DJ-1 interacts with RACK1 and protects neurons from oxidative-stress-induced apoptosis

The early release of adenosine following traumatic brain injury (TBI) suppresses seizures and brain inflammation; thus, it is important to elucidate the cellular sources of adenosine following injurious stimuli triggered by TBI so that therapeutics for enhancing the early adenosine-release response can be optimized. Using mass spectrometry with 13C-labelled standards, we investigated in cultured rat neurons, astrocytes, and microglia the effects of oxygen-glucose deprivation (OGD; models energy failure), H2O2 (produces oxidative stress), and glutamate (induces excitotoxicity) on intracellular and extracellular levels of 5′-AMP (adenosine precursor), adenosine, and inosine and hypoxanthine (adenosine metabolites). In neurons, OGD triggered increases in intracellular 5′-AMP (2.8-fold), adenosine (2.6-fold), inosine (2.2-fold), and hypoxanthine (5.3-fold) and extracellular 5′-AMP (2.2-fold), adenosine (2.4-fold), and hypoxanthine (2.5-fold). In neurons, H2O2 did not affect intracellular or extracellular purines; yet glutamate increased intracellular adenosine, inosine, and hypoxanthine (1.7-fold, 1.7-fold, and 1.6-fold, respectively) and extracellular adenosine, inosine, and hypoxanthine (2.9-fold, 2.1-fold, and 1.6-fold respectively). In astrocytes, neither H2O2 nor glutamate affected intracellular or extracellular purines, and OGD only slightly increased intracellular and extracellular hypoxanthine. Microglia were unresponsive to OGD and glutamate, but were remarkably responsive to H2O2, which increased intracellular 5′-AMP (1.6-fold), adenosine (1.6-fold), inosine (2.1-fold), and hypoxanthine (1.6-fold) and extracellular 5′-AMP (5.9-fold), adenosine (4.0-fold), inosine (4.3-fold), and hypoxanthine (1.9-fold). Conclusion: Under these particular experimental conditions, cultured neurons are the main contributors to adenosine production/release in response to OGD and glutamate, whereas cultured microglia are the main contributors upon oxidative stress. Developing therapeutics that recruit astrocytes to produce/release adenosine could have beneficial effects in TBI.

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“…), induces oxidative stress (Ma et al . ) and represents a reasonable pathophysiological level of H 2 O 2 (Lei et al . ).…”

Section: Discussionmentioning

confidence: 99%

“…), and Ma and coworkers reported that 50 μM H 2 O 2 induces oxidative stress in DIV6 cortical neurons (Ma et al . ). Therefore, in this study, we selected 40 μM H 2 O 2 as the best approximation of a pathophysiological relevant and oxidative stress‐inducing level of H 2 O 2 .…”

Section: Methodsmentioning

confidence: 97%

Adenosine production by brain cells

Jackson

Kotermanski

Menshikova

et al. 2017

Journal of Neurochemistry

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“…RACK1 was originally identified as an intercellular scaffold protein of the Trp-Asp (WD) repeat protein family (23). RACK1 can recruit and bind many kinases and receptors, such as PKC and DJ-1, thereby impacting a wide range of signal transduction pathways (24,25). RACK1 plays an important role in multiple cellular functions, including cell growth, migration and differentiation (26,27).…”

Section: Introductionmentioning

confidence: 99%

RACK1 modulates apoptosis induced by sorafenib in HCC cells by interfering with the IRE1/XBP1 axis

Zhou

Guo

et al. 2015

Oncology Reports

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Abstract. Sorafenib is one of the preferred drugs for the treatment of advanced primary hepatocellular carcinoma (HCC). However, its side-effects and acquired resistance limit its use. The unfolded protein response (UPR) induced by chemotherapeutics has been demonstrated to be required for tumor cells to maintain malignancy and therapy resistance. Activation of the IRE1/XBP1 pathway during the UPR is important for tumor survival under pathophysiological conditions. In the present study, we found that the UPR was activated and RACK1 was overexpressed in three human HCC cell lines and in HCC samples. Activation of the IRE1/XBP1 signaling pathway plays a protective role when HCC cells encounter endoplasmic reticulum (ER) stress due to in vitro sorafenib treatment. We then found that the interaction between IRE1 and RACK1 was essential for the activation of IRE1 signaling in sorafenibtreated cells. Exogenous overexpression of RACK1 enhanced the phosphorylation level of IRE1 and increased XBP1 mRNA splicing activity, which protected the HCC cells from sorafenib-induced apoptosis. However, the re-expression of RACK1 led HCC cells to regain susceptibility to sorafenibinduced apoptosis. Taken together, the present study suggests that the RACK1/IRE1 complex may contribute to activation of the UPR in HCC cells. Targeting RACK1 in combination with sorafenib administration is a potential strategy for clinical trials of advanced HCC treatment.

