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Janet Deane

Lipid processing defects in neurodegenerative disease

Elucidating how disruptions to lipid recycling in the lysosome cause severe neurodegenerative diseases.

Efficient signal transduction in the nervous system requires that neurons be wrapped in a membrane-rich insulating sheath known as myelin. This sheath is highly enriched with a special class of lipids known as glycosphingolipids. If the recycling and turnover of these lipids is defective the myelin sheath becomes damaged and the neurons degenerate causing severe diseases.

Our research focuses on a rare autosomal recessive disorder, Krabbe disease, which primarily affects infant children. It is caused by deficiencies in the enzyme galactocerebrosidase (GALC), which is responsible for the degradation of the myelin lipid galactocerebroside (GalCer, Fig. 1A). GALC is produced in the ER-Golgi complex and then traffics to the lysosome where it is essential for lipid recycling. Defects in GALC lead to the accumulation of cytotoxic metabolites that elicit complex, and still only partially understood, cellular events that result in apoptosis of myelin-forming cells and rapid neurodegeneration. We have identified several molecular mechanisms by which GALC function can be disrupted including loss of catalytic activity, misfolding and mistrafficking of GALC. However, GALC does not function in isolation and requires an additional protein Saposin A (SapA) which is a lipid-binding and lipid-transfer protein (Fig. 1B).

In order to understand how GALC and SapA function together to coordinate the efficient recycling of sphingolipids we determined the structure of the GALC-SapA complex in the presence of lipid mimics (Fig. 1C). A central SapA dimer binds the hydrophobic tails of the lipids, shielding them from the aqueous solvent (Fig. 1D). This arrangement allows the glycosylated headgroup of the lipid to remain exposed in order to bind the active site of GALC to allow efficient cleavage to occur. This structure provides new insights into the mechanism by which these hydrophobic substrates can be cleaved by soluble enzymes: SapA extracts the sphingolipid from the bilayer and presents it to the GALC active site for processing. A number of disease-relevant mutations are buried in this interface, identifying the critical role of this interaction for correct sphingolipid processing.

We are using a range of techniques from protein biochemistry and structural biology to cell-based assays and proteomics to address how these defects in GALC result in changes in the cell that trigger demyelination and neurodegeneration.

 

Deane GALC

 

Figure 1. Glycosphingolipid processing by GALC and SapA

(A) The cleavage of galactocerebroside (GalCer) by GALC produces galactose and ceramide. (B) Lipid substrates must be extracted from the membrane in order to be processed. (C) Our crystal structure of the GALC-SapA complex reveals that a central dimer of SapA (yellow and orange) is enclosed by two molecules of GALC (cyan and magenta). (D) A cross-section through the structure reveals a continuous channel from the GALC active site into the SapA core, into which hydrophobic lipid substrates can bind.

 

Deane lab

          

Key papers:

Hill CH, Cook GM, Spratley SJ, Fawke S, Graham SC & Deane JE. The mechanism of glycosphingolipid degradation revealed by a GALC-SapA complex structure. Nature Comm. 9:151 (2018) 

Demydchuk M, Hill CH, Zhou A, Bunkóczi G, Stein PE, Marchesan D, Deane JE & Read RJ. Insights into Hunter syndrome from the structure of iduronate-2-sulfatase. Nature Comm. 8:15786 (2017)

Neerincx A, Hermann C, Antrobus R, van Hateren A, Cao H, Trautwein N, Stevanović S, Elliott T, Deane JE & Boyle LH. TAPBPR bridges UDP-glucose:glycoprotein glucosyltransferase 1 onto MHC class I to provide quality control in the antigen presentation pathway. Elife 6. pii: e23049 (2017).

Spratley SJ & Deane JE. New therapeutic approaches for Krabbe disease: The potential of pharmacological chaperones. J Neurosci Res. 94:1203-19 (2016).

Spratley SJ, Hill CH, Viuff AH, Edgar JR, Skjødt K & Deane JE. Molecular mechanisms of disease pathogenesis differ in Krabbe disease variants. Traffic 17, 908-922 (2016).

Hermann C, van Hateren A, Trautwein N, Neerincx A, Duriez PJ, Stevanovic S, Trowsdale J, Deane JE, Elliott E & Boyle LH. TAPBPR alters MHC class I peptide presentation by functioning as a peptide exchange catalyst. eLife 10.7554/eLife.09617 (2015).

Hill CH, Viuff AH, Spratley SJ, Salamone S, Christensen SH, Read RJ, Moriarty NW, Jensen HH & Deane JE. Azasugar Inhibitors as Pharmacological Chaperones for Krabbe Disease. Chemical Science 6, 3075-3086, DOI: 10.1039/C5SC00754B (2015).

Hill CH, Graham SC, Read RJ & Deane JE. Structural snapshots illustrate the catalytic cycle of β-galactocerebrosidase, the defective enzyme in Krabbe disease. Proc. Natl Acad. Sci. USA. 110, 20479–20484 (2013).

 

Janet Deane 

Dr Janet Deane

Royal Society University Research Fellow

Department of Pathology

e-mail: jed55@cam.ac.uk

01223 762 815

Plain English

Our cells can be thought of like a house: there are different rooms for different purposes. In the cell, these ‘rooms’ are known as organelles and the ‘walls’ are made of lipids. But cells are self-repairing and can synthesize, breakdown and recycle their own lipids. When lipid recycling becomes defective it results in severe human diseases, often due to rapid and fatal neurodegeneration. Our lab uses a range of techniques to probe how lipids are recycled in the cell and what happens when this recycling breaks down.

 

Group members

Stuart Fawke · Maria Shamin

Funding

The Royal Society

Wellcome Trust

Medical Research Council