ER-Stress Markers and Ubiquitin-Proteasome Pathway Activity in Response to 200-Km Run.

PURPOSE:: This study investigated whether a 200-km run modulates signalling 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 (UPP). METHODS:: Eight men ran 200-km (28h 03min+/-2h 01min). Two muscle biopsies were obtained from the vastus lateralis muscle, 2 weeks prior to and 3 hours after the race. MAPK, UPP, ER-stress, inflammation and oxidative stress markers were assayed by western blotting or by RT-qPCR. Chymotrypsin-like activity of the proteasome was measured by a fluorimetric assay. RESULTS:: Phosphorylation states of ERK1/2 (+401+/-173.8%, P=0.027) and JNK (+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 mRNA level of total (+138+/-31.2%, P=0.002) and spliced XBP1 (+241+/-53.3%, P=0.001) indicated the presence of ER-stress. Transcripts of inflammatory markers -IL-6 (+403+/-96.1%, P=0.002) and TNF- square (+233+/-58.4%, P=0.003)- as well as oxidative stress markers -MT1F (+519 +/- 258.3%, P=0.042), MT1H (+666 +/- 157.5 %, P=0.002) and NADPHox (+162 +/- 60.5%, P=0.016)- were increased. mRNA level of the ubiquitin-ligases MuRF-1 (+583+/-244.3%; P=0.024) and MAFbx (+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 USP28 deubiquitinase was increased (+81+/-37.9%; P=0.034). CONCLUSION:: In skeletal muscle, a 200-km run activates the expression of ubiquitin-ligases MuRF-1 and MAFbx as well as various cellular stresses amongst which 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