Myosin light chain phosphorylation is required for peak power output of mouse fast skeletal muscle in vitro

Abstract

The skeletal myosin light chain kinase (skMLCK) catalyzed phosphorylation of the myosin regulatory light chain (RLC) is associated with potentiation of force, work, and power in rodent fast twitch muscle. The purpose of this study was to compare concentric responses of EDL from wild-type (WT) and skMLCK devoid (skMLCK) muscles at a range of shortening speeds (0.05 to 0.70 V ) around that expected to produce maximal power (in vitro, 25 °C) both before (unpotentiated) and after (potentiated) a potentiating stimulus (PS). When collapsed across all speeds tested, neither unpotentiated force, work, or power differed between genotypes (all data n = 10, P< 0.05). In contrast, although both genotypes displayed speed-dependent increases, these increases were greater for WT than skMLCK muscles. For example, when collapsed across the six fastest speeds we tested, both concentric force and power were increased 30-34 % in WT but only 15-17 % in skMLCK muscles. In contrast, at the two slowest speeds, these parameters were increased in WT but decreased in skMLCK muscles (8-10 and 7-9 %, respectively). Intriguingly, potentiation of concentric force and power was optimal near speeds producing maximal power in both genotypes. Because the PS elevated RLC phosphorylation above resting levels in WT but not in skMLCK muscles, our data suggest that skMLCK-catalyzed phosphorylation of the RLC is required for maximal concentric power output of mouse EDL muscle stimulated at high frequency in vitro.

Epinephrine augments posttetanic potentiation in mouse skeletal muscle with and without myosin phosphorylation

“…Thus, our data suggest that a more robust potentiation of concentric force is provided when the RLCs are phosphorylated compared to when they are not (see also Bowslaugh et al. ; Bunda et al. ; Gittings et al.…”

“…Indeed, at a time when concentric twitch force of skMLCK À/À muscles had returned to baseline in the control condition (i.e., 120 sec) wild-type muscles still displayed considerable potentiation. Thus, our data suggest that a more robust potentiation of concentric force is provided when the RLCs are phosphorylated compared to when they are not (see also Bowslaugh et al 2016;Bunda et al 2018;Gittings et al 2017;Gittings et al 2018;Vandenboom et al 2013). Because peak values for RLC phosphorylation were similar in both the untreated and treated conditions, it seems unlikely that epinephrine influences skMLCK activity in wild-type muscles.…”

Myosin phosphorylation potentiated steady state work output without altering contractile economy of mouse fast skeletal muscles

“…No fiber-type differences between WT and skMLCK −/− mice have been detected . For a recent discussion of issues related to genetic drift and our use of skMLCK −/− mice, see Bowslaugh et al (2016). Identical experiments were conducted on the two genotypes to investigate the contractile economy of skeletal muscles with and without the capacity (skMLCK expression) to phosphorylate the RLC.…”

“…Mechanical work (J kg −1 ) was determined by calculating the integral of the force-displacement plot for each stimulus period and normalizing it to muscle wet mass. The kinetics of force development (+dP/dt) and relaxation (−dP/dt) were sampled at the start and end of each stimulation period from the first derivative function of the active force trace, expressed in mN s −1 [as described previously in Bowslaugh et al (2016) and Gittings et al (2012)].…”

“…Muscles were frozen with liquid nitrogen-cooled tongs and stored at −80°C until needed. RLC phosphorylation analysis was conducted using urea-glycerol PAGE as previously described Bowslaugh et al, 2016;Xeni et al, 2011), in accordance with the method conceived and subsequently described by Dr Jim Stull and co-workers (Manning and Stull, 1982;Zhi et al, 2005). The primary myosin RLC antibody used in the current work was originally purified by Dr Jim Stull (Department of Physiology, University of Texas Southwestern Medical Center) and was provided as a gift.…”