Enhanced Gαq Signaling in <i>TSC2</i> -Deficient Cells Is Required for Their Neoplastic Behavior

Inherited or sporadic loss of the <i>TSC2</i> gene can lead to pulmonary lymphangioleiomyomatosis (LAM), a rare cystic lung disease caused by protease-secreting interstitial tumor nodules. The nodules arise by metastasis of cells that exhibit features of neural crest and smooth muscle lineage (LAM cells). Their aberrant growth is attributed to increased activity of mechanistic target of rapamycin complex 1 (mTORC1), an anabolic protein kinase that is normally suppressed by the TSC1-TSC2 protein complex. The mTORC1 inhibitor rapamycin slows the progression of LAM but fails to eradicate disease, indicating a role for mTORC1-independent mechanisms in LAM pathogenesis. Our previous studies revealed G protein-coupled urotensin-II receptor (UT) signaling as a candidate mechanism, but how it promotes oncogenic signaling in <i>TSC2</i>-deficient cells remained unknown. Using a human pluripotent stem cell-derived <i>in vitro</i> model of LAM, we now show hyperactivation of UT, which was required for their enhanced migration and proneoplastic signaling in a rapamycin-insensitive mechanism that required heterotrimeric Gαq/11 (Gαq). Bioluminescence resonance energy transfer assays in HEK 293T cells lacking <i>TSC2</i> demonstrated selective and enhanced activation of Gαq and its RhoA-associated effectors compared with wild-type control cells. By immunoprecipitation, recombinant UT was physically associated with Gαq and TSC2. The augmented Gαq signaling in <i>TSC2</i>-deleted cells was independent of mTOR activity and associated with increased endosomal targeting of p63RhoGEF, a known RhoA-activating effector of Gαq. These studies identify potential mTORC1-independent proneoplastic mechanisms that can be targeted for prevention or eradication of pulmonary and extrapulmonary LAM tumors.