Involvement of the p38/MK2 Pathway in MCLR Hepatotoxicity Revealed through MAPK Pharmacological Inhibition and Phosphoproteomics in HepaRG Cells

Katherine D Lynch; Dayne T Iverson; Namrata K Bachhav; Michael Ridge Call; Guihua Eileen Yue; Bhagwat Prasad; John D Clarke

doi:10.3390/ijms241311168

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Involvement of the p38/MK2 Pathway in MCLR Hepatotoxicity Revealed through MAPK Pharmacological Inhibition and Phosphoproteomics in HepaRG Cells

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Involvement of the p38/MK2 Pathway in MCLR Hepatotoxicity Revealed through MAPK Pharmacological Inhibition and Phosphoproteomics in HepaRG Cells

Katherine D Lynch, Dayne T Iverson, Namrata K Bachhav, Michael Ridge Call, Guihua Eileen Yue, Bhagwat Prasad and John D Clarke

International journal of molecular sciences, Vol.24(13), p.11168

07/06/2023

DOI: https://doi.org/10.3390/ijms241311168

PMID: 37446360

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Abstract

Chemical and Drug Induced Liver Injury - metabolism

Cytoskeleton - metabolism

Humans

Microcystins - toxicity

Mitogen-Activated Protein Kinases - metabolism

p38 Mitogen-Activated Protein Kinases - metabolism

Phosphoprotein Phosphatases - metabolism

Phosphorylation

Microcystin-leucine arginine (MCLR) is one of the most common and toxic microcystin variants, a class of cyanotoxins produced by cyanobacteria. A major molecular mechanism for MCLR-elicited liver toxicity involves the dysregulation of protein phosphorylation through protein phosphatase (PP) inhibition and mitogen-activated protein kinase (MAPK) modulation. In this study, specific pharmacological MAPK inhibitors were used in HepaRG cells to examine the pathways associated with MCLR cytotoxicity. SB203580 (SB), a p38 inhibitor, rescued HepaRG cell viability, whereas treatment with SP600125 (JNK inhibitor), MK2206 (AKT inhibitor), or N-acetylcysteine (reactive oxygen species scavenger) did not. Phosphoproteomic analysis revealed that phosphosites-which were altered by the addition of SB compared to MCLR treatment alone-included proteins involved in RNA processing, cytoskeletal stability, DNA damage response, protein degradation, and cell death. A closer analysis of specific proteins in some of these pathways indicated that SB reversed the MCLR-mediated phosphorylation of the necroptosis-associated proteins, the mixed lineage kinase domain-like protein (MLKL), receptor-interacting serine/threonine kinase 1 (RIP1), DNA damage response proteins, ataxia telangiectasia and Rad3-related kinase (ATR), and checkpoint kinase 1 (CHK1). Overall, these data implicate p38/MK2, DNA damage, and necroptosis in MCLR-mediated hepatotoxicity, and suggest these pathways may be targets for prevention prior to, or treatment after, MCLR toxicity.

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Title: Involvement of the p38/MK2 Pathway in MCLR Hepatotoxicity Revealed through MAPK Pharmacological Inhibition and Phosphoproteomics in HepaRG Cells
Creators: Katherine D Lynch - Department of Pharmaceutical Sciences, Washington State University, Spokane, WA 99202, USA
Dayne T Iverson - Washington State University Spokane
Namrata K Bachhav - Washington State University Spokane
Michael Ridge Call - Washington State University Spokane
Guihua Eileen Yue - Washington State University, Department of Pharmaceutical Sciences
Bhagwat Prasad - Washington State University, Department of Pharmaceutical Sciences
John D Clarke - Washington State University, Department of Pharmaceutical Sciences
Publication Details: International journal of molecular sciences, Vol.24(13), p.11168
Academic Unit: Department of Pharmaceutical Sciences
Grant note: R01ES032558 / NIEHS NIH HHS
Identifiers: 99901181236801842
Language: English
Resource Type: Journal article