Genomic organization, alternative splicing, and promoter analysis of human dynamin-like protein gene.
The human dynamin-like protein, HdynIV, has recently been cloned and shown to be involved in the formation and trafficking of coated vesicles. In particular, one of the HdynIV variant overexpressions has been suggested to contribute to the pathogenesis of brain tumors. In this paper, we report on the genomic organization of the human HdynIV gene. The gene was found to correspond to 20 exons of genomic sequence on human chromosome 12, distributed over 64kb of genomic DNA. The two exons, numbers 15 and 16, are subjected to differential splicing, generating four different transcripts of a perfect match to our recent report on the four different spliced HdynIV variants [DNA Cell Biol. 19 (2000) 189]. We have also characterized the 5(') regulatory region of the HdynIV gene in order to understand the molecular mechanisms regulating its expression. The transcriptional initiation site was identified by 5(')-RACE. The 5(')-flanking sequence of the HdynIV gene contains three GC boxes that concatenate Ap2- and Sp1-binding motifs, but that does not contain either the TATA or CAAT consensus sequence. A region between -140 and +92 contributed to high promoter activity. Deletion analysis demonstrated that the minimal promoter activity required the region of -110 to -100. Electrophoretic mobility shift assay demonstrated that a putative transcriptional factor bound to the region of -119 to -90. Site-directed mutagenesis analysis of this region revealed that nucleotides at -108 to -100 were essential for transactivation mediated by this transcriptional factor. In conclusion, we have characterized the minimal HdynIV promoter and shown that CTCCCAGCA (-108 to -100) sequence may act as a novel transcriptional element for regulating HdynIV gene expression.
Howng
Shen-Long
SL
Neurosurgery Department, Kaohsiung Medical University and Hospital, 80708, ROC, Kaohsiung, Taiwan.
Sy
Wei-Di
WD
Cheng
Tai-Shan
TS
Lieu
Ann-Shung
AS
Wang
Chihuei
C
Tzou
Wen-Shyong
WS
Cho
Chung-Lung
CL
Hong
Yi-Ren
YR
D017398:Alternative Splicing; D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D002460:Cell Line; D017931:DNA Primers / Q000235:genetics; D034281:Dynamins; D024202:Electrophoretic Mobility Shift Assay; D020558:GTP Phosphohydrolases / Q000235:genetics* / Q000378:metabolism; D040461:Gene Components / Q000235:genetics; D017930:Genes, Reporter / Q000235:genetics; D015894:Genome, Human; D006801:Humans; D008156:Luciferases / Q000378:metabolism; D008869:Microtubule-Associated Proteins; D024101:Mitochondrial Proteins; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009687:Nuclear Proteins / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics; D011506:Proteins / Q000235:genetics* / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014162:Transfection
D017931:DNA Primers; D008869:Microtubule-Associated Proteins; D024101:Mitochondrial Proteins; D009687:Nuclear Proteins; D011506:Proteins; D011994:Recombinant Proteins; D008156:Luciferases; D020558:GTP Phosphohydrolases; C110768:DNM1L protein, human; D034281:Dynamins