Effect of resistance exercise contraction mode and protein supplementation on members of the STARS signalling pathway

Abstract

The striated muscle activator of Rho signalling (STARS) pathway is suggested to provide a link between external stress responses and transcriptional regulation in muscle. However, the sensitivity of STARS signalling to different mechanical stresses has not been investigated. In a comparative study, we examined the regulation of the STARS signalling pathway in response to unilateral resistance exercise performed as either eccentric (ECC) or concentric (CONC) contractions as well as prolonged training; with and without whey protein supplementation. Skeletal muscle STARS, myocardian-related transcription factor-A (MRTF-A) and serum response factor (SRF) mRNA and protein, as well as muscle cross-sectional area and maximal voluntary contraction, were measured. A single-bout of exercise produced increases in STARS and SRF mRNA and decreases in MRTF-A mRNA with both ECC and CONC exercise, but with an enhanced response occurring following ECC exercise. A 31% increase in STARS protein was observed exclusively after CONC exercise (P < 0.001), while pSRF protein levels increased similarly by 48% with both CONC and ECC exercise (P < 0.001). Prolonged ECC and CONC training equally stimulated muscle hypertrophy and produced increases in MRTF-A protein of 125% and 99%, respectively (P < 0.001). No changes occurred for total SRF protein. There was no effect of whey protein supplementation. These results show that resistance exercise provides an acute stimulation of the STARS pathway that is contraction mode dependent. The responses to acute exercise were more pronounced than responses to accumulated training, suggesting that STARS signalling is primarily involved in the initial phase of exercise-induced muscle adaptations.

Figure 1. Exercise-habituation and single-bout exercise protocols

A, exercise-habituation protocol. Three days prior to a harvesting of a pre-exercise-habituation biopsy, subjects refrained from exercise (−3 days). A pre-exercise-habituation biopsy (arrow) was then sampled from a randomly selected leg (0 days). Subjects were then allowed to recover for 3 days prior to initiation of the exercise-habituation period (3 days). The exercise-habituation period included 3 exercise sessions during a 7 day period, with 48 h interspacing each exercise session. Exercise sessions were conducted as 6 sets of 10 maximal eccentric or concentric repetitions at 30 deg s⁻¹ angular velocity, with 1 min of recovery between sets (10–13 days). Three days of recovery was then interspaced prior to sampling of post-exercise-habituation biopsies (B_ecc and B_conc;13 days). Another 3 days of recovery were then interspaced before completion of single-bout exercise trial (16 days). All pre- and post-exercise biopsies were sampled after an overnight fast. B, single-bout exercise protocol. After an overnight fast, the subjects reported to the laboratory at 08.30 h (−1 h). After 30 min of supine rest (−1/2 to 0 h), subjects completed a single-bout exercise session, conducted exactly as during exercise habituation. Immediately after exercise, a protein or a placebo supplement was ingested (0 h) and during the post-exercise recovery, biopsies were sampled at 1, 3 and 5 h from both eccentrically and concentrically working legs.

Figure 2. Effect of exercise habituation on STARS pathway mRNA and signalling proteins

Contraction mode-specific effects (ECC versus CONC) of exercise habituation (pre-habituation basal versus post-habituation basal) are shown as % change (mean ± SEM)from basal set to a fixed value of 100% for: STARS (A and B); MRTF-A (C and D); SRF and pSRF (E–G). Difference from basal level: *P < 0.05, ***P < 0.001.

Figure 3. Examples of Western blots

Examples of representative Western blots are shown for STARS, MRTF-A, SRF, pSRF and GAPDH for contraction mode-specific effects of exercise habituation and single-bout exercise.

Figure 4. Effect of single-bout exercise on STARS pathway mRNA and signalling proteins

Contraction mode-specific effects (ECC versus CONC) of single-bout exercise (post-habituation basal versus post-exercise recovery time points) are shown as % change (mean ± SEM) from basal set to a fixed value of 100% for: STARS (A and B); MRTF-A (C and D); SRF and pSRF (E–G). Difference from ECC or CONC basal level: *P < 0.05, **P < 0.01, ***P < 0.001. Difference between contraction modes, #P < 0.05, ##P < 0.01, ###P < 0.001.

Figure 5. Effect of single-bout exercise on SRF gene targets

Contraction mode-specific effects (ECC versus CONC) of single-bout exercise (post-habituation basal versus post-exercise recovery time points) are shown as % change (mean ± SEM) from basal set to a fixed value of 100% for: α actin (A); JUNB (B); IGF-1 (C) and CPT-1β (D). Difference from ECC or CONC basal level: *P < 0.05, **P < 0.01, ***P < 0.001. Difference between contraction modes: ##P < 0.01, ###P < 0.001.

Figure 6. Training-induced changes in STARS pathway protein expression

Contraction mode-specific effects (ECC versus CONC) of prolonged training (pre-habituation basal versus post-training basal) are shown as % change (mean ± SEM) from basal set to a fixed value of 100% for: STARS (A); MRTF-A (B); SRF (C). Different from basal level: *P < 0.05, **P < 0.01.