Pro-oxidative DEP chemicals induce heat shock proteins and an unfolding protein response in a bronchial epithelial cell line as determined by DIGE analysis.
Ambient particulate matter (PM) induces adverse health effects through the ability of pro-oxidative chemicals to induce the production of oxygen radicals and oxidant injury. Utilizing a proteomics strategy involving 2-D DIGE, immunoblotting, and real-time PCR, we demonstrate that organic diesel exhaust particle (DEP) chemicals induce an unfolding protein response (UPR) and proinflammatory effects in the human bronchial epithelial cell line, BEAS-2B. DIGE and MS showed the induction of at least 14 proteins, among which heat shock protein 70 (HSP70), HSP40, TPR2, and T-complex protein 1 (zeta-subunit) are known to play a role in the UPR. Demonstrating increased HSP70 mRNA expression and nuclear translocation of HSF1, the key transcription factor responsible for HSP expression, further strengthened this notion. Immunoblotting demonstrated increased expression of ATF4, an ER stress-associated transcriptional enhancer responsible for differential protein translation under conditions of ER stress. Finally, the DEP extract induced the expression of IL-6 and IL-8 in the culture supernatant. The role of oxidative stress was demonstrated further by response subtraction in the presence of the thiol antioxidant, N-acetyl cysteine. Our data suggest that pro-oxidative DEP chemicals induce protein unfolding/misfolding that lead to UPR and proinflammatory effects in a cell type that is targeted by PM in the lung.
Jung
EunMi Juliana
EJ
Department of Medicine, Division of Clinical Immunology and Allergy, and David Geffen School of Medicine, University of California-Los Angeles, CA 90095, USA.
Avliyakulov
Nuraly K
NK
Boontheung
Pinmanee
P
Loo
Joseph A
JA
Nel
Andre E
AE
D051701:Activating Transcription Factor 4 / Q000096:biosynthesis / Q000187:drug effects; D001483:Base Sequence; D001980:Bronchi / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D002460:Cell Line; D017931:DNA Primers / Q000235:genetics; D015180:Electrophoresis, Gel, Two-Dimensional; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism; D018840:HSP70 Heat-Shock Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000187:drug effects / Q000235:genetics; D006360:Heat-Shock Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000187:drug effects* / Q000235:genetics; D006801:Humans; D018836:Inflammation Mediators / Q000378:metabolism; D016877:Oxidants / Q000633:toxicity; D018384:Oxidative Stress / Q000187:drug effects; D040081:Protein Array Analysis; D011489:Protein Denaturation / Q000187:drug effects; D040901:Proteomics; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D001335:Vehicle Emissions / Q000633:toxicity*
C497109:ATF4 protein, human; D017931:DNA Primers; D018840:HSP70 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D018836:Inflammation Mediators; D016877:Oxidants; D012333:RNA, Messenger; D001335:Vehicle Emissions; D051701:Activating Transcription Factor 4