Abstract

1. Galactosialidosis is a lysosomal storage disease associated with a combined deficiency of β-galactosidase and neuraminidase, secondary to a defect of another lysosomal protein, the protective protein/cathepsin A (PPCA).

   
   

2. All patients have clinical manifestations typical of a lysosomal disorder, such as coarse facies, cherry-red spots, vertebral changes, foam cells in the bone marrow, and vacuolated lymphocytes. Three phenotypic subtypes are recognized. The early infantile form is associated with fetal hydrops, edema, ascites, visceromegaly, skeletal dysplasia, and early death. Hepatosplenomegaly, growth retardation, cardiac involvement, and rare occurrence of important neurologic signs characterize the late infantile type. The majority of reported patients belong to the juvenile/adult group and are mainly of Japanese origin. Myoclonus, ataxia, angiokeratoma, mental retardation, neurologic deterioration, absence of visceromegaly, and long survival are quite characteristic of this subtype.

   
   

3. Sialyloligosaccharides accumulate in lysosomes and are excreted in body fluids. Ultrastructural analysis of central, peripheral, and autonomic nervous systems, retina, liver, kidney, skin, and lymphocytes shows numerous membrane-bound vacuoles and, in some tissues, many membranous cytoplasmic bodies.

   
   

4. The autosomal gene encoding human PPCA that has been localized on chromosome 20q13.1 transmits the disorder. The PPCA cDNA and genomic sequences have been used to analyze the mutations causing galactosialidosis. Mainly single-base substitutions or splice-junction defects have been identified in patients with different clinical phenotypes. Correlation of genotype with disease severity has emerged from structure/function studies of the mutant enzyme.

   
   

5. PPCA associates with β-galactosidase and neuraminidase in an early biosynthetic compartment. By virtue of this association the two glycosidases are correctly routed to the lysosome and are protected against rapid intralysosomal proteolysis. Mammalian PPCA is a multifunctional enzyme with distinct protective and catalytic activities. It is synthesized as a 54-kDa precursor that is processed into two disulfide-linked chains of 32 and 20 kDa by endoproteolytic removal of a 2-kDa peptide. The enzyme is active at both acidic and neutral pH and functions as cathepsin A/deamidase/esterase on a subset of neuropeptides, including substance P, oxytocin, and endothelin I. This activity was found to be deficient in all patients with galactosialidosis so far tested.

   
   

6. The three-dimensional structure of the PPCA precursor has been solved by a combination of molecular replacement and twofold density averaging. The structure revealed an unusual inactivation mechanism of the zymogen that explains how removal of the 2-kDa peptide triggers activation. A number of amino acid substitutions found in defective PPCA from different galactosialidosis patients have been localized in the three-dimensional structure. There appears to be a significant correlation between the effect of each mutation on the integrity of protein structure and the general severity of the clinical phenotype.

   
   

7. The disease can be suspected in patients with clinical features of a lysosomal storage disease showing sialyloligosacchariduria on thin-layer chromatography. The demonstration of a combined deficiency of β-galactosidase and neuraminidase in lymphocytes and/or cultured skin fibroblasts is the preferred method of biochemical diagnosis. Assaying only the cathepsin A activity of PPCA might give a false negative result because, in principle, a mutation could affect the protective function but not the catalytic activity. However, carrier detection should be based on assays involving the primary defect.

   
   

8. Prenatal diagnosis has been established by demonstrating a combined β-galactosidase/neuraminidase deficiency in cultured amniotic fluid cells. Furthermore, sialyloligosaccharides in deproteinized amniotic fluid supernatant were detected in the case of an affected fetus.

   
   

9. Mice homozygous for a null mutation at the PPCA locus develop a phenotype closely resembling human patients with the severe form of galactosialidosis. Excessive excretion of sialyloligosaccharides in urine is diagnostic of the disease. Systemic organ pathology can be fully corrected by transplanting null mutants with bone marrow from transgenic mice overexpressing human PPCA in specific hemopoietic lineages.

   
   

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