Vascular stiffness mechanoactivates YAP/TAZ-dependent glutaminolysis to drive pulmonary hypertension
Thomas Bertero, … , Joshua Fessel, Stephen Y. Chan
Thomas Bertero, … , Joshua Fessel, Stephen Y. Chan
Published August 22, 2016
Citation Information: J Clin Invest. 2016;126(9):3313-3335. https://doi.org/10.1172/JCI86387.
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Research Article Pulmonology Vascular biology

Vascular stiffness mechanoactivates YAP/TAZ-dependent glutaminolysis to drive pulmonary hypertension

Abstract

Dysregulation of vascular stiffness and cellular metabolism occurs early in pulmonary hypertension (PH). However, the mechanisms by which biophysical properties of the vascular extracellular matrix (ECM) relate to metabolic processes important in PH remain undefined. In this work, we examined cultured pulmonary vascular cells and various types of PH-diseased lung tissue and determined that ECM stiffening resulted in mechanoactivation of the transcriptional coactivators YAP and TAZ (WWTR1). YAP/TAZ activation modulated metabolic enzymes, including glutaminase (GLS1), to coordinate glutaminolysis and glycolysis. Glutaminolysis, an anaplerotic pathway, replenished aspartate for anabolic biosynthesis, which was critical for sustaining proliferation and migration within stiff ECM. In vitro, GLS1 inhibition blocked aspartate production and reprogrammed cellular proliferation pathways, while application of aspartate restored proliferation. In the monocrotaline rat model of PH, pharmacologic modulation of pulmonary vascular stiffness and YAP-dependent mechanotransduction altered glutaminolysis, pulmonary vascular proliferation, and manifestations of PH. Additionally, pharmacologic targeting of GLS1 in this model ameliorated disease progression. Notably, evaluation of simian immunodeficiency virus–infected nonhuman primates and HIV-infected subjects revealed a correlation between YAP/TAZ–GLS activation and PH. These results indicate that ECM stiffening sustains vascular cell growth and migration through YAP/TAZ-dependent glutaminolysis and anaplerosis, and thereby link mechanical stimuli to dysregulated vascular metabolism. Furthermore, this study identifies potential metabolic drug targets for therapeutic development in PH.

Authors

Thomas Bertero, William M. Oldham, Katherine A. Cottrill, Sabrina Pisano, Rebecca R. Vanderpool, Qiujun Yu, Jingsi Zhao, Yiyin Tai, Ying Tang, Ying-Yi Zhang, Sofiya Rehman, Masataka Sugahara, Zhi Qi, John Gorcsan III, Sara O. Vargas, Rajan Saggar, Rajeev Saggar, W. Dean Wallace, David J. Ross, Kathleen J. Haley, Aaron B. Waxman, Victoria N. Parikh, Teresa De Marco, Priscilla Y. Hsue, Alison Morris, Marc A. Simon, Karen A. Norris, Cedric Gaggioli, Joseph Loscalzo, Joshua Fessel, Stephen Y. Chan

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Figure 2

A metabolic switch induced by ECM stiffening is coordinated by the mechanoactivation of YAP/TAZ.

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A metabolic switch induced by ECM stiffening is coordinated by the mecha...
(A) Immunoblot analysis confirmed the knockdown of YAP and TAZ by 2 independent siRNA sequences in PAECs. (B–E) PAECs were cultured in soft or stiff matrix. Released lactate was increased in stiff matrix, but such increase was blunted by siRNA knockdown of YAP/TAZ (B). In stiff matrix, YAP/TAZ knockdown also blunted specific metabolite alterations reflective of anaplerotic (C) and glycolytic activity as reflected by the lactate/pyruvate ratio (D). MitoTracker labeling confirmed that YAP/TAZ knockdown reversed the alteration of mitochondrial membrane potential triggered by stiff matrix (E). (F) Immunoblot analysis confirmed the forced expression of YAP in PAECs infected with a lentiviral vector containing the YAP coding sequences (pYAP) compared with cells infected with a control vector (pGFP). (G) In PAECs cultivated on soft matrix, forced expression of YAP (pYAP) increased lactate compared with control vector (pGFP). (H and I) In similarly treated PAECs, YAP decreased glutamine, pyruvate, and succinate, increased glutamate and aspartate (H), and increased lactate/pyruvate ratio (I). (J) MitoTracker labeling confirmed that YAP (pYAP) in soft matrix altered PAEC mitochondrial membrane potential. In all panels, mean expression in control groups (si-NC, pGFP cultivated on soft matrix) was assigned a fold change of 1, to which relevant samples were compared. Data are expressed as mean ± SEM (*P < 0.05, §P < 0.01, #P < 0.001) of at least 3 independent experiments performed in triplicate. Paired samples were compared by 2-tailed Student’s t test, while 1-way ANOVA and post-hoc Tukey’s tests were used for group comparisons. Scale bars: 20 μm. See also Supplemental Figure 2 (similar results were found in PASMCs).