Tribbles 3 Regulates Skeletal Muscle Mass in Fasting‐induced Atrophy

Abstract

Tribbles homolog 3 (TRB3) is a pseudokinase that has been found in multiple tissues, including skeletal muscle, in response to various stress stimuli, such as insulin resistance, nutrient deprivation, and endoplasmic reticulum stress. TRB3 has been mainly known as a negative regulator of Akt, which is an essential protein kinase to control protein synthesis and breakdown. Recently, our lab has demonstrated that TRB3 decreases protein synthesis and increases protein degradation in mouse skeletal muscle at basal state. However, it still remains to be determined whether TRB3 regulates skeletal muscle mass under atrophic conditions. Here, we hypothesized that TRB3 has a detrimental effect on skeletal muscle mass under 48hr fasting‐induced atrophy. In order to test the hypothesis, we utilized C57BL/6 wild type (WT) mice and fasted them for 48hr to induce skeletal muscle atrophy. Our fasting strategy reduced body weight by 20% and effectively decreased skeletal muscle mass by 15–25% compared to fed controls in multiple muscles (TA; 26%, EDL; 26%, GAS;14%, all P<0.05). In addition, TRB3 protein (60%) and mRNA (49%, P<0.05) expression was significantly increased in fasted muscles, which was associated with elevated atrogin−1 (750%, P<0.01) and MuRF‐1 (1840%, P<0.001) mRNA expression. Next, we analyzed Akt and its downstream proteins associated with protein synthesis and degradation. Phosphorylation of Akt (T308) was significantly decreased after 48hr fasting (37%, P<0.05) and FOXO1 (69%, P<0.01) and FOXO3a (47%, P=0.10) were activated whereas protein synthesis, mTOR and p70S6K1, was not affected. Next, we fasted muscle‐specific TRB3 transgenic (TG) and WT mice for 48hr in order to determine if TRB3 overexpression worsens the fasting‐induced atrophy. Body weight was significantly reduced by 22% after 48hr fasting in both WT and TG groups. Interestingly, we found a significant reduction in muscle mass in TG mice compared to WT (WT vs. TG in EDL; P<0.05, in GAS; P=0.07). Further, Akt and its downstream proteins were determined in EDL muscle. In TG mice, Akt phosphorylation (T308) was significantly decreased after fasting (18%, P<0.05) in line with suppressed S6K1 (10%, P=0.065) and activated FOXO1 (76%, P<0.001) and FOXO3a (32%, P=0.068) signaling. Additionally, protein synthesis rate was assessed by the incorporation of puromycin into nascent peptide chains. 48hr fasting significantly decreased protein synthesis rate in both WT and TG (P<0.01) but the degree of reduction was prominent in TG mice compared to WT (P=0.08). Lastly, we studied 48hr fasting‐induced atrophy in TRB3 knockout (KO) mice to determine if the deletion of TRB3 could prevent fasting‐induced atrophy. Body weight was significantly decreased by 20% in both WT and KO groups. Although both genotypes significantly reduced body weight, KO mice interestingly preserved ~10% more muscle mass in fasted TA, EDL, and SOL compared to fasted WT muscles (TA; 8.5%, EDL;15%, P<0.05, SOL;3.6%). These data suggest that TRB3 could be a key regulator of protein synthesis and breakdown through the Akt/mTOR/FOXO signaling under 48hr fasting‐induced atrophy.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.