Altered lipid homeostasis is associated with cerebellar neurodegeneration in SNX14 deficiency.

Dysregulated lipid homeostasis is emerging as a potential cause of neurodegenerative disorders. However, evidence of errors in lipid homeostasis as a pathogenic mechanism of neurodegeneration remains limited. Here, we show that cerebellar neurodegeneration caused by Sorting Nexin 14 (SNX14) deficiency is associated with lipid homeostasis defects. Recent studies indicate that SNX14 is an interorganelle lipid transfer protein that regulates lipid transport, lipid droplet (LD) biogenesis, and fatty acid desaturation, suggesting that human SNX14 deficiency belongs to an expanding class of cerebellar neurodegenerative disorders caused by altered cellular lipid homeostasis. To test this hypothesis, we generated a mouse model that recapitulates human SNX14 deficiency at a genetic and phenotypic level. We demonstrate that cerebellar Purkinje cells (PCs) are selectively vulnerable to SNX14 deficiency while forebrain regions preserve their neuronal content. Ultrastructure and lipidomic studies reveal widespread lipid storage and metabolism defects in SNX14-deficient mice. However, predegenerating SNX14-deficient cerebella show a unique accumulation of acylcarnitines and depletion of triglycerides. Furthermore, defects in LD content and telolysosome enlargement in predegenerating PCs suggest lipotoxicity as a pathogenic mechanism of SNX14 deficiency. Our work shows a selective cerebellar vulnerability to altered lipid homeostasis and provides a mouse model for future therapeutic studies.

Transmembrane 163 (TMEM163) protein effluxes zinc.

Recent investigations of rodent Tmem163 suggest that it binds to and transports zinc as a dimer, and that alanine mutagenesis of its two species-conserved aspartate (D123A/D127A) residues proposed to bind zinc, perturbs protein function. Direct corroboration, however, is lacking whether it is an influx or efflux transporter in cells. We hypothesized that human TMEM163 is a zinc effluxer based on its predicted protein characteristics. We used cultured human cell lines that either stably or transiently expressed TMEM163, and pre-loaded the cells with zinc to determine transport activity. We found that TMEM163-expressing cells exhibited significant reduction of intracellular zinc levels as evidenced by two zinc-specific fluorescent dyes and radionuclide zinc-65. The specificity of the fluorescence signal was confirmed upon treatment with TPEN, a high-affinity zinc chelator. Multiple sequence alignment and phylogenetic analyses showed that TMEM163 is related to distinct members of the cation diffusion facilitator (CDF) protein family. To further characterize the efflux function of TMEM163, we substituted alanine in two homologous aspartate residues (D124A/D128A) and performed site-directed mutagenesis of several conserved amino acid residues identified as non-synonymous single nucleotide polymorphism (S61R, S95C, S193P, and E286K). We found a significant reduction of zinc efflux upon cellular expression of D124A/D128A or E286K protein variant when compared with wild-type, suggesting that these particular amino acids are important for normal protein function. Taken together, our findings demonstrate that TMEM163 effluxes zinc, and it should now be designated ZNT11 as a new member of the mammalian CDF family of zinc efflux transporters.