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CLN5 and CLN8 protein association with ceramide synthase: biochemical and proteomic approaches

Saria El Haddad1, Marwan Khoury, Mohammad Daoud

  • 1Department of Pediatric, American University of Beirut, Beirut, Lebanon.

Electrophoresis
|November 20, 2012
PubMed

Insights

Juvenile neuronal ceroid lipofuscinoses previously classified as CLN9 variant are reclassified as CLN5 disease. CLN5 protein deficiency impacts ceramide synthesis and cell adhesion, with CLN5 and CLN8 proteins potentially activating ceramide synthases.

Area of Science:

  • Biochemistry
  • Cell Biology
  • Genetics

Background:

  • Juvenile neuronal ceroid lipofuscinoses (JNCL) are a group of childhood neurodegenerative disorders.
  • A variant previously known as CLN9 is now reclassified as CLN5 disease based on new genetic findings.

Purpose of the Study:

  • To reclassify patients with a CLN9 variant as CLN5 disease.
  • To investigate the cellular and molecular mechanisms underlying CLN5 deficiency.
  • To explore the relationship between CLN5 and CLN8 proteins in sphingolipid metabolism.

Main Methods:

  • Fibroblast cell culture from CLN5-deficient patients.
  • Biochemical assays measuring ceramide, sphingomyelin, and glycosphingolipid levels.
  • Protein interaction studies using immunoprecipitation, differential gel electrophoresis, and mass spectrometry.
  • Homozygosity mapping and microarray analysis for gene identification.

Main Results:

  • CLN5-deficient cells exhibit adhesion defects, altered growth, increased apoptosis, and reduced levels of key sphingolipids.
  • CLN8 protein partially corrects growth and apoptosis defects in CLN5-deficient cells.
  • Ceramide synthase 1 (CerS1) partially complements CLN5-deficient cells, suggesting CLN5 protein activates CerS1.
  • Absence of gamma-actin and associated proteins (vimentin, histones) in CLN5-deficient cells.
  • Identification of CLN5 as the causative gene for previously classified CLN9-defective cases.

Conclusions:

  • The CLN9 variant is reclassified as CLN5 disease.
  • CLN5 and CLN8 proteins are closely related and may function as activators of (dihydro)ceramide synthases.
  • Defects in CLN5 protein function, potentially involving gamma-actin binding, contribute to the cellular phenotype in CLN5 disease.

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