Cycloheximide stimulated SOCS-3 gene expression in 3T3-L1 adipocyte via the ERK pathway

• Main Authors: Pei-hua Tsai, 蔡沛樺
• Format: Others
• Language: zh-TW
• Published: 2011
• Online Access: http://ndltd.ncl.edu.tw/handle/29066217623604543117

碩士 === 國立中央大學 === 生命科學研究所 === 99 === SOCS-3 (suppressors of cytokine signaling-3) is a 24.7 kDa protein that plays a very important role in the signaling transduction of insulin resistance caused by resistin in 3T3-L1 adipocytes. However, we found that cycloheximide, a protein synthesis inhibitor, could stimulate SOCS-3 mRNA expression in 3T3-L1 adipocytes. Despite of these, the exact mechanism of cycloheximide’s action on SOCS-3 gene expression and its effect on adipocyte are still unknown. Therefore, this study was to investigate how cycloheximide could affect the expression of adipocyte SOCS-3 gene. We found that cycloheximide stimulated SOCS-3 mRNA expression in a time- and dose-dependent manner. Actinomycin-D (a transcription inhibitor) blocked the cycloheximide-stimulated SOCS-3 mRNA expression, suggesting the need of a new mRNA synthesis, but not due to changes in the mRNA stability. Interestingly, cycloheximide did not significantly alter the SOCS-3 protein levels. Pretreatment with U0126 (an ERK MAPK inhibitor) reduced the cycloheximide-stimulated SOCS-3 mRNA levels by 50%. This suggests the ERK-dependent effect of cycloheximide. Treatment with an additional protein inhibitor, such as anisomycin, also stimulated SOCS-3 mRNA expression in C3H10T1/2 adipocyte and C2C12 myoblast, but it had no effect on H4IIEC3 hepatoma cell. This suggests that the cycloheximide effects vary with the cell types. Moreover, cycloheximide stimulated gene expression of other SOCS family members, such as SOCS-1，SOCS-2，SOCS-4，SOCS-5，SOCS-6，SOCS-7 and CIS；However, the different peak time of the individual SOCS mRNA level occurred among SOCSs. Finally, we found that the dose of cycloheximide at 10 μg/ml for 6-24 h and 5 μg/ml for 12-24 h, but not 5 μg/ml within 6 h, reduced cell number and cell viability of the preadipocytes and adipocytes. We conclude that cycloheximide stimulates 3T3-L1 adipocyte SOCS-3 mRNA expression via the MEK1/ERK-dependent pathway. Further study is needed to demonstrate whether the effects of cycloheximide on SOCS-3 gene expression are related to its action on fat cell viability.