Different Regulation Mechanisms Of FoxO Transcription Factors And NF-?B Pathway In Different Type Skeletal Muscles In Hibernating Daurian Ground Squirrels (Spermophilus Dauricus)

Atrogin-1 (also named MAFbx/FBXO32) and MuRF-1 (also named Trim63) are two muscle specific E3 ubiquitin ligases identified as differentially up-regulated following disuse-induced atrophy. During disuse, the increase of intracellular activited, non-phosphorylated FoxO transcription factors (FoxO1 and Fox03a) promote increase of the atrogin-1 and MuRF-1 mRNA expressions, and NF-?B pathway (IKK?/p50) is only involved in the regulation of MuRF-1 mRNA expression. Moreover, the extent of muscle atrophy is effect by muscle types. Hibernation showed a minmier or limited atrophy after prolonged hibernation inactivity. However, whether atrogin-1 and MuRF-1 expressions are regulated by FoxO transcription factors and NF-?B pathway in hibernating manmmals, and whether the regulation of FoxO transcription factors and NF-?B pathway on atrogin-1 and MuRF-1 mRNA expressions was effect by muscle types is still unclear. In this study, we determine the changes of atrogin-1, MuRF-1 and upstream FoxO transcript factors and NF-?B pathway in different type skeletal muscles (the slow twitch soleus muscle, the relative mixed gastrocnemius muscle and the fast twitch extensor digitorum longus muscle) and further explored the molecular mechanism involved in preventing muscle atrophy in different muscle types in mammalian hibernators.Total FoxOs and p-FoxOs were repressed the relative expressions in intracellular total FoxOs and phosphorylated FoxOs, respectively. P-FoxOs/total FoxOs were repressed the intracellular phosphorylation level in FoxOs, and negatively related to the transcriptional activities of atrogin-1 and MuRF-1. The results showed that hibernating ground squirrels inhibited the increase of FoxO transcript factors and phosphonation, maintained the intracellular phosphorylated level of FoxOs, and finally inhibited the increase of atrogin-1 mRNA experiment in soleus muscle; the increased phosphonation inhibited the phosphonation of FoxOs, and inhibited atrogin-1 mRNA experiment increased in gastrocnemius muscle; the phosphonation of FoxO3a unchanged and the increased FoxOs in total protein expressions were not enough to lead to the increase of phosphorylation in FoxOs in extensor digitorum longus muscle, both of which were benefit to the mainteinance of atrogin-1 mRNA expersions. These results indicated that different mechanisms in inhibition of the FoxOs' transcriptional activities of atrogin-1 and protein synthesis might be exsited in different type skeletal muscles of ground squirrels during prolonged hibernation. Moreover, the changes of FoxOs in translation level were in accord with atrogin-1 mRNA experissions. Ground squirrels inhibited the increase of atrogin-1 mRNA expressions via increasing the phosphorylation of FoxOs in gastrocnemius and extensor digitorum longus muscles awaking from months of hibernation.NF-?B pathway (IKK?/p50 pathway) were determined in mRNA and protein levels in ground squirrels' gastrocnemius muscle. The results from the study and the previous findings in our research group showed that hibernating ground squirrels inhibited MuRF-1 mRNA expressions by the maintaining the phosphorylation of FoxOs and expressions of NF-?B pathway in soleus muscle; the activities of NF-?B pathway might result to the increase of MuRF-1 mRNA expressions, and finally decrease muscle mass in gastrocnemius muscle; the NF-?B pathway were also activited in extensor digitorum longus muscle, while the increase of MuRF-1 mRNA expressions wasn't observed. These results indicated that FoxOs and NF-?B pathway regulated the MuRF-1 mRNA expressions lonely in both gastrocnemius and extensor digitorum longus muscles, and together in soleus muscle. In addition, the regulation mechanism in intracellular atrogin-1 and MuRF-1 mRNA expressions in extensor digitorum longus and gastrocnemius muscles were more complex than that in soleus muscle.Intracellular atrogin-1 and MuRF-1 expressions have tissue-specific and are only detected in skeletal muscle and cardiac muscle. Hibernators maintain contraction function of cardiac muscle under low oxygen and low metabolism occurred in hibernation, which may be closely assosicted to the mainteinance of muscle mass during hibernation. The study found that atrogin-1 and MuRF-1 mRNA experiments maintained in lower level in cardiac muscle in hibernation, which is benefit to the mainteinace of cardiac muscle mass. As like as skeletal muscle (gastrocnemius and extensor digitorum longus muscles), NF-?B pathway in cardiac muscle was activated in hibernating ground squirrels. However, the activated NF-?B pathway wasn't enough to induce the increase of MuRF-1 mRNA expression, which was different from that in skeletal muscle. These results indicated that the regulation of NF-?B pathway to MuRF-1 mRNA expressions in cardiac muscle was relatively weak.