| Summary: | 碩士 === 國立臺灣大學 === 分子與細胞生物學研究所 === 102 === The Bcl-2 family is the best characterized protein family involved in the regulation of apoptotic cell death, consisting of anti-apoptotic and pro-apoptotic members. These proteins determine the life or dead of cells by altering the VDAC permeability, which is located on the mitochondrial outer membrane. However, recent studies suggest that bcl-2 family proteins have cellular functions beyond regulation of apoptosis.
The BAD protein is a pro-apoptotic member of Bcl-2 family whose ability to heterodimerize with Bcl-2 and Bcl-xL, and increase permeability of VDAC. On the mitochondrial outer membrane, Bad assemble a complex together with PKA, PP1c, WAVE-1, and glucokinase. BAD complex and the phosphorylation state of BAD regulate the glucokinase activity in hepatocytes and the insulin secretion in beta cells, raising the possibility that BAD may be involved in nutrient metabolism. The mitochondrial anaplerosis activity is in response to cellular nutrient availability. To investigate the interaction between BAD activity and anaplerosis, we treated U2OS cells with anaplerosis inhibitor, aminooxyacetic acid(AOA), and measured the phosphorylation levels of BAD. We found that AOA induces BAD(S136) phosphorylation and reduces BAD(S155) phosphorylation. The treatments with BPTES result in the same BAD phosphorylation state, suggest that BAD phosphorylation states correspond to mitochondrial anaplerosis activity. In addition, soluble adenylyl cyclase inhibitor, KH7 treatments have the same effect. Suggest that mitochondrial anaplerosis activity may be coordinated by BAD complex through the cAMP/PKA signaling.
Treatment with AOA, BPTES, KH7, both reduce the activity of mTORC1(p-S6K1-T389 decrease),and induce the activity of mTORC2(p-Akt-S473 decrease). mTORC2-Akt(S474) phosphorylates BAD at S136, may occur in membrane raft. The phosphorylation of BAD(S136)may regulate WAVE-1(the component of BAD complex), and make mitochondria move to plama membrane in order to acquire nutrient. Sequentially, cAMP-PKA signaling phosphorylate BAD(S155).
The results suggest that BAD phosphorylation states correspond to mitochondrial anaplerosis activity, and mitochondrial anaplerosis activity may be coordinated by BAD complex through the cAMP/PKA signaling; furthermore affect mTORC1 and mTORC2 activity to regulate metabolism and cell growth.
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