Defective lysosomal egress of free sialic acid (N-acetylneuraminic acid) in fibroblasts of patients with infantile free sialic acid storage disease.
Egress of free NeuAc from normal lysosome-rich granular fractions was assessed at NeuAc concentrations of up to 221 pmol/hexosaminidase unit, achieved by exposure of growing fibroblasts to 40-125 nM N-acetylmannosamine for up to 7 days. The normal velocity of NeuAc egress increased with NeuAc loading and with temperature, exhibiting a Q10 of 2.4, characteristic of carrier-mediated transport. Fibroblasts cultured from five patients with infantile free sialic acid storage disease (ISSD) contained approximately 139 nmol of free NeuAc/mg of whole cell protein, or 100 times the normal level. Differential centrifugation, as well as density gradient analysis using 25% Percoll, showed that the stored NeuAc cosedimented with the lysosomal enzyme beta-hexosaminidase. The velocity of appearance of free NeuAc outside ISSD granular fractions was negligible, even at initial loading levels of up to 3500 pmol/hexosaminidase unit. The lack of egress from ISSD granular fractions was found for both endogenous and N-acetylmannosamine-derived NeuAc. Fibroblasts from ISSD parents did not accumulate excess free NeuAc and did not display a velocity of NeuAc egress significantly different from normal. The defect in ISSD, like that in Salla disease, appears to be an impairment of carrier-mediated transport of free NeuAc across the lysosomal membrane. Clinical and biochemical differences between Salla disease and ISSD may reflect differences in the amount of residual NeuAc transport capacity.
Laboratory of Molecular and Cell Biology, National Institute of Diabetes and Digestive and Kidney Diseases, Bethesda, Maryland 20892.
F
F
Tietze
R
R
Seppala
M
M
Renlund
J J
JJ
Hopwood
G S
GS
Harper
G H
GH
Thomas
W A
WA
Gahl
D002239:Carbohydrate Metabolism, Inborn Errors / Q000235:genetics / Q000378:metabolism*; D002458:Cell Fractionation / Q000379:methods; D002478:Cells, Cultured; D002499:Centrifugation, Density Gradient / Q000379:methods; D005347:Fibroblasts / Q000378:metabolism; D006580:Genetic Carrier Screening; D006801:Humans; D007223:Infant; D007700:Kinetics; D008247:Lysosomes / Q000378:metabolism* / Q000648:ultrastructure; D019158:N-Acetylneuraminic Acid; D012016:Reference Values; D012794:Sialic Acids / Q000378:metabolism*
D012794:Sialic Acids; D019158:N-Acetylneuraminic Acid