Endoplasmic Reticulum Stress Markers and Ubiquitin-Proteasome Pathway Activity in Response to a 200-km Run

KIM, H. J., C. JAMART, L. DELDICQUE, G.-L. AN, Y. H. LEE, C. K. KIM, J.-M. RAYMACKERS, and M. FRANCAUX. Endoplasmic Reticulum Stress Markers and Ubiquitin-Proteasome Pathway Activity in Response to a 200-km Run. Med. Sci. Sports Exerc., Vol. 43, No. 1, pp. 18-25, 2011. Purpose: This study investigated whether a 200-km run modulates signaling pathways implicated in cellular stress in skeletal muscle, with special attention paid to the endoplasmic reticulum (ER) stress and to the activation of the ubiquitin-proteasome pathway. Methods: Eight men ran 200 km (28 h 03 min + 2 h 01 min). Two muscle biopsies were obtained from the vastus lateralis muscle 2 wk before and 3 h after the race. Mitogen-activated protein kinase, ubiquitin-proteasome pathway, ER stress, inflammation, and oxidative stress markers were assayed by Western blot analysis or by quantitative real-time polymerase chain reaction. Chymotrypsin-like activity of the proteasome was measured by a fluorimetric assay. Results: Phosphorylation states of extracellular signal-related kinase 1/2 (+401% +/- 173.8%, P = 0.027) and c-Jun N-terminal (+149% +/- 61.9%, P = 0.023) increased after the race, whereas p38 phosphorylation remained unchanged. Increases in BiP (+235% +/- 94.7%, P = 0.021) and in the messenger RNA level of total (+138% +/- 31.2%, P = 0.002) and spliced X-box binding protein 1 (+241% +/- 53.3%, P = 0.001) indicated the presence of ER stress. Transcripts of inflammatory markers interleukin-6 (+403% +/- 96.1%, P = 0.002) and tumor necrosis factor-alpha (+233% +/- 58.4%, P = 0.003) as well as oxidative stress markers metallothionein 1F (+519% +/- 258.3%, P = 0.042), metallothionein 1H (+666% +/- 157.5%, P = 0.002), and nicotinamide adenine dinucleotide phosphate-oxidase (NADPH oxidase) (+162% +/- 60.5%, P = 0.016) were increased. The messenger RNA level of the ubiquitin ligases muscle-specific RING finger 1 (+583% +/- 244.3%, P = 0.024) and muscle atrophy F-box (+249% +/- 83.8%, P = 0.011) and the C2 proteasome subunit (+116% +/- 40.6%, P = 0.012) also increased. Surprisingly, the amount of ubiquitin-conjugated proteins and the chymotrypsin-like activity of the proteasome were decreased by 20% +/- 8.3% (P = 0.025) and 21% +/- 4.4% (P = 0.001), respectively. The expression of ubiquitin-specific protease 28 deubiquitinase was increased (+81% +/- 37.9%, P = 0.034). Conclusions: In the skeletal muscle, a 200-km run activates the expression of ubiquitin ligases muscle-specific RING finger 1 and muscle atrophy F-box as well as various cellular stresses, among which are ER stress, oxidative stress, and inflammation. Meanwhile, compensatory mechanisms seem also triggered: the unfolded protein response is up-regulated, and the chymotrypsin-like activity of the proteasome is repressed.