Specific sphingolipid content decrease in Cerkl knockdown mouse retinas

• An exhaustive sphingolipidome of WT and Cerkl −/− mouse retinas is provided.
• Cerkl −/− retinas showed a statistically significant sphingolipid content decrease.
• Other retinal lipids, such as neutral lipids and phospholipids, remain unaltered.
• CERKL is not involved in the transcription regulation of the SPL-related genes.
• CERKL binds to GlcCer, GalCer and sphingomyelin in protein–lipid overlay assays.

Sphingolipids (SPLs) are finely tuned structural compounds and bioactive molecules involved in membrane fluidity and cellular homeostasis. The core sphingolipid, ceramide (CER), and its derivatives, regulate several crucial processes in neuronal cells, among them cell differentiation, cell–cell interactions, membrane conductance, synaptic transmission, and apoptosis. Mutations in Ceramide Kinase-Like (CERKL) cause autosomal recessive Retinitis Pigmentosa and Cone Rod Dystrophy. The presence of a conserved lipid kinase domain and the overall similarity with CERK suggested that CERKL might play a role in the SPL metabolism as a CER kinase. Unfortunately, CERKL function and substrate(s), as well as its contribution to the retinal etiopathology, remain as yet unknown. In this work we aimed to characterize the mouse retinal sphingolipidome by UPLC-TOF to first, thoroughly investigate the SPL composition of the murine retina, compare it to our Cerkl −/− model, and finally assess new possible CERKL substrates by phosphorus quantification and protein-lipid overlay. Our results showed a consistent and notable decrease of the retinal SPL content (mainly ranging from 30% to 60%) in the Cerkl −/− compared to WT retinas, which was particularly evident in the glucosyl/galactosyl ceramide species (Glc/GalCer) whereas the phospholipids and neutral lipids remained unaltered. Moreover, evidence in favor of CERKL binding to GlcCer, GalCer and sphingomyelin has been gathered. Altogether, these results highlight the involvement of CERKL in the SPL metabolism, question its role as a kinase, and open new scenarios concerning its function.