title: Novel HSAN1 mutation in serine palmitoyltransferase resides at a putative phosphorylation site that is involved in regulating substrate specificity.
abstract: 1-Deoxysphingolipids (1-deoxySL) are atypical sphingolipids that are formed by the enzyme serine palmitoyltransferase (SPT) due to a promiscuous use of L-alanine over its canonical substrate L-serine. Several mutations in SPT are associated with the hereditary sensory and autonomic neuropathy type I (HSAN1). The current hypothesis is that these mutations induce a permanent shift in the affinity from L-serine toward L-alanine which results in a pathologically increased 1-deoxySL formation in HSAN1 patients. Also, wild-type SPT forms 1-deoxySL under certain conditions, and elevated levels were found in individuals with the metabolic syndrome and diabetes. However, the molecular mechanisms which control the substrate shift of the wild-type enzyme are not understood. Here, we report a novel SPTLC2-S384F variant in two unrelated HSAN1 families. Affected patients showed elevated plasma 1-deoxySL levels and expression of the S384F mutant in HEK293 cells increased 1-deoxySL formation. Previously, S384 has been reported as one of the two (S384 and Y387) putative phosphorylation sites in SPTLC2. The phosphorylation of wild-type SPTLC2 was confirmed by isoelectric focusing. The impact of an S384 phosphorylation on SPT activity was tested by creating mutants mimicking either a constitutively phosphorylated (S384D, S384E) or non-phosphorylated (S384A, Y387F, Y387F+S384A) protein. The S384D but not the S384E variant was associated with increased 1-deoxySL formation. The other mutations had no influence on activity and substrate affinity. In summary, our data show that S384F is a novel mutation in HSAN1 and that the substrate specificity of wild-type SPT might by dynamically regulated by a phosphorylation at this position.
authors.affiliation: Institute for Clinical Chemistry, University Hospital Zurich, CH-8091, Zurich, Switzerland.
authors.forename: Daniela
authors.initials: D
authors.lastname: Ernst
authors.forename: Sinéad M
authors.initials: SM
authors.lastname: Murphy
authors.forename: Karthik
authors.initials: K
authors.lastname: Sathiyanadan
authors.forename: Yu
authors.initials: Y
authors.lastname: Wei
authors.forename: Alaa
authors.initials: A
authors.lastname: Othman
authors.forename: Matilde
authors.initials: M
authors.lastname: Laurá
authors.forename: Yo-Tsen
authors.initials: YT
authors.lastname: Liu
authors.forename: Anke
authors.initials: A
authors.lastname: Penno
authors.forename: Julian
authors.initials: J
authors.lastname: Blake
authors.forename: Michael
authors.initials: M
authors.lastname: Donaghy
authors.forename: Henry
authors.initials: H
authors.lastname: Houlden
authors.forename: Mary M
authors.initials: MM
authors.lastname: Reilly
authors.forename: Thorsten
authors.initials: T
authors.lastname: Hornemann
mesh_terms: D000368:Aged; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D017124:Conserved Sequence; D015180:Electrophoresis, Gel, Two-Dimensional; D005260:Female; D057809:HEK293 Cells; D009477:Hereditary Sensory and Autonomic Neuropathies / Q000235:genetics*; D006579:Heterozygote; D006720:Homozygote; D006801:Humans; D007525:Isoelectric Focusing; D008297:Male; D008875:Middle Aged; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009431:Neural Conduction; D010375:Pedigree; D010766:Phosphorylation; D010768:Phosphoserine / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D051102:Serine C-Palmitoyltransferase / Q000737:chemistry / Q000235:genetics* / Q000502:physiology; D013045:Species Specificity; D013107:Sphingolipids / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D011993:Recombinant Fusion Proteins; D013107:Sphingolipids; D010768:Phosphoserine; C487784:SPTLC1 protein, human; D051102:Serine C-Palmitoyltransferase