HIF signalling and mitochondrial biogenesis

Using the RCC cell line, UOK257, Preston et al. (2011) demonstrated that FLCN inhibits hypoxia-inducible factor (HIF) signalling. Additionally, HIF-1a expression is increased in cystic lung tissue resected from a BHD patient, also suggesting that under normal conditions, FLCN inhibits HIF signalling (Nishii et al., 2013). The HIF signalling pathway regulates a number of different genes that are involved in angiogenesis, erythropoiesis, cell survival and metastasis (Semenza, 2012). Interestingly, increased vascularisation of sub-pleural cysts expressing increased amounts of HIF-1a has been observed (Nishii et al., 2013).

Preston et al. (2011) observed that increased HIF-signalling in FLCN-null cells lead to the increased activity of glycolytic enzymes, thus causing these cells to favour glycolytic rather than lipid metabolism, as seen in the Warburg effect (Warburg, 1956). The resulting increased lactate production could also be activitating a second HIF-independent hypoxia pathway mediated by NDRG3 (Lee et al., 2015).

The Warburg effect states that cancerous cells produce energy by glycolysis and lactic fermentation in the cytosol rather than by pyruvate oxidation in mitochondria (Warburg, 1956). Thus, Warburg also postulated that cancer could be interpreted as a mitochondrial disease. Indeed, Klomp et al. (2010) found that the loss of FLCN in BHD-associated renal tumours resulted in mitochondrial dysfunction, as indicated by an increased expression of the mitochondrial genes PGC1A and TFAM in these tumours. Additionally, Hasumi et al. (2012) show that when FLCN is deleted in murine muscle tissues, there is an increase in mitochondrial gene expression and a subsequent metabolic shift towards mitochondrial oxidative phosphorylation, and this is regulated by PGC1A (Hasumi et al., 2012).

A study by Yan et al. (2014) drew many of these observations together. They found that AMPK is constitutively activated in FLCN-null mouse embryonic fibroblasts, leading to increased PGC1A activity and a consequent increase in mitochondrial biogenesis and ROS production. This ultimately activates HIF signalling and leads to metabolic changes consistent with the Warburg Effect within FLCN-null cells. Re-addition of FLCN into this cell line reduced the number of colonies formed in a soft agar assay, indicating that the metabolic transformation as a result of FLCN-loss provides these cells with a tumorigenic advantage.

Biallelic FLCN inactivation in murine heart muscle causes cardiac hypertrophy, cardiac dysfunction and significantly reduces lifespan compared to wild type littermates (Hasumi et al., 2014). This effect was mediated by PGC1a, as double FLCN/PGC1a cardiac knock out animals did not show this phenotype (Hasumi et al., 2014). FLCN deletion led to overexpression of PGC1a, leading to increased mitochondrial mass and high intracellular ATP levels, which ultimately led to AMPK inactivation and mTORC1 hyperactivity (Hasumi et al., 2014).

Conversely, biallelic inactivation of FNIP1 leads to reduced mitochondrial mass and lower basal ATP levels in iNKT cells (Park et al., 2014).

Interestingly, mitochondrial hyperplasia is a common characteristic of oncocytic tumours, and is commonly observed in tumours resected from BHD patients (Lindor et al., 2012; Pradella et al., 2013; Raymond et al., 2014).