).Previous studies have revealed that in order to protect the human body from invading pathogens and endogenous damage-associated molecules, neutrophils sense these threats through innate immune receptors, such as toll-like receptors (TLRs) and other pattern recognition receptors, to upregulate the secretion of inflammatory cytokines (4,5). The recognition of lipopolysaccharide (LPS) from gram-negative bacteria by TLR4 is a well-characterized example with the initiation of proinflammatory gene transcription through multiple signaling pathways, including nuclear factor (NF)-κB) (6,7). The exposure of neutrophils to bacterial LPS may contribute to tissue damage, under certain conditions, including endotoxic shock and infection-induced adult respiratory distress syndrome (8).Neddylation is a novel-type protein post-translational modification, a multistep enzymatic process that adds the ubiquitin-like molecule neural precursor cell expressed developmentally downregulated protein 8 (Nedd8) to target proteins in an ATP-dependent manner. Neddylation is vital for a range of processes, including cell viability, growth and development (9-12). To the best of our knowledge, the only known E1 for neddylation is a heterodimer comprised of the amyloid precursor protein-binding protein and ubiquitin-like modifier activating enzyme 3 (9-12). A previous study identified the ubiquitin-conjugating enzymes Ube2M and Ube2F as Nedd8 E2s (13). Several targets for N...

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Section: Discussionsupporting

confidence: 81%

Section: Mln4924 Pretreatment Followed By Lps Stimulation For 4 H Revmentioning

confidence: 99%

MLN4924 suppresses lipopolysaccharide‑induced proinflammatory cytokine production in neutrophils in a dose‑dependent manner

Jin

Jing²,

Ye³

et al. 2018

Oncol Lett

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“…For ATPlite assay, cells were seeded at 3,000 cells per well in 96-well black plates, cultured overnight, and treated with MLN4924 or DMSO for 96 h. Cell viability was determined using the ATPlite assay (PerkinElmer, 6016731) according to the manufacturer's instruction. [53][54][55] Cycloheximide-chase assay and immunoblotting For cycloheximide (CHX)-chase experiments, cells were pretreated with 1 mM MLN4924 for 12 h, washed twice with PBS, and treated with 50 mg/ml CHX (Sigma-Aldrich, C4859) in combination with 0.3 mM MLN4924 or DMSO for indicated time points. Cell lysates were prepared and analyzed by immunoblotting.…”

Section: Cell Lines and Reagentsmentioning

confidence: 99%

“…Antibodies against NEDD8 (2754), CDT1 (8064), Cell-cycle profile analysis Cell-cycle profile was evaluated by PI staining and FACS analysis as described previously. 17,55 Briefly, cells with the treatment of MLN4924 or DMSO were fixed in 70% ethanol at ¡20 C overnight, stained with PI (36 mg/mL; Sigma-Aldrich, P4170) containing RNase A (10 mg/mL; Sigma-Aldrich, R6513) at 37 C for 15 min, and analyzed for cell-cycle profile by CyAn ADP (Beckman Coulter). Data were analyzed with ModFit LT software (Verity Software House).…”

Section: Cell Lines and Reagentsmentioning

confidence: 99%

Targeting protein neddylation with an NEDD8-activating enzyme inhibitor MLN4924 induced apoptosis or senescence in human lymphoma cells

