Recent studies have revealed various functions for the small ubiquitin-related modifier (SUMO) in diverse biological phenomena, such as regulation of cell division, DNA repair and transcription, in yeast and animals. In contrast, only a limited number of proteins have been characterized in plants, although plant SUMO proteins are involved in many physiological processes, such as stress responses, regulation of flowering time and defense reactions to pathogen attack. Here, we reconstituted the Arabidopsis thaliana SUMOylation cascade in Escherichia coli. This system is rapid and effective for the evaluation of the SUMOylation of potential SUMO target proteins. We tested the ability of this system to conjugate the Arabidopsis SUMO isoforms, AtSUMO1, 2, 3 and 5, to a model substrate, AtMYB30, which is an Arabidopsis transcription factor. All four SUMO isoforms tested were able to SUMOylate AtMYB30. Furthermore, SUMOylation sites of AtMYB30 were characterized by liquid chromatography-tandem mass spectrometry (LC-MS/MS) followed by mutational analysis in combination with this system. Using this reconstituted SUMOylation system, comparisons of SUMOylation patterns among SUMO isoforms can be made, and will provide insights into the SUMO isoform specificity of target modification. The identification of SUMOylation sites enables us to investigate the direct effects of SUMOylation using SUMOylation-defective mutants. This system will be a powerful tool for elucidation of the role of SUMOylation and of the biochemical and structural features of SUMOylated proteins in plants.
New Findings r What is the central question of this study?Does an acute bout of exercise alter vitamin D receptor expression in rat skeletal muscle? r What is the main finding and its importance?Resistance exercise but not endurance exercise increased intramuscular vitamin D receptor expression. Thus, resistance exercise may be an effective way to increase muscle vitamin D receptor expression.Vitamin D and vitamin D receptor (VDR) are involved in the maintenance of skeletal muscle mass and function. Although resistance exercise is well known to enhance muscle growth and improve muscle function, the effect of resistance exercise on VDR has been unclear. We investigated intramuscular VDR expression in response to an acute bout of resistance exercise or endurance exercise. Male adult Sprague-Dawley rats were subjected to either resistance exercise (isometrically exercised via percutaneous electrical stimulation for five sets of ten 3 s contractions, with a 7 s interval between contractions and 3 min rest intervals between sets) or endurance exercise (treadmill at 25 m min −1 for 60 min). Rats were killed immediately or 1, 3, 6 or 24 h after completion of the resistance or endurance exercise, and gastrocnemius muscles were removed. Non-exercised control animals were killed in a basal state (control group). Intramuscular VDR expression was significantly higher immediately after resistance exercise and elevated for 3 h after exercise compared with the control group (P < 0.05), and the resistance exercise significantly increased phosphorylated ERK1/2 and Mnk1 expression (P < 0.05), which may be associated with VDR expression, immediately after exercise. Additionally, intramuscular expression of cytochrome P450 27B1, an enzyme related to vitamin D metabolism, was significantly higher at 1 and 3 h after exercise (P < 0.05) compared with the control group. In contrast, endurance exercise had no effect on any of the measured proteins. Our results indicate that resistance exercise may be an efficient way to increase intramuscular VDR and related enzyme expression.
Skeletal muscle is an important organ for controlling the development of type 2 diabetes. We discovered Panax notoginseng roots as a candidate to improve hyperglycemia through in vitro muscle cells screening test. Saponins are considered as the active ingredients of ginseng. However, in the body, saponins are converted to dammarane-type triterpenes, which may account for the anti-hyperglycemic activity. We developed a method for producing a dammarane-type triterpene extract (DTE) from Panax notoginseng roots and investigated the extract's potential anti-hyperglycemic activity. We found that DTE had stronger suppressive activity on blood glucose levels than the saponin extract (SE) did in KK-A mice. Additionally, DTE improved oral glucose tolerance, insulin sensitivity, glucose uptake, and Akt phosphorylation in skeletal muscle. These results suggest that DTE is a promising agent for controlling hyperglycemia by enhancing glucose uptake in skeletal muscle.
Summary Insulin resistance reduces insulin-induced muscle protein synthesis and accelerates muscle protein degradation. Ginseng ingestion has been reported to improve insulin resistance through the phosphoinositide 3-kinase (PI3K)/Akt signaling pathway. We hypothesized that panaxatriol (PT) derived from ginseng in combination with aerobic exercise (EX) may further promote protein synthesis and suppress protein degradation, and subsequently maintain muscle mass through the amelioration of insulin resistance. KKAy insulin-resistant mice were divided into control, panaxatriol only (PT), exercise only (EX), and EXϩPT groups. EX and EXϩPT ran on the treadmill for 45 min at 15 m/min 5 d/wk for 6 wk. PT and EXϩPT groups were fed a standard diet containing 0.2% PT for 6 wk. Homeostasis model assessment for insulin resistance (HOMA-R) values was significantly improved after exercise for 6 wk. Moreover, EXϩPT mice showed improved HOMA-R as compared to EX mice. p70S6K phosphorylation after a 4 h fast was significantly higher in EX than in the non-exercise control, and it was higher in EXϩPT mice than in EX mice. Atrogin1 mRNA expression was significantly lower in EX than in the non-exercise control, and was significantly lowered further by PT treatment. EX and EXϩPT mice showed higher soleus muscle mass and cross-sectional area (CSA) of the soleus myofibers than control animals, with higher values noted for both parameters in EXϩPT than in EX. These results suggest that aerobic exercise and PT ingestion may contribute to maintain skeletal muscle mass through the amelioration of insulin resistance.
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