Modulation of the Autophagic Process by S1P Under Hypoxia: Dissecting the Role of S1PR1 and S1PR5 Receptors

The active biolipid sphingosine-1-phosphate (S1P) regulates cellular survival and stress responses, emerging also as a potential modulator of autophagy and mitophagy. Given the well-established role of S1P signalling in hypoxia, this study investigated how S1P modulates autophagic processes in a neuronal hypoxic model, mimicking such process in vitro through the chemical stabilization of the Hypoxia-inducible factor 1-alpha (HIF-1α) subunit via DMOG-mediated prolyl hydroxylase inhibition. Hypoxia-like conditions reshaped S1P metabolism by downregulating its major degradative enzymes, Sphingosine-1-phosphate phosphatase 1 (SGPP1) and Sphingosine-1-phosphate lyase 1 (SGPL1), potentially prolonging its signalling. Conversely, exogenous S1P administration determined a compensatory upregulation of these catabolic genes, likely promoting signal termination. After characterizing the distinct morphological rearrangements induced by DMOG and S1P, we examined their combined impact on the autophagic machinery. While DMOG alone activated autophagy, exogenous S1P acted as an inhibitor. Notably, selective activation of S1PR1 and S1PR5 showed divergent effects; however, their co-administration with DMOG led to a significant accumulation of Microtubule-associated protein 1 light chain 3 (LC3) and SQSTM1/p62. This molecular signature, further supported by mitochondrial-lysosomal colocalization analyses and thresholded Manders’ coefficients, is suggestive of altered mitophagy. Our findings demonstrate that S1P signalling is a key modulator of the neuronal hypoxic response, acting also mitochondrial quality control. The convergence between hypoxia-induced stress and S1P receptor activation during mitophagy suggests a primary role for S1P in protecting cells from metabolic exhaustion. Consequently, S1P biosynthesis and S1P receptors (S1PR) emerge as promising therapeutic targets for neurological conditions associated with hypoxic stress, dysregulated autophagy, and mitochondrial dysfunction. Divergent roles of S1PR1 and S1PR5 in the neuronal hypoxic response. HIF-1α upregulates GLUT1 , GLUT3 , B N IP3 , and SP HK1 . Under hypoxic conditions, exogenous S1P downregulates SPHK2 , but also SGPP1 and SGPL1 , which are involved in S1P catabolism, thus prolonging S1P signalling. Selective activation of S1PR1 and S1PR5 receptors induces divergent responses in cytoskeletal dynamics and degradative pathways compared to the pan-receptor effect of S1P. Concomitant HIF-1α stabilization further reprogrammes these responses