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“…RACK1 was originally identified to bind and activate protein kinase C and is now recognized as a multi-functional scaffold protein (11,12). Evidence has indicated that RACK1 protects from oxidative stress-induced cell death in various types of cells, including fission yeasts (13), shrimp cells (14), neurons (15), HeLa cells (16) and HL60 cells (17). However, such a role for RACK1 has not been reported in HCC cells to the best of our knowledge.…”

Section: Introductionmentioning

confidence: 76%

RACK1 promotes hepatocellular carcinoma cell survival via CBR1 by suppressing TNF-α-induced ROS generation

Zhou¹,

Cao²,

Zhao³

et al. 2016

Oncology Letters

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Abstract. It has been reported that intracellular accumulation of reactive oxygen species (ROS) has a significant role in tumor necrosis factor (TNF)-α-induced cell apoptosis and necrosis; however, the key molecules regulating ROS generation remain to be elucidated. The present study reports that knockdown of endogenous receptor for activated C kinase 1 (RACK1) increases the intracellular ROS level following TNF-α or H 2 O 2 stimulation in human hepatocellular carcinoma (HCC) cells, leading to promotion of cell death. Carbonyl reductase 1 (CBR1), a ubiquitous nicotinamide adenine dinucleotide phosphate-dependent enzyme, is reported to protect cells from ROS-induced cell damage. The present study reports that RACK1 is a regulator of CBR1 that interacts with and sustains the protein stability of CBR1. Overexpression of CBR1 reverses the enhanced cell death due to RACK1 knockdown. Taken together, the results of the present study suggest that RACK1 protects HCC cells from TNF-α-induced cell death by suppressing ROS generation through interacting with and regulating CBR1. IntroductionEscape from tumor necrosis factor (TNF)-α-induced cell apoptosis and necrosis is of importance in tumor development (1-3).This process is regulated by a number of intracellular signaling pathways, including c-jun N-terminal kinase (JNK) and IκB kinase (IKK), as well as reactive oxygen species (ROS) (4,5). Extensive studies have indicated that reduced levels of oxidant stress and ROS promote malignant transformation and oncogenic growth in hepatocellular carcinoma (HCC) cells (6-9). However, the key molecules regulating ROS in HCC remain to be elucidated. It has been reported that scaffolding protein receptor for activated C kinase 1 (RACK1) enhances JNK activation in HCC, leading to promotion of the malignant growth of HCC (10). Therefore, it may be assumed that RACK1 affects other aspects of HCC. RACK1 was originally identified to bind and activate protein kinase C and is now recognized as a multi-functional scaffold protein (11,12). Evidence has indicated that RACK1 protects from oxidative stress-induced cell death in various types of cells, including fission yeasts (13), shrimp cells (14), neurons (15), HeLa cells (16) and HL60 cells (17). However, such a role for RACK1 has not been reported in HCC cells to the best of our knowledge. In the present study, it was demonstrated that RACK1 knockdown leads to increased cell death in TNF-α-treated HCC cells in the presence of cycloheximide (CHX), a protein synthesis inhibitor. Subsequently, it was observed that RACK1 knockdown promotes intracellular ROS accumulation upon TNF-α or H 2 O 2 stimulation. A combination of co-immunoprecipitation (co-IP) and mass spectrometry analysis indicated that carbonyl reductase 1 (CBR1), a ubiquitous nicotinamide adenine dinucleotide phosphate-dependent enzyme, acts as a RACK1-interacting partner in HCC cells. CBR1 has been reported to provide protection from ROS-induced cellular damage in HCC and leukemia (4,18), which suggests that CBR1 serves a role in c...

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DJ-1 interacts with RACK1 and protects neurons from oxidative-stress-induced apoptosis

Cited by 29 publications

References 66 publications

Adenosine production by brain cells

Adenosine production by brain cells

RACK1 modulates apoptosis induced by sorafenib in HCC cells by interfering with the IRE1/XBP1 axis

RACK1 promotes hepatocellular carcinoma cell survival via CBR1 by suppressing TNF-α-induced ROS generation

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