Regulation of the cell cycle and growth signaling pathways by a sensing mechanism for Vitamin B5-Coenzyme A metabolism

摘要 细胞生长和分裂的决定依赖于对生长因子和营养物质的感知机制。 多个细胞传感通路会聚在机械靶点的中央信号节点， 雷帕霉素复合物1（mTORC 1），代谢的主要调节剂。而氨基酸和葡萄糖剥夺 已知通过明确定义的机制抑制mTORC 1，但仍不清楚是否感测 维生素和辅助因子的机制存在，它们对中心碳代谢同样重要。在这里， 丰富的必需营养素维生素B5（VB 5）被证明影响细胞周期进程， mTORC 1激活。VB 5是辅酶A（CoA）的前体，辅酶A是中枢神经系统必需的酰基链载体。 碳代谢然而，对VB 5/CoA代谢扰动的响应不是由于细胞能量 这表明表型并不简单地反映碳源氧化的减少。而 未转化的细胞在选择性VB 5耗竭下经历稳定的细胞周期停滞，缺乏肿瘤的细胞系 参与细胞周期调节的抑制子和mTORC 1反而经历细胞死亡。因此，Cell 周期阻滞和mTORC 1抑制似乎是受干扰的VB 5/CoA代谢的存活所必需的。如果 VB 5是通过下游代谢物感知的，那么患有缺乏特异性肿瘤细胞的癌细胞的患者 抑制者，或mTORC 1失调的结节性硬化症患者，可能是潜在的 以CoA合成抑制剂为靶点。所提出的目标是：1）确定细胞周期 在VB 5耗竭中生存所必需的调节机制和2）确定 通过研究mTORC 1调节蛋白复合物的作用， 代谢物。这两个目标都将表征缺乏肿瘤细胞中CoA合成抑制的敏感性 抑制剂参与每个途径。这些研究问题将通过基因消融的关键 细胞周期和mTORC 1调节剂以及通过选择性VB 5耗竭干扰VB 5/CoA代谢 和CoA合成的药理学抑制。将使用生化和质谱技术 研究VB 5/CoA丰度对信号蛋白和代谢产物的影响。拟议工作 将揭示一种新的维生素敏感机制，并确定其对关键细胞生长的调节， 增殖途径，具有揭示靶向癌细胞代谢脆弱性的潜力。

Abstract The cellular decision to grow and divide relies on sensing mechanisms for growth factors and nutrients. Multiple cellular sensing pathways converge at the central signaling node of the mechanistic target of rapamycin complex 1 (mTORC1), a master regulator of metabolism. While amino acid and glucose deprivation are known to inactivate mTORC1 through well-defined mechanisms, it remains unknown whether sensing mechanisms exist for vitamins and cofactors, which are equally important for central carbon metabolism. Here, abundance of the essential nutrient Vitamin B5 (VB5) was shown to influence cell cycle progression and mTORC1 activation. VB5 is the precursor for Coenzyme A (CoA), an acyl chain carrier necessary for central carbon metabolism. However, response to perturbation of VB5/CoA metabolism is not due to cellular energy depletion, suggesting that the phenotype does not simply reflect decreased carbon source oxidation. While non-transformed cells undergo stable cell cycle arrest under selective VB5 depletion, cell lines lacking tumor suppressors involved in regulation of the cell cycle and mTORC1 instead undergo cell death. Therefore, cell cycle arrest and mTORC1 inhibition appear to be necessary for survival of perturbed VB5/CoA metabolism. If VB5 is sensed through a downstream metabolite, then patients with cancer cells lacking specific tumor suppressors, or tuberous sclerosis complex patients in which mTORC1 is dysregulated, could be potentially targeted with inhibitors of CoA synthesis. The proposed aims are designed to 1) identify the cell cycle regulatory machinery necessary for survival in VB5 depletion and 2) determine the molecular mechanism of VB5-mediated mTORC1 regulation by investigating the roles of mTORC1-regulating protein complexes and metabolites. Both aims will characterize sensitivity to inhibition of CoA synthesis in cells lacking the tumor suppressors involved in each pathway. These research questions will be addressed by genetic ablation of key cell cycle and mTORC1 regulators and perturbation of VB5/CoA metabolism through selective VB5 depletion and pharmacological inhibition of CoA synthesis. Biochemical and mass spectrometry techniques will be used to investigate the influence of VB5/CoA abundance on signaling proteins and metabolites. The proposed work will shed light on a novel vitamin-sensing mechanism and define its regulation of critical cellular growth and proliferative pathways, with the potential to reveal targetable cancer cell metabolic vulnerabilities.