Structure of the 68-kDa neurofilament gene and regulation of its expression.
The complete structure of the mouse 68-kDa neurofilament (NF-L) gene was elucidated. We cloned cDNAs corresponding to 3.5- and 2.3-kb NF-L mRNA, including their polyadenylation sites. Sequence analysis revealed that these NF-L mRNAs arose from the alternative use of two polyadenylation sites in exon 4. Promoter analysis using NF-L promoter-beta-galactosidase fusion plasmids determined regions responsible for its basic promoter activity, which were located between -328 and -36 base pairs from the transcription initiation site. These promoter fusion plasmids induced a significant level of beta-galactosidase in NF-nonproducing C6 cells as well as in NF-producing PC12h cells. The in vitro transcription assay using HeLa cell extract also showed that this promoter exhibited strong transcriptional activity. Little difference in NF-L mRNA stability was observed between the two cells. However, nuclear run-off assay revealed that the NF-L gene was not transcribed in NF-nonproducing C6 cells. These data suggest that the strong promoter activity of the NF-L gene is repressed in vivo at the transcription initiation level in a tissue-specific manner.
Nakahira
K
K
Division of Regulation of Macromolecular Functions, Osaka University, Japan.
Ikenaka
K
K
Wada
K
K
Tamura
T
T
Furuichi
T
T
Mikoshiba
K
K
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D015246:Deoxyribonuclease EcoRI; D005091:Exons; D005786:Gene Expression Regulation; D006367:HeLa Cells; D006801:Humans; D007381:Intermediate Filament Proteins / Q000235:genetics*; D051379:Mice; D008969:Molecular Sequence Data; D016900:Neurofilament Proteins; D009693:Nucleic Acid Hybridization; D011061:Poly A / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics; D015183:Restriction Mapping; D014158:Transcription, Genetic; D014162:Transfection
D007381:Intermediate Filament Proteins; D016900:Neurofilament Proteins; D012333:RNA, Messenger; C044882:neurofilament protein L; D011061:Poly A; D015246:Deoxyribonuclease EcoRI