👤 A N Siakotos

Hereditary ceroid-lipofuscinosis in English setters has been proposed to be the canine equivalent of human juvenile ceroid-lipofuscinosis, which results from defects in the CLN3 gene. Analyses were performed to determine whether the disease in English setters is also the consequence of a CLN3 gene mutation. Canine CLN3 cDNA was found to contain a 1,314-bp open reading frame predicting a derived amino acid sequence which is 89%, 85%, and 84% identical to the predicted amino acid sequences for the human, mouse, and rabbit CLN3 proteins, respectively. The canine gene has sixteen exons. No differences were detected when cDNA nucleotide sequences from an English setter with ceroid-lipofuscinosis and from a normal dog were compared. Moreover, alleles of the canine CLN3 gene distinguished by an intragenic marker segregated independently from the disease in an English setter family, eliminating CLN3 as the locus for the canine disease. A ceroid-lipofuscinosis-affected Tibetan terrier was homozygous for a Gly70Glu CLN3 variant; however, this allele is common in dog breeds considered free of ceroid-lipofuscinosis. Show less

The identification of the genetic defect in CLN1 as a palmitoyl-protein thioesterase deficiency initiated a search for the lysosomal storage material. Pulse-chase labelling of fibroblasts and lymphoblastoid cell lines with [35S]cysteine revealed the presence of lipid [35S]cysteine material in CLN1 fibroblasts and not in controls, CLN2 or CLN3 patients or other patients with lipidosis. A single band comigrated with the acylcysteine standard and labelling with [3H]palmitate showed a band of material which eluted from the silicic acid column with the phospholipid fraction and which co-migrated with the lipid-[35S]cysteine band. The storage material is tentatively identified as palmitoylcysteine. Show less

The storage of subunit c of mitochondrial ATP synthase, other hydrophobic peptides, and autofluorescent pigment in both late infantile (CLN2) and juvenile (CLN3) neuronal ceroid lipofuscinosis, but not in infantile (CLN1), has raised the question of abnormal mitochondrial function. We now report a partial deficiency in three types of fatty acid oxidation in intact skin fibroblasts from CLN2 and CLN3 patients, but not CLN1. We observed a statistically significant 33% reduction in palmitate (beta-oxidation; mainly mitochondrial) and lignocerate (beta-oxidation; mainly peroxisomal), and a 50% reduction in phytanic acid (alpha-oxidation; mainly peroxisomal) in the absence of exogenous carnitine. In contrast, when we measured fatty acid beta-oxidation (lignoceric acid and palmitic acid), in the same human skin fibroblasts, following lysis in the presence of carnitine, we found no difference in enzyme activity among normal, CLN1, CLN2, and CLN3. However, we did observe a 40% reduction in peroxisomal particulate (bound) catalase activity in CLN1 and CLN2 fibroblasts, which typically results from organellar lipid accumulation or a membrane abnormality. However, total catalase levels were normal, and Western blot analysis of this and three other major oxidant protective enzymes (Mn-dependent superoxide dismutase [MnSOD], CuZn-dependent superoxide dismutase [CuZnSOD], and glutathione peroxidase) were normal in CLN1, CLN2, and CLN3, as well as in liver from an animal (English Setter dog) model for CLN, which shows similar pathology and subunit c storage. Our data showing differences between CLN1 and forms CLN2 and CLN3 suggest some type of mitochondrial membrane abnormality as the source of the pathology in CLN2 and CLN3. Show less