De novo gene expression and antisense inhibition in cultured cells of BmTRN-1, cloned from the midgut of the silkworm, Bombyx mori, which is homologous with mammalian TIA-1/R.
A cDNA encoding a 388 amino acid TIA-1-type RNA-binding protein (BmTRN-1) was isolated from midgut cDNAs of the silkworm, Bombyx mori, via homologous cloning, in order to characterize its function. The deduced amino acid sequence, most likely encoded by a single copy gene, has significant homology with human TIA-1 and TIAR known as apoptotic regulators and recently reported to function as important factors for either splicing or translation. RT-PCR analysis showed that the BmTRN-1 gene was vigorously transcribed in the midgut at the gut purge stage, indicating a possible relation to the tissue-decomposing process in larval-pupal metamorphosis. We also show that inhibition of the expression of BmTRN-1 by a transfected oligonucleotide encoding the antisense sequence caused a remarkable rise in protein expression from artificially constructed cDNAs encoded by plasmid vectors in Bombyx cells, depending on the constructed ORF sequences of the introduced cDNAs. Furthermore, it was shown that the transcripts from the cDNAs introduced into the cells increased under the antisense-inhibition of BmTRN-1 when the protein levels of these cDNAs also rose, demonstrating that BmTRN-1 could act as a regulator especially of the mechanism eliminating transcripts with possible targets for BmTRN-1 recognition in the authentic post-transcription process.
Kotani
Eiji
E
Department of Applied Biology, Kyoto Institute of Technology, Sakyo 606-0962, Japan. kotani@ipc.kit.ac.jp
Ohba
Tadashi
T
Niwa
Takeshi
T
Storey
Kenneth B
KB
Storey
Janet S
JS
Hara
Saburo
S
Saito
Hitoshi
H
Sugimura
Yukio
Y
Furusawa
Toshiharu
T
D000595:Amino Acid Sequence; D000818:Animals; D012831:Bombyx / Q000166:cytology / Q000235:genetics* / Q000254:growth & development; D002478:Cells, Cultured; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D015870:Gene Expression; D018507:Gene Expression Regulation, Developmental; D006801:Humans; D019476:Insect Proteins / Q000235:genetics / Q000378:metabolism; D007814:Larva / Q000235:genetics / Q000254:growth & development; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D008675:Metamorphosis, Biological / Q000235:genetics; D008969:Molecular Sequence Data; D016376:Oligonucleotides, Antisense / Q000235:genetics*; D039102:Poly(A)-Binding Proteins; D011506:Proteins; D011679:Pupa / Q000235:genetics / Q000254:growth & development; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D000074019:T-Cell Intracellular Antigen-1; D014162:Transfection
D018076:DNA, Complementary; D019476:Insect Proteins; D008565:Membrane Proteins; D016376:Oligonucleotides, Antisense; D039102:Poly(A)-Binding Proteins; D011506:Proteins; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D000074019:T-Cell Intracellular Antigen-1; C071558:TIA1 protein, human