Endoplasmic reticulum stress markers and ubiquitin–proteasome pathway activity in response to a 200-km run.
OBJECTIVE
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-α (+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.
Kim
Hyo Jeong
HJ
Human Physiology, Korea National Sport University, Seoul, South Korea.
Jamart
Cécile
C
Deldicque
Louise
L
An
Gang-li
GL
Lee
Yoon Hee
YH
Kim
Chang Keun
CK
Raymackers
Jean-marc
JM
Francaux
Marc
M
D000328:Adult; D015415:Biomarkers / Q000378:metabolism*; D001706:Biopsy; D015153:Blotting, Western; D004268:DNA-Binding Proteins / Q000096:biosynthesis; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D005470:Fluorometry; D006801:Humans; D007249:Inflammation / Q000378:metabolism; D020935:MAP Kinase Signaling System / Q000502:physiology; D008297:Male; D008875:Middle Aged; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009124:Muscle Proteins / Q000096:biosynthesis; D018384:Oxidative Stress / Q000502:physiology; D063150:Polycomb Repressive Complex 1; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D011494:Protein Kinases / Q000378:metabolism; D052097:Quadriceps Muscle / Q000378:metabolism* / Q000473:pathology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012420:Running / Q000502:physiology*; D044843:SKP Cullin F-Box Protein Ligases / Q000096:biosynthesis; D015398:Signal Transduction / Q000502:physiology*; D013312:Stress, Physiological / Q000502:physiology*; D043743:Ubiquitin-Protein Ligase Complexes / Q000378:metabolism*
D015415:Biomarkers; D004268:DNA-Binding Proteins; D009124:Muscle Proteins; D043743:Ubiquitin-Protein Ligase Complexes; C440907:FBXO32 protein, human; D063150:Polycomb Repressive Complex 1; C080355:RING1 protein, human; D044843:SKP Cullin F-Box Protein Ligases; D011494:Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D046988:Proteasome Endopeptidase Complex