Rat nicastrin gene: cDNA isolation, mRNA variants and expression pattern analysis.
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.
Confaloni
Annamaria
A
Department of Cellular Biology and Neuroscience, Istituto Superiore di Sanità, 299 Viale Regina Elena, 00161 Rome, Italy. annamar@iss.it
Crestini
Alessio
A
Albani
Diego
D
Piscopo
Paola
P
Campeggi
Lorenzo Malvezzi
LM
Terreni
Liana
L
Tartaglia
Marco
M
Forloni
Gianluigi
G
D053829:Amyloid Precursor Protein Secretases; D000818:Animals; D015152:Blotting, Northern; D015153:Blotting, Western / Q000379:methods; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology; D003001:Cloning, Molecular / Q000379:methods; D003513:Cycloheximide / Q000494:pharmacology; D018076:DNA, Complementary / Q000302:isolation & purification*; D004622:Embryo, Mammalian; D015870:Gene Expression / Q000187:drug effects / Q000502:physiology; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology*; D014644:Genetic Variation; D006801:Humans; D008562:Membrane Glycoproteins / Q000235:genetics* / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D009447:Neuroblastoma; D009457:Neuroglia; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D017386:Sequence Homology, Amino Acid; D013997:Time Factors
D018076:DNA, Complementary; D008562:Membrane Glycoproteins; D020033:Protein Isoforms; D011500:Protein Synthesis Inhibitors; D012333:RNA, Messenger; C413604:nicastrin protein; D003513:Cycloheximide; D053829:Amyloid Precursor Protein Secretases