Mutation of β-glucosidase 2 causes glycolipid storage disease and impaired male fertility
J. Clin. Invest. Yildiz Yildiz, et al. 116:2985 doi:10.1172/JCI29224 [
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Figure 6Analysis of glycolipids in
Gba2 wild-type and knockout mice.
(
A) Glycolipids were extracted from testes of 6-, 20-, and 24-month-old mice, resolved by 1-dimensional TLC, and visualized by staining with orcinol. The positions of the origin and glucosylceramide standards are indicated at left. A glycolipid that accumulated with age in knockout tissue is marked “X” at right. The positions to which several fucosylated glycolipids migrated to are indicated at left. (
B) Structural analysis of glycolipid X shown in
A. Glycolipids were extracted from the testes of
Gba2–/– mice and resolved by 1-dimensional TLC, and glycolipid X was analyzed by mass spectrometry. An
m/z 264 precursor ion scan revealed compounds of molecular masses (700.9 and 728.3) characteristic of monohexosylceramides containing sphingosine (d18:1) and either 16:0 or 18:0 fatty acyl groups. A neutral loss scan for
m/z 180 diagnostic for glucose- or galactose-containing monohexosylceramides revealed a major compound of mass 700.9. (
C) Expression of mouse and human GBA2 in COS cells and measurement of glucosylceramidase and bile acid glucosidase enzyme activities. Cells were transfected with the indicated plasmid DNA, and lysates were assayed for enzyme activity using fluorescently labeled glucosylceramide or bile acid glucoside substrates. (
D) Expression of mouse GBA2 (mGBA2) in human embryonic kidney 293 cells and measurement of glucosylceramidase activity. Cells were transfected and assayed for enzyme activity using [
14C]glucosylceramide. The positions of the origin and to which authentic glucosylceramide and ceramide standards migrated to are indicated at left. pCMV-mGBA2, expression plasmid encoding mouse GBA2 enzyme.
