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。 多条细胞感知通路汇聚在机械靶的中央信号节点 雷帕霉素复合体1(MTORC1)，代谢的主要调节剂。而氨基酸和葡萄糖的缺乏 已知通过定义明确的机制使mTORC1失活，目前尚不清楚感知 维生素和辅因子的作用机制是存在的，它们对中枢碳代谢同样重要。这里, 大量的基本营养物质维生素B5(VB5)被证明影响细胞周期进程和 MTORC1激活。VB5是辅酶A(CoA)的前体，辅酶A是 碳代谢。然而，对VB5/CoA代谢扰动的反应不是由于细胞能量 耗竭，表明表型并不简单地反映碳源氧化减少。而当 未转化的细胞在选择性VB5耗尽下经历稳定的细胞周期停滞，细胞系无肿瘤 参与调节细胞周期和mTORC1的抑制子反而经历细胞死亡。因此，细胞 周期停滞和mTORC1抑制似乎是VB5/CoA代谢紊乱的生存所必需的。如果 VB5是通过下游的代谢物感觉到的，然后癌细胞缺乏特异性肿瘤的患者 抑制子，或mTORC1表达失调的结节性硬化症患者，可能是潜在的 以辅酶A合成抑制剂为靶点。提出的目标是：1)确定细胞周期 在VB5耗竭的情况下生存所需的调节机制和2)确定 VB5通过研究mTORC1调节蛋白复合体的作用调节mTORC1 代谢物。这两个目标都将表征缺乏肿瘤的细胞对CoA合成抑制的敏感性 参与每条途径的抑制物。这些研究问题将通过基因消融关键基因来解决 细胞周期和mTORC1调节因子与VB5/CoA代谢的干扰 药物对辅酶A合成的抑制作用。将使用生化和质谱学技术 研究VB5/CoA丰度对信号蛋白和代谢产物的影响。拟议中的工作 将阐明一种新的维生素敏感机制，并定义其对关键细胞生长和 增殖途径，有可能揭示靶向癌细胞代谢的脆弱性。

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.