Stable Overexpression of the Constitutive Form of Heat Shock Protein 70 Confers Oxidative Protection

Chong, Kowit-Yu; Lai, Chen-Ching; Lille, Sean; Chang, Chawnshang; Su, Ching‐Yuan

doi:10.1006/jmcc.1997.0623

Cited by 104 publications

(61 citation statements)

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“…COS-1 cells are widely used in transfection experiments, including transfection with Hsp. 45,46 From our transfectants, 1 clone with a high expression of Hsp70 was selected. This clone, as expected, was protected against toxic stimuli such as H 2 O 2 , heat shock, and UV irradiation, in agreement with in vitro and in vivo data on the role of this protein in protecting from cellular damage.…”

Section: Discussionmentioning

confidence: 99%

Overexpression of Inducible Heat Shock Protein 70 in COS-1 Cells Fails to Protect From Cytotoxicity of Oxidized LDLs

Pirillo

Norata

Zanelli

et al. 2001

ATVB

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Abstract-Oxidized low density lipoproteins (OxLDLs) are believed to play a central role in atherogenesis and to possess a wide variety of biological properties; among them, OxLDLs are cytotoxic to cultured vascular cells in that they induce necrosis and apoptosis. Moreover, OxLDLs are known to induce the expression of heat shock protein 70 (Hsp70), a protein that protects cells from several cytotoxic stimuli. To determine whether Hsp70 can protect cells against OxLDL-induced cytotoxicity, COS-1 cells were transfected with a construct containing human Hsp70. A number of cell lines permanently expressing Hsp70 were obtained, 1 of which (cos-Hsp70/10, with high Hsp70 expression) was selected for further studies. Hsp70 overexpression protected cells from toxic stimuli, such as H 2 O 2 , UV irradiation, and heat shock, suggesting that the overexpressed protein was functional. When incubated with OxLDLs, however, the clone overexpressing Hsp70 showed a significant decrease in viability, as determined by the [ 3 H]adenine release assay (319.8Ϯ3.16% of control for transfected cells versus 217.6Ϯ6.08% for control cells exposed to 100 g protein/mL of OxLDL), 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay (12.5Ϯ0.9% versus 28.9Ϯ1.99% of control, respectively), and LDH release (48.4Ϯ0.04% versus 15.2Ϯ0.06% of control cells). The increased expression of Bax and the decreased expression of Bcl-2 (a proapoptotic and an antiapoptotic protein, respectively) in cos-Hsp70/10 cells and in control cells on incubation with OxLDLs suggested that overexpression of Hsp70 did not confer protection against apoptosis induced by OxLDLs. The analysis of nucleosome content and the nuclear staining with Hoechst 33258 confirmed this finding. These data suggest that overexpression of Hsp70 not only fails to protect COS-1 cells against OxLDL-induced apoptosis but rather confers a higher sensitivity to the cytotoxic action of these lipoproteins. Thus, the Hsp70 response, although induced by OxLDLs, cannot protect cells from lipoprotein toxicity. Key Words: low density lipoproteins Ⅲ apoptosis Ⅲ necrosis Ⅲ Bcl-2 Ⅲ Bax W hen exposed to stresses, such as high temperature or other toxic stimuli, cells react by synthesizing stress proteins, also named heat shock proteins (Hsps). [1][2][3][4][5][6] Hsps include constitutive and inducible forms. Constitutive Hsps play important biological roles, including facilitation of the correct folding of newly synthesized polypeptides 7 and their transport across cellular membranes 8 -10 ; inducible forms, expressed by cells under stress conditions, are believed to protect cellular proteins from denaturation. 11 Hsps show a very high degree of homology among different organisms 12 ; Hsp70, the major Hsp, is the most conserved throughout evolution 13 and plays a key role in protecting cells from environmental stresses. 14 In fact, overexpression of Hsp70 confers thermotolerance and increases cell survival in the presence of noxious stimuli. [15][16][17][18][19] Interestingly, the cellular conten...

