title: Lon protease and eiF2α are involved in acute, but not prolonged, antiretroviral induced stress response in HepG2 cells.
abstract: Lon protease, an ATP dependent mitochondrial protease, is important in mitochondrial protein maintenance. Disruption of protein homeostasis and mitochondrial dysfunction is associated with lipodystrophy, metabolic syndrome and accelerated aging, and are commonly observed in patients on long term antiretroviral therapy. Sirtuin 3 (SIRT3) is a post-translational regulator of Lon and regulates antioxidant response. We previously showed the nucleoside analogues (NRTIs), Zidovudine (AZT; 7.1 μM), Stavudine (d4T; 4 μM), and Tenofovir (TFV; 1.2 μM) induced oxidative stress and mitochondrial dysfunction in human hepatoma (HepG2) cells at 24 h (h) and 120 h. We conducted a mitochondrial proteomic assessment of homeostasis in the same model, using the same NRTIs. Protein expression of Lon, SIRT3, heat shock protein (HSP) 60, phospho-eukaryotic translation initiation factor 2α (p-eIF2α; Ser51) and phospho-c-jun N-terminal kinase (p-JNK; Thr183/Tyr185) were quantified by western blots. The data showed all stress responses were significantly increased in HepG2 cells by all antiretroviral drugs at 24 h (p < 0.0001); however, at 120 h, a significant depletion in the ATP-dependent proteins Lon (p = 0.00013) and HSP60 (p < 0.0001) was observed. Proteins initiated by endoplasmic reticulum stress: p-eIF2α (p = 0.001) and p-JNK (p = 0.0029), were significantly reduced following prolonged treatment. SIRT3 was maintained at elevated levels in the treated cells following prolonged exposure (p < 0.001). We conclude that the ATP dependent proteins are more relevant to acute toxicity, while SIRT3 confers protection over prolonged periods of toxicity.
authors.affiliation: 3rd Floor George Campbell Building, Discipline of Medical Biochemistry, Howard College, University of KwaZulu Natal, King George Avenue, Durban, 4041, South Africa. Electronic address: 208509769@ukzn.stu.az.za.
authors.forename: Savania
authors.initials: S
authors.lastname: Nagiah
authors.affiliation: Department of Physiology, School of Medicine, Prinshof Campus, Dr Savage Road, 0083, Prinshof, Pretoria, Gauteng, South Africa. Electronic address: phulukdaree@up.ac.za.
authors.forename: Alisa
authors.initials: A
authors.lastname: Phulukdaree
authors.affiliation: Discipline of Medical Biochemistry, School of Laboratory Medicine and Medical Sciences, College of Health Science, University of KwaZulu Natal, Durban, South Africa; 3rd Floor George Campbell Building, Discipline of Medical Biochemistry, Howard College, University of KwaZulu Natal, King George Avenue, Durban, 4041, South Africa. Electronic address: CHUTUR@ukzn.ac.za.
authors.forename: Anil A
authors.initials: AA
authors.lastname: Chuturgoon
mesh_terms: D044966:Anti-Retroviral Agents / Q000009:adverse effects*; D018834:Chaperonin 60 / Q000378:metabolism; D015852:Eukaryotic Initiation Factor-2 / Q000378:metabolism*; D056945:Hep G2 Cells; D022781:Hepatocytes / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D024101:Mitochondrial Proteins / Q000378:metabolism; D018384:Oxidative Stress / Q000187:drug effects; D049070:Protease La / Q000378:metabolism*; D056566:Sirtuin 3 / Q000378:metabolism; D018119:Stavudine / Q000009:adverse effects*; D013312:Stress, Physiological / Q000187:drug effects; D000068698:Tenofovir / Q000009:adverse effects*; D015215:Zidovudine / Q000009:adverse effects*
chemical_list: D044966:Anti-Retroviral Agents; D018834:Chaperonin 60; D015852:Eukaryotic Initiation Factor-2; C060293:HSPD1 protein, human; D024101:Mitochondrial Proteins; D015215:Zidovudine; D000068698:Tenofovir; D018119:Stavudine; D049070:Protease La; D056566:Sirtuin 3
keywords: Lon; Mitochondrial stress; NRTI; SIRT3; eIF2α