Endurance training lowers ribosome density despite increasing ribosome biogenesis markers in rodent skeletal muscle

Abstract

Objective: The purpose of this study was to examine if: (a) high sugar/high fat Western diet (WD)-feeding affects skeletal muscle ribosome biogenesis markers in hyperphagic, diabetic-prone Otsuka Long-Evans Tokushima Fatty (OLETF) rats, and (b) 12 weeks of treadmill training rescued potential detriments that WD feeding exerted on these markers.

Methods: Eight week-old male OLETF rats were fed a low-fat control diet (O-CON, n = 10) or high/sucrose/cholesterol Western diet (WD). At weeks 20-32 of age, WD-fed rats were divided into WD sedentary (O-WD/SED, n = 16), or WD treadmill trained (5 days/week, 60 min/day) (O-WD/EX, n = 10) conditions.

Results: Interestingly, total RNA (i.e., ribosome density) was 2.3-fold greater in O-WD/SED versus O-WD/EX rats (p = 0.003) despite levels of upstream binding factor protein, RNA polymerase I protein and pre-45S rRNA being greater in O-WD/EX rats. Ribophagy (USP10 and G3BP1) and TRAMP-exosome rRNA degradation pathway (EXOSC10 and SKIV2L2) proteins were assayed to determine if these pathways were involved with lower ribosome density in O-WD/EX rats. While USP10 was higher in O-CON versus O-WD/SED and O-WD/EX rats (p < 0.001 and p < 0.001, respectively), G3BP1, EXOSC10 and SKIV2L2 did not differ between groups. Nop56 and Ncl mRNAs, ribosome assembly markers, were highest in O-WD/EX rats. However, Fbl mRNA and 28S rRNA, downstream ribosome processing markers, were lowest in O-WD/EX rats. Collectively these data suggest that, in WD-fed rats, endurance training increases select skeletal muscle ribosome biogenesis markers. However, endurance training may reduce muscle ribosome density by interfering with rRNA processing and/or export through mechanisms independent of ribophagy or rRNA degradation.

Keywords: Endurance training; Ribophagy; Ribosome biogenesis; Western diet; rRNA degradation.

Fig. 1

Effects of diet and exercise on markers of total ribosome content and function. The effects of the intervention on skeletal muscle total RNA levels (i.e., ribosome content) (a), myofibrillar protein levels (b), muscle soluble protein levels (c), myofibrillar (MF): total RNA ratio as a surrogate of ribosome function (d) and muscle soluble protein: total RNA ratio as an additional surrogate of ribosome function (e). All data are presented as mean ± SD (n = 10–16 rats per group), group mean values are presented within each bar, 95% confidence intervals are presented in parentheses within each bar, and bars with different superscript letters indicate between-treatment differences (p < 0.05). Due to limited tissue, only a subset of animals were assayed for myofibrillar protein (O-CON n = 7, O-WD/SED n = 14, O-WD/EX n = 8). O-CON OLETF rats consuming a control diet during weeks 20–32 and were not treadmill-trained, O-WD/SED OLETF rats consuming a Western diet during weeks 20–32 and were not treadmill-trained, O-WD/EX OLETF rats consuming a Western diet during weeks 20–32 and were treadmill-trained 5 days/week

Fig. 2

Effects of diet and exercise on markers of ribosome biogenesis. The effects of the intervention on c-Myc protein levels (a), UBF protein levels (b), RNA Pol I protein levels (c) and pre-45S rRNA transcript levels (d). e Representative images of Western blotting targets. All data are presented as mean ± SD (n = 10–16 rats per group), group mean values are presented within each bar, 95% confidence intervals are presented in parentheses within each bar, and bars with different superscript letters indicate between-treatment differences (p < 0.05). O-CON OLETF rats consuming a control diet during weeks 20–32 and were not treadmill-trained, O-WD/SED OLETF rats consuming a Western diet during weeks 20–32 and were not treadmill-trained, O-WD/EX OLETF rats consuming a Western diet during weeks 20–32 and were treadmill-trained 5 days/week

Fig. 3

Effects of diet and exercise on markers of ribosome degradation. The effects of the intervention on USP10 protein levels (a) and G3BP1 protein levels (b) (both markers being critical for ribophagy), as well as EXOSC10 protein levels (c) and SKIV2L2 protein levels (d) (both markers being the TRAMP exosome rRNA degradation pathway). e Representative images of Western blotting targets. All data are presented as mean ± SD (n = 10–16 rats per group), group mean values are presented within each bar, 95% confidence intervals are presented in parentheses within each bar, and bars with different superscript letters indicate between-treatment differences (p < 0.05). O-CON OLETF rats consuming a control diet during weeks 20–32 and were not treadmill-trained, O-WD/SED OLETF rats consuming a Western diet during weeks 20–32 and were not treadmill-trained, O-WD/EX OLETF rats consuming a Western diet during weeks 20–32 and were treadmill-trained 5 days/week

Fig. 4

Effects of diet and exercise on the expression of genes associated with rRNA processing and ribosome export. The effects of the intervention on Nop56 mRNA levels (a) and Ncl mRNA levels (b) (both genes being involved in rRNA processing and ribosome assembly), Npm1 mRNA levels (c) and Fbl mRNA levels (d) (genes involved with nuclear ribosome export), and 28S rRNA transcript levels (e) and 18S rRNA transcript levels (f) (both markers being indicative of downstream rRNA processing from the pre-45S transcript). All data are presented as mean ± SD (n = 10–16 rats per group), group mean values are presented within each bar, 95% confidence intervals are presented in parentheses within each bar, and bars with different superscript letters indicate between-treatment differences (p < 0.05). O-CON OLETF rats consuming a control diet during weeks 20–32 and were not treadmill-trained, O-WD/SED OLETF rats consuming a Western diet during weeks 20–32 and were not treadmill-trained, O-WD/EX OLETF rats consuming a Western diet during weeks 20–32 and were treadmill-trained 5 days/week