Animal Models for the Study of Gaucher Disease

Abstract

In Gaucher disease (GD), a relatively common sphingolipidosis, the mutant lysosomal enzyme acid β-glucocerebrosidase (GCase), encoded by the GBA1 gene, fails to properly hydrolyze the sphingolipid glucosylceramide (GlcCer) in lysosomes, particularly of tissue macrophages. As a result, GlcCer accumulates, which, to a certain extent, is converted to its deacylated form, glucosylsphingosine (GlcSph), by lysosomal acid ceramidase. The inability of mutant GCase to degrade GlcSph further promotes its accumulation. The amount of mutant GCase in lysosomes depends on the amount of mutant ER enzyme that shuttles to them. In the case of many mutant GCase forms, the enzyme is largely misfolded in the ER. Only a fraction correctly folds and is subsequently trafficked to the lysosomes, while the rest of the misfolded mutant GCase protein undergoes ER-associated degradation (ERAD). The retention of misfolded mutant GCase in the ER induces ER stress, which evokes a stress response known as the unfolded protein response (UPR). GD is remarkably heterogeneous in clinical manifestation, including the variant without CNS involvement (type 1), and acute and subacute neuronopathic variants (types 2 and 3). The present review discusses animal models developed to study the molecular and cellular mechanisms underlying GD.

Keywords: ER stress; GBA1; glucocerebrosidase (GCase); glucosylceramide (GlcCer); inflammation; knockin animals; knockout animals; misfolding; unfolded protein response (UPR).

Figure 1

Two pathologies in GD. In GD, there are two pathologies: A. ER retention of misfolded GCase molecules in the ER, leading to ER stress and ER stress response, known as the unfolded protein response (UPR) [36,37], and B. Due to the decreased number of GCase molecules in the lysosomes and their decreased ability to normally degrade the substrate, there is substrate accumulation.Illustration was produced using BioRender.

Figure 2

Multiple sequence alignment of active GCases between different organisms. MSA analysis of human, fly, mouse, and fish GCase sequences. The known human and predicted animal leader sequences are highlighted in blue. The two amino acids in the active site are highlighted in purple and the amino acids involved in stabilizing it are highlighted in yellow. The prevalent GD mutations E326, N370, V394, D409, L444, and R496 are shown in orange.

Figure 3

Mutant zebrafish models. Abnormalities noted in glycolipids, presence of Gaucher cells, and abnormal motor coordination (data from Lelieveld et al., 2019 and 2022). gba1 encodes lysosomal GCase converting GlcCer to Cer and glucose; gba2 encodes cytosol-facing GCase converting GlcCer to Cer and generating GlcChol via transglycosylation of cholesterol; asha1b encodes acid ceramidase isoform capable of converting GlcCer to GlcSph in lysosomes. N—normal; nd—not determined; ↑—elevated levels; ↑↑—high levels; ↓—decreased levels. ++—severely discordant movement.

Figure 4

Expression of a foreign gene (transgene) in flies. Through a cross between two flies, a line is established in which the transcription factor GAL4 is expressed from a native promoter coupled to it. GAL4 binds to its promoter, UAS, and activates the expression of a foreign gene coupled to the UAS.

Figure 5

Drosophila GBA1 proteins. Multiple sequence alignment analysis comparing human GCase and the two Drosophila GBA1-encoded protein sequences (Dmel GBA1b, GBA1a). The two amino acids in the active site are highlighted in green. Amino acids that stabilize the active site are highlighted in purple. The known human and predicted fly leader sequences are highlighted in blue and orange, respectively.

Figure 6

Drosophila locus containing the GBA1 orthologs. Schematic representation of the chromosomal region containing the fly GBA1 orthologs. Gba1a is located 2 Kb upstream of Gba1b. CG31413 is a non-relevant gene. Exons of Gba1a appear in maroon and those of Gba1b in green. Red arrow represents the deletion created by Davis et al. [134] (see KO Drosophila models). Illustration made using BioRender.

Figure 7

Drosophila GBA1 orthologs containing the Minos insertions. Schematic representation of the Minos insertion site in each gene. Exons of Gba1a appear in maroon and those of Gba1b in green. The lengths of Gba1a- and Gba1b-translated proteins are indicated in maroon and green, respectively. The lengths of the truncated proteins due to Minos insertion (Gba1a^m and Gba1b^m) are indicated in red. The Minos insertion sites are indicated using white triangles. Illustration made using BioRender.

Figure 8

Different animal models used to study mutant GBA1 variants associated with GD. Shown are key features of animal models used to study GD.