Wang

Luo

Pan

et al. 2015

Cancer Biology & Therapy

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These authors equally contributed to this work.Abbreviations: NAE, NEDD8-activating enzyme; CRL, cullin-RING E3 ligase; NHL, non-Hodgkin lymphoma; GCB-DLBCL, germinal-center B cell-like diffuse large B-cell lymphoma; SA-b-gal, senescence-associated b-galactosidase; IAP, inhibitor of apoptosis.Recent studies indicate that post-translational protein neddylation is required for the maintenance of cell viability in several lymphoma cell lines, while inhibition of the neddylation pathway with an NEDD8-activating enzyme (NAE) inhibitor MLN4924 induces apoptosis in lymphoma cells. However, the mechanism by which neddylation inhibition induces apoptosis in lymphoma cells has not been fully elucidated. Moreover, it is unknown whether neddylation inhibition triggers non-apoptotic cell-killing responses, such as cell senescence, in lymphoma cells. Here, we report that MLN4924 specifically inhibited protein neddylation, inactivated cullin-RING E3 ligase (CRL), the best-known neddylation substrate, and induced the accumulation of tumor-suppressive CRL substrates in lymphoma cells. Moreover, MLN4924 potently suppressed the growth of lymphoma cells by inducing G2 cell-cycle arrest, followed by apoptosis or senescence in a cell line-dependent manner. MLN4924-induced apoptosis was mediated by intrinsic apoptotic signaling with substantial up-regulation of pro-apoptotic Bik and Noxa as well as down-regulation of anti-apoptotic XIAP, c-IAP1 and c-IAP2, while senescence induction upon neddylation inhibition seemed dependent on the expression of tumor suppressor p21/p27. Together, these findings expand our understanding on how lymphoma cells respond to neddylation inhibition and support the development of neddylation inhibitors (e.g. MLN4924) for the treatment of lymphoma.

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“…8,18 Recent data have suggested that targeting neddylation with MLN4924 regulated the inflammatory functions of dendritic cells and macrophages, whereas inhibition of neddylation by knocking down Ubc12 mitigated CD4 T-cell responses. 1,19,20 Collectively, these studies point to an emerging role for neddylation in the regulation of immune cell subsets. However, the exclusive utilization of either the small molecule or the shRNA knockdown approaches does not rule out the possibility of potential off-target effects.…”

mentioning

confidence: 96%

SAG/Rbx2-Dependent Neddylation Regulates T-Cell Responses

Mathewson

Fujiwara

et al. 2016

The American Journal of Pathology

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Neddylation is a crucial post-translational modification that depends on the E3 cullin ring ligase (CRL). The E2-adapter component of the CRL, sensitive to apoptosis gene (SAG), is critical for the function of CRL-mediated ubiquitination; thus, the deletion of SAG regulates neddylation. We examined the role of SAG-dependent neddylation in T-cellemediated immunity using multiple approaches: a novel T-celle specific, SAG genetic knockout (KO) and chemical inhibition with small-molecule MLN4924. The KO animals were viable and showed phenotypically normal mature T-cell development. However, in vitro stimulation of KO T cells revealed significantly decreased activation, proliferation, and T-effector cytokine release, compared with WT. Using in vivo clinically relevant models of allogeneic bone marrow transplantation also demonstrated reduced proliferation and effector cytokine secretion associated with markedly reduced graft-versus-host disease. Similar in vitro and in vivo results were observed with the small-molecule inhibitor of neddylation, MLN4924. Mechanistic studies demonstrated that SAG-mediated effects in T cells were concomitant with an increase in suppressor of cytokine signaling, but not NF-kB translocation. Our studies suggest that SAG is a novel molecular target that regulates T-cell responses and that inhibiting neddylation with the clinically available small-molecule MLN4924 may represent a novel strategy to mitigate T-cellemediated immunopathologies, such as graft-versushost disease. (Am J Pathol 2016 http://dx

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Neddylation pathway regulates the proliferation and survival of macrophages

Cited by 40 publications

References 28 publications

MLN4924 suppresses lipopolysaccharide‑induced proinflammatory cytokine production in neutrophils in a dose‑dependent manner

MLN4924 suppresses lipopolysaccharide‑induced proinflammatory cytokine production in neutrophils in a dose‑dependent manner

Targeting protein neddylation with an NEDD8-activating enzyme inhibitor MLN4924 induced apoptosis or senescence in human lymphoma cells

SAG/Rbx2-Dependent Neddylation Regulates T-Cell Responses

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