An intron is required for dihydrofolate reductase protein stability.
We compared the expression of dihydrofolate reductase minigenes with and without an intron. The levels of protein were significantly higher in the presence of dihydrofolate reductase intron 1. However, mRNA levels in both constructs were comparable. In addition, the RNA transcribed from either construct was correctly polyadenylated and exported to the cytoplasm. The intron-mediated increase in dihydrofolate reductase protein levels was position-independent and was also observed when dihydrofolate reductase intron 1 was replaced by heterologous introns. The translational rate of dihydrofolate reductase protein was increased in transfectants from the intron-containing minigene. In addition, the protein encoded by the intronless construct was unstable and subject to lysosomal degradation, thus showing a shorter half-life than the protein encoded by the intron-containing minigene. We conclude that an intron is required for the translation and stability of dihydrofolate reductase protein.
Noé
Véronique
V
Department of Biochemistry and Molecular Biology, School of Pharmacy, University of Barcelona E-08028, Spain. noe@farmacia.far.ub.es
MacKenzie
Simon
S
Ciudad
Carlos J
CJ
D017398:Alternative Splicing; D000818:Animals; D016466:CHO Cells; D002467:Cell Nucleus / Q000378:metabolism; D006224:Cricetinae; D003593:Cytoplasm / Q000378:metabolism; D005091:Exons; D007438:Introns; D008247:Lysosomes / Q000378:metabolism; D011233:Precipitin Tests; D014176:Protein Biosynthesis; D012313:RNA / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012260:Ribonucleases / Q000378:metabolism; D013347:Subcellular Fractions; D013762:Tetrahydrofolate Dehydrogenase / Q000737:chemistry* / Q000235:genetics*; D013997:Time Factors; D014162:Transfection
D012333:RNA, Messenger; D012313:RNA; D013762:Tetrahydrofolate Dehydrogenase; D012260:Ribonucleases