Damir Khabibullin, MS
Pulmonary and Clinical Care Medicine
Damir Khabibullin*, Yongfeng Luo, Hui Chen, Wei Shi and Lisa Henske
Birt-Hogg-Dube (BHD) syndrome is a rare autosomal disorder characterized by benign skin tumors, kidney cancer and cystic lung disease with lung collapse (pneumothorax). BHD is caused by germline loss-of-function mutations in the Folliculin (FLCN) gene. FLCN has been linked to multiple signaling networks including mTOR, AMPK and WNT and many of its functions are mediated by the transcription factors TFEB and TFE3, which we previously linked to WNT signaling.
The pathogenesis of pulmonary cystic disease in BHD is poorly understood. In unpublished data, we developed a novel lung mesenchymal stromal cell (MSC) line derived from a floxed-Flcn and mT/mG reporter mouse. In this model, Flcn deletion was induced by adenoviralCre expression and Tfeb or Tfe3 was downregulated using shRNA. FLCN-deficient MSCs grew 60% faster compared to control cells and shRNA downregulation of Tfe3 or Tfeb prevented this increase, indicating that TFE3/TFEB may be directly involved in BHD-associated cystic lung disease.
RNA-sequencing of the MSCs revealed that expression of Wnt pathway ligands and receptors was significantly increased in Flcn-deficient cells, including Wnt7a (over 1,000-fold increase), Wnt10a (37-fold increase), Wnt9a (2.5-fold increase), Lgr5 (26-fold increase). Wnt signaling plays a critical role during lung development, regulating mesenchyme/epithelium crosstalk in the alveoli. TFE3 downregulation in FLCN-deficient cells reversed 75-80% of the FLCN-dependent transcriptional changes, including all Wnt ligands and receptors. In contrast, TFEB downregulation reversed only 40% of the FLCN-dependent changes. Interestingly, in this cellular context, some FLCN-dependent transcriptional changes were equally dependent on TFE3 and TFEb (such as Igfbp5 and Serpinb7), with near complete normalization by either TFE3 or TFEB downregulation, while other genes were primarily dependent on TFE3.
These data suggest that TFE3 is a critical downstream target of FLCN in lung mesenchyme and may play a role in BHD-associated pulmonary cystic disease via dysregulation of Wnt signaling during lung development.
While up to 90% of BHD patients develop lung cysts and 30-35% develop pneumothorax, the pathogenesis of pulmonary cystic disease in BHD is poorly understood. We hope that our work will provide insights into the mechanisms of how FLCN deficiency impacts lung mesenchyme cells that were previously not considered as potential drivers of cystic lung disease in BHD. Moreover, our data suggests that abnormal activation of TFE3 and WNT abnormalities downstream of FLCN deficiency could be a critical step in disrupting normal alveolar homeostasis in BHD. If confirmed in vivo, it will necessitate the development of novel therapeutic approaches to target TFE3.