title: The Niemann-Pick C1 protein in feline fibroblasts.
abstract: Niemann-Pick type C (NPC) disease is a rare inherited metabolic disorder characterized by hepatosplenomegaly, progressive neurodegeneration, and storage of lipids such as cholesterol and glycosphingolipids in most tissues. The current study was conducted to characterize the Niemann-Pick C1 (NPC1) protein in feline fibroblasts. This was accomplished by generating rabbit polyclonal antibodies against a peptide corresponding to amino acids 1256-1275 of the feline NPC1 protein. The results obtained using immunoblot analysis identified two major proteins that migrated at approximately 140 and 180 kDa. These two proteins were absent when immunoblots were incubated in the presence of feline NPC1 antibody and immunizing peptide, or preimmune serum. Fluorescence microscopy of feline fibroblasts incubated with the feline NPC1 antibody revealed granular staining within the perinuclear region of the cell. This granular staining was diminished when feline fibroblasts were incubated in the presence of feline NPC1 antibody and immunizing peptide, or was completely absent when feline fibroblasts were incubated in the presence of preimmune serum. Additional studies using double-labeled fluorescence microscopy indicated that feline NPC1 partially colocalized with markers for late endosomes/lysosomes, endoplasmic reticulum, and microtubules, but not the trans-Golgi network. In summary, the results presented in this report demonstrate that the NPC1 protein in feline fibroblasts has a similar distribution as that previously described for human and murine fibroblasts.
authors.affiliation: Department of Pediatrics, Arizona Health Sciences Center, The University of Arizona, Tucson 85724, USA. wgarver@peds.arizona.edu
authors.forename: William S
authors.initials: WS
authors.lastname: Garver
authors.forename: Kyra
authors.initials: K
authors.lastname: Somers
authors.forename: Kumar
authors.initials: K
authors.lastname: Krishnan
authors.forename: Thomas
authors.initials: T
authors.lastname: Mitchell
authors.forename: Randall A
authors.initials: RA
authors.lastname: Heidenreich
authors.forename: Mary Anna
authors.initials: MA
authors.lastname: Thrall
mesh_terms: D000818:Animals; D000906:Antibodies; D002352:Carrier Proteins / Q000032:analysis; D002415:Cats; D005347:Fibroblasts / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D015151:Immunoblotting; D047908:Intracellular Signaling Peptides and Proteins; D008562:Membrane Glycoproteins / Q000032:analysis; D008856:Microscopy, Fluorescence; D000077296:Niemann-Pick C1 Protein; D009542:Niemann-Pick Diseases / Q000378:metabolism*
chemical_list: D000906:Antibodies; D002352:Carrier Proteins; D047908:Intracellular Signaling Peptides and Proteins; D008562:Membrane Glycoproteins; C106881:NPC1 protein, human; D000077296:Niemann-Pick C1 Protein