RSK4 and the regulation of cellular metabolism in lung cancer

Abstract

RSK4 is a member of the p90 ribosomal S6 kinase (RSK) family, which promotes metastasis and chemoresistance in non-small cell lung cancer (NSCLC), and its inhibition improves therapeutic outcomes in the in vivo and ex vivo models of this disease. However, the effect of RSK4 inhibition on cancer metabolism was unknown.

Here, I have demonstrated that RSK4 downregulation using RNA interference in A549 NSCLC cell line enhances glycolysis, glutaminolysis, and reductive carboxylation. It achieves this through increasing the levels of key enzymes that catalyse these metabolic processes, including hexokinase 2, glutaminase kidney isoform, and isocitrate dehydrogenases 1 and 2. RSK4 silencing sensitises NSCLC cells grown in 3D culture to 2-deoxy-D-glucose (glycolytic inhibitor) and 6-Diazo-5-oxo-L-norleucine (glutaminase inhibitor). Moreover, it triggers the elongation of mitochondrial networks in NSCLC cells.

Interestingly, downregulation of RSK4 is associated with the p53-dependent increase in hypoxia-inducible factor 1 alpha (HIF-1α) protein levels in normoxia, a phenomenon compatible with the observed glucose and glutamine metabolic rewiring. This is, at least, partly achieved through Serine 125 dephosphorylation of HIF prolyl hydroxylase 2 (PHD2), thereby inactivating this enzyme that targets HIF-1α for degradation. I found that RSK4 interacts with HIF-1α and PHD2. RSK4 also phosphorylates HIF-1α in vitro on T276, S464, S465 and S692. The biological significance of RSK4-mediated HIF-1α phosphorylation on these residues remains unknown and requires further investigation. Finally, I demonstrated that HIF-1α is responsible for cell cycle arrest in RSK4-silenced A549 cells by transactivating p21, an inhibitor of cyclin-dependent kinases.

In summary, this research project reveals a connection between RSK4 and HIF-1α, a key metabolic regulator downstream of cellular oxygen-sensing. RSK4 inhibition promotes pseudo-hypoxic metabolic responses in lung adenocarcinoma, and these alterations may offer novel metabolic dependencies to improve the outcome of RSK4-targeting therapies.