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Section: Discussionmentioning

confidence: 99%

Overexpression of Inducible Heat Shock Protein 70 in COS-1 Cells Fails to Protect From Cytotoxicity of Oxidized LDLs

Pirillo

Norata

Zanelli

et al. 2001

ATVB

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“…HSP70s are fundamental in developmental processes and function in environmental stress, including heat, cold, heavy metal, water deficit, oxidative stress, wounding and so on [15][16][17][18][19]. In addition, HSP70 also are important in pathological processes.…”

Section: Introductionmentioning

confidence: 99%

Characterization of a wheat HSP70 gene and its expression in response to stripe rust infection and abiotic stresses

et al. 2010

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Members of the family of 70-kD heat shock proteins (HSP70 s) play various stress-protective roles in plants. In this study, a wheat HSP70 gene was isolated from a suppression subtractive hybridization (SSH) cDNA library of wheat leaves infected by Puccinia striiformis f. sp. tritici. The gene, that was designated as TaHSC70, was predicted to encode a protein of 690 amino acids, with a molecular mass of 73.54 KDa and a pI of 5.01. Further analysis revealed the presence of a conserved signature that is characteristic for HSP70s and phylogenetic analysis demonstrated that TaHSC70 is a homolog of chloroplast HSP70s. TaHSC70 mRNA was present in leaves of both green and etiolated wheat seedlings and in stems and roots. The transcript level in roots was approximately threefold less than in leaves but light-dark treatment did not charge TaHSC70 expression. Following heat shock of wheat seedlings at 40°C, TaHSC70 expression increased in leaves of etiolated seedlings but remained stable at the same level in green seedlings. In addition, TaHSC70 was differentially expressed during an incompatible and compatible interaction with wheat-stripe rust, and there was a transient increase in expression upon treatment with methyl jasmonate (MeJA) treatment. Salicylic acid (SA), ethylene (ET) and abscisic acid (ABA) treatments had no influence on TaHSC70 expression. These results suggest that TaHSC70 plays a role in stress-related responses, and in defense responses elicited by infection with stripe rust fungus and does so via a JA-dependent signal transduction pathway.

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“…We also observed that PKCι regulated oxidative sensitivity by positively regulating Hsc70, a pro-survival protein that confers oxidative protection (Chong et al 1998;Su et al 1999). We found that Hsc70 is involved in a complex formed by PKCι-p62/LC3 that functions in autophagic protein degradation (Behrends et al 2010).…”

Section: Discussionmentioning

confidence: 68%

“…3f). Hsc70 has been shown to protect cells from oxidative stress (Su et al 1999;Chong et al 1998), and we demonstrated that PKCι served as a positive regulator of Hsc70. Therefore, we hypothesized that PKCι also regulated this oxidative protection mediated by Hsc70.…”

Section: Pkcι Regulates Hsc70 In An Autophagy-dependent Mannermentioning

confidence: 64%

“…Hsc70 is also involved in Akt (Shiota et al 2010;Chen et al 2011) and NF-κB signaling (Lim et al 2008) and has been shown to regulate the Dbl oncoprotein (Kauppinen et al 2005) and Bim mRNA (Matsui et al 2007). Importantly, Hsc70 also regulates cell survival (Powers et al 2008) and confers protection from several forms of cellular stresses, such as viral infection (Yan et al 2010), metabolic stress (Williams et al 1993), and oxidative stress (Chong et al 1998;Dastoor and Dreyer 2000;Su et al 1999). Thus far, the mechanism of regulation of Hsc70 remains unclear, but it appears that its expression is regulated by KLF4 (Liu et al 2008).…”

Section: Introductionmentioning

confidence: 99%

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PKCι counteracts oxidative stress by regulating Hsc70 in an esophageal cancer cell line

Wang¹,

Yang²,

Yang³

et al. 2013

Cell Stress and Chaperones

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Using a glutathione S-transferase pull-down liquid chromatography-coupled tandem mass spectrometry approach and immunoprecipitation/immunoblot analysis, we found that heat shock cognate protein 70 (Hsc70) was involved in the complex formed by atypical protein kinase Cι (PKCι) and LC3 in the esophageal cancer cell line KYSE30. Further study indicated that Hsc70 was targeted by autophagic degradation, and knockdown of PKCι downregulated Hsc70 by promoting autophagy. PKCι knockdown sensitized cells to oxidative stress-induced apoptosis, whereas forced PKCι expression counteracted the oxidative stress-induced apoptosis via Hsc70.

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Stable Overexpression of the Constitutive Form of Heat Shock Protein 70 Confers Oxidative Protection

Cited by 104 publications

References 0 publications

Overexpression of Inducible Heat Shock Protein 70 in COS-1 Cells Fails to Protect From Cytotoxicity of Oxidized LDLs

Overexpression of Inducible Heat Shock Protein 70 in COS-1 Cells Fails to Protect From Cytotoxicity of Oxidized LDLs

Characterization of a wheat HSP70 gene and its expression in response to stripe rust infection and abiotic stresses

PKCι counteracts oxidative stress by regulating Hsc70 in an esophageal cancer cell line

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