Pulmonary, Allergy and Critical Care Division, Department of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
Mol Cell Biol. 2011 Jun;31(12):2484-98. doi: 10.1128/MCB.01061-10. Epub 2011 Apr 11.
Mutational inactivation of the tumor suppressor tuberous sclerosis complex 2 (TSC2) constitutively activates mTORC1, increases cell proliferation, and induces the pathological manifestations observed in tuberous sclerosis (TS) and in pulmonary lymphangioleiomyomatosis (LAM). While the role of mTORC1 in TSC2-dependent growth has been extensively characterized, little is known about the role of mTORC2. Our data demonstrate that mTORC2 modulates TSC2-null cell proliferation and survival through RhoA GTPase and Bcl2 proteins. TSC2-null cell proliferation was inhibited not only by reexpression of TSC2 or small interfering RNA (siRNA)-induced downregulation of Rheb, mTOR, or raptor, but also by siRNA for rictor. Increased RhoA GTPase activity and P-Ser473 Akt were inhibited by siRNA for rictor. Importantly, constitutively active V14RhoA reversed growth inhibition induced by siRNA for rictor, siRNA TSC1, reexpression of TSC2, or simvastatin. While siRNA for RhoA had a modest effect on growth inhibition, downregulation of RhoA markedly increased TSC2-null cell apoptosis. Inhibition of RhoA activity downregulated antiapoptotic Bcl2 and upregulated proapoptotic Bim, Bok, and Puma. In vitro and in vivo, simvastatin alone or in combination with rapamycin inhibited cell growth and induced TSC2-null cell apoptosis, abrogated TSC2-null tumor growth, improved animal survival, and prevented tumor recurrence by inhibiting cell growth and promoting apoptosis. Our data demonstrate that mTORC2-dependent activation of RhoA is required for TSC2-null cell growth and survival and suggest that targeting both mTORC2 and mTORC1 by a combination of proapoptotic simvastatin and cytostatic rapamycin shows promise for combinational therapeutic intervention in diseases with TSC2 dysfunction.
肿瘤抑制因子结节性硬化复合物 2 (TSC2) 的突变失活可使 mTORC1 持续激活，促进细胞增殖，并导致结节性硬化症 (TS) 和肺淋巴管平滑肌瘤病 (LAM) 中观察到的病理表现。虽然 mTORC1 在 TSC2 依赖性生长中的作用已被广泛研究，但对 mTORC2 的作用知之甚少。我们的数据表明，mTORC2 通过 RhoA GTPase 和 Bcl2 蛋白调节 TSC2 缺失细胞的增殖和存活。不仅 TSC2 的重新表达或 Rheb、mTOR 或 raptor 的 siRNA 诱导下调可抑制 TSC2 缺失细胞的增殖，而且 rictor 的 siRNA 也可抑制 TSC2 缺失细胞的增殖。ric tor 的 siRNA 抑制 RhoA GTPase 活性增加和 P-Ser473 Akt。重要的是，组成性激活的 V14RhoA 逆转了 rictor、TSC1、TSC2 的重新表达或辛伐他汀的 siRNA 诱导的生长抑制。虽然 RhoA 的 siRNA 对生长抑制有轻微影响，但下调 RhoA 可显著增加 TSC2 缺失细胞的凋亡。RhoA 活性的抑制下调抗凋亡的 Bcl2 并上调促凋亡的 Bim、Bok 和 Puma。在体外和体内，辛伐他汀单独或与雷帕霉素联合抑制细胞生长并诱导 TSC2 缺失细胞凋亡，消除 TSC2 缺失肿瘤生长，提高动物存活率，并通过抑制细胞生长和促进凋亡来预防肿瘤复发。我们的数据表明，mTORC2 依赖性 RhoA 激活对于 TSC2 缺失细胞的生长和存活是必需的，并表明通过联合促凋亡的辛伐他汀和细胞抑制的雷帕霉素靶向 mTORC2 和 mTORC1，为 TSC2 功能障碍相关疾病的联合治疗干预提供了希望。
