A rat brain mRNA encoding a transcriptional activator homologous to the DNA binding domain of retroviral integrases.
We have isolated a rat cDNA, named FE65, hybridizing to an mRNA of about 2,300 nucleotides present in rat brain, undetectable in rat liver and very poorly represented in other tissues. An mRNA of the same size is present in human neuroblastoma cells and is absent from other human cell lines. The FE65 cDNA contains an open reading frame (ORF) coding for a polypeptide of 499 amino acids in which 143 residues can be aligned with the DNA binding domain of the integrases encoded by mammalian immunodeficiency viruses. The remaining part of the FE65 ORF is not homologous with the correspondent regions of the integrases; the first 206 residues of the FE65 ORF show numerous negative charges and a short sequence not dispensable for the function of the transactivating acidic domain of the jun family transcriptional factors. A plasmid which expresses FE65 amino acids 1-232 fused to the yeast GAL4 DNA binding domain was co-transfected with a plasmid containing five GAL4 binding sites upstream of a minimal Adenovirus promoter controlling the expression of the CAT gene. This experiment showed that the fused protein GAL4-FE65 is able to obtain a 30-40 fold increase of the CAT gene expression compared to the expression observed in the presence of the GAL4 DNA binding domain alone. Two types of FE65 mRNA are present in rat brain, differing only for six nucleotides. We demonstrate that this is the consequence of a neuron-specific alternative splicing of a six-nucleotide miniexon, which is also present in the human genome, in an intron/exon context very similar to that of the rat FE65 gene.
Dipartimento di Biochimica e Biotecnologie Mediche, Università degli Studi di Napoli, II Facoltà di Medicina e Chirurgia, Italy.
A
A
Duilio
N
N
Zambrano
A R
AR
Mogavero
R
R
Ammendola
F
F
Cimino
T
T
Russo
D000595:Amino Acid Sequence; D000818:Animals; D001482:Base Composition; D015152:Blotting, Northern; D001921:Brain / Q000378:metabolism*; D004254:DNA Nucleotidyltransferases / Q000235:genetics*; D005786:Gene Expression Regulation / Q000235:genetics; D006678:HIV / Q000201:enzymology; D006367:HeLa Cells; D006801:Humans; D019426:Integrases; D008969:Molecular Sequence Data; D009447:Neuroblastoma / Q000378:metabolism; D016366:Open Reading Frames / Q000235:genetics; D012326:RNA Splicing / Q000502:physiology; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology; D016415:Sequence Alignment; D012689:Sequence Homology, Nucleic Acid; D014157:Transcription Factors / Q000235:genetics*
D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; D004254:DNA Nucleotidyltransferases; D019426:Integrases