Rat nicastrin gene: cDNA isolation, mRNA variants and expression pattern analysis

• Published in: Brain research. Molecular brain research. 2005-05, Vol.136 (1), p.12-22
• Main Authors: Confaloni, Annamaria, Crestini, Alessio, Albani, Diego, Piscopo, Paola, Campeggi, Lorenzo Malvezzi, Terreni, Liana, Tartaglia, Marco, Forloni, Gianluigi
• Format: Article
• Language: English
• Subjects: Alternative splicing; Amyloid Precursor Protein Secretases; Animals; Biological and medical sciences; Blotting, Northern; Blotting, Western - methods; Cells, Cultured; Cerebral Cortex - cytology; Cloning, Molecular - methods; Culture expression; Cycloheximide - pharmacology; DNA, Complementary - isolation & purification; Embryo, Mammalian; Fundamental and applied biological sciences. Psychology; Gene Expression - drug effects; Gene Expression - physiology; Gene Expression Regulation, Enzymologic - physiology; Genetic Variation; Humans; Membrane Glycoproteins - genetics; Membrane Glycoproteins - metabolism; Mice; Molecular Sequence Data; Neuroblastoma; Neuroglia; Nicastrin; Nonsense-mediated mRNA decay; Protein Isoforms - genetics; Protein Isoforms - metabolism; Protein Synthesis Inhibitors - pharmacology; Rats; Rattus norvegicus; Reverse Transcriptase Polymerase Chain Reaction - methods; RNA, Messenger - metabolism; Sequence Homology, Amino Acid; Time Factors; Tissue expression; Vertebrates: nervous system and sense organs
• ISSN: 0169-328X; 1872-6941
• DOI: 10.1016/j.molbrainres.2004.12.022

Nicastrin is a type 1 transmembrane glycoprotein that interacts with presenilin, Aph-1, and Pen-2 proteins to form a high molecular complex with gamma secretase activity. Then, nicastrin has a central role in presenilin-mediated processing of beta-amyloid precursor protein and in some aspects of Notch/glp-1 signaling in vivo. Here, we isolated a rat nicastrin cDNA and investigated gene expression in embryonic and adult rat tissues. The predicted amino acid sequence is comprised of 708 residues and showed a high degree of identity with other vertebrate orthologs. Besides full-length nicastrin mRNA, we identified an alternative spliced variant lacking the whole exon 3 and predicted to encode a 62-residue-long truncated protein. Full-length nicastrin mRNA was observed to be ubiquitously expressed, while the spliced variant was preferentially transcribed in the nervous system, whether in embryonic or adult neural tissues. Studies performed on primary cell cultures demonstrated that the short isoform was expressed in neurons, but not in astrocyte and microglial cells. Further experiments performed to verify the presence of the variant in neuroblastoma culture failed to show any truncated protein. Treatments by cyclohexamide showed the involvement of a quality control-based surveillance mechanism, which selectively degrades the exon 3-skipped isoform. In summary, this is the first report describing a novel skipped isoform of nicastrin which may suggest a new possible control mechanism based on the alternative splicing and nonsense-mediated mRNA decay to regulate brain protein expression and provide newer insights into potential implication in Alzheimer's disease.