Transcriptional regulatory sequences within the first intron of the chicken apolipoproteinAI (apoAI) gene.

Previous studies demonstrated that the -82 to +87 nucleotides (nt) 5'-upstream region of the chicken apolipoprotein (apoAI) gene are necessary for maximum reporter chloramphenicol acetyl transferase (cat) gene activation in chicken hepatocarcinoma (LMH) cells [Bhattacharyya, N., Chattapadhyay, R., Oddoux, C., Banerjee, D., 1993. Characterisation of the chicken apolipoprotein A-I gene 5'-flanking region. DNA Cell Biol. 12, 597-604]. The -82 to +87nt contain the 5'-untranslated nt, part of the first intron, and the upstream regulatory sequences. In this study, we examined the role of the first intron in the transcriptional regulation of the chicken apoAI gene. Six different reporter cat gene constructs with or without part of the first intron were prepared and transfected into LMH, normal rat kidney (NRK) and human hepatocarcinoma (HepG2) cells. Cell extracts were prepared from each transfected cell line, and CAT activities were measured. All three cell-lines readily expressed CAT, indicating that transcriptional regulatory sequences are present within the first intron region of the chicken apoAI gene. In an enhancer assay, the first intron containing cat construct exhibited a 5.4-fold increase of reporter activity in NRK cells when compared to a SV 40 promoter containing cat plasmid, suggesting the presence of a moderate enhancer element within +29 to +87nt of the first intron. DNase I protection assays, electrophoretic mobility shift assays and binding experiments with nuclear proteins isolated from different chicken tissues and LMH cells showed interaction with +29 to +87nt. Nuclear proteins isolated from tissues like liver and intestine, that actively express apoAI gene, failed to interact with +29 to +87nt, whereas nuclear proteins isolated from tissues that are less active in apoAI gene expression readily interacted with this region. To show the binding of the LMH-specific trans-acting factors to the +50 to +68nt intron region, DNA-affinity chromatography step was performed by using 3H-labeled nuclear proteins. These studies demonstrate that the first intron region of the apoAI gene interacts with trans-acting proteins and plays an important role in transcriptional regulation of the apoAI gene.

[1]  D. Granner,et al.  Regulation of gene expression of rat skeletal muscle/liver 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase. Isolation and characterization of a glucocorticoid response element in the first intron of the gene. , 1992, The Journal of biological chemistry.

[2]  C. Oddoux,et al.  Characterization of the chicken apolipoprotein A-I gene 5'-flanking region. , 1993, DNA and cell biology.

[3]  K. Hashinaka,et al.  Identification of transcriptional cis-elements in introns 7 and 9 of the myeloperoxidase gene. , 1993, The Journal of biological chemistry.

[4]  S. Hedrick,et al.  A transcriptional silencer controls the developmental expression of the CD4 gene. , 1994, The EMBO journal.

[5]  B. Levy-Wilson,et al.  Characterization of tissue-specific enhancer elements in the second intron of the human apolipoprotein B gene. , 1991, The Journal of biological chemistry.

[6]  U. K. Laemmli,et al.  Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4 , 1970, Nature.

[7]  R. Gaynor,et al.  Human immunodeficiency virus type 1 LTR TATA and TAR region sequences required for transcriptional regulation. , 1989, The EMBO journal.

[8]  W. Gilbert,et al.  Sequencing end-labeled DNA with base-specific chemical cleavages. , 1980, Methods in enzymology.

[9]  F. Ruddle,et al.  Use of a protein-blotting procedure and a specific DNA probe to identify nuclear proteins that recognize the promoter region of the transferrin receptor gene. , 1985, Proceedings of the National Academy of Sciences of the United States of America.

[10]  J. Darnell,et al.  Liver-enriched transcription factor HNF-4 is a novel member of the steroid hormone receptor superfamily. , 1990, Genes & development.

[11]  A. Hirsch,et al.  Isolation, characterization and sequencing of the chicken apolipoprotein-AI-encoding gene. , 1991, Gene.

[12]  J. Mathis,et al.  Glucocorticoid regulation of the rat cytochrome P450c (P450IA1) gene: receptor binding within intron I. , 1989, Archives of biochemistry and biophysics.

[13]  D. Littman,et al.  A lineage-specific transcriptional silencer regulates CD4 gene expression during T lymphocyte development , 1994, Cell.

[14]  J. Ordovás,et al.  Linkage, evolution, and expression of the rat apolipoprotein A-I, C-III, and A-IV genes. , 1986, The Journal of biological chemistry.

[15]  P. Dawson,et al.  Structure, evolution, and regulation of chicken apolipoprotein A-I. , 1987, The Journal of biological chemistry.

[16]  M. Dreyfus,et al.  Isolation of DNA-protein complexes based on streptavidin and biotin interaction. , 1987, European journal of biochemistry.

[17]  M. Jung,et al.  Identification of multiple repeat sequences and transcription-related elements within introns 4, 8 and 9 of human Raf-1. , 1993, Biochemical and biophysical research communications.

[18]  H. Stunnenberg,et al.  A major thyroid hormone response element in the third intron of the rat growth hormone gene. , 1990, The EMBO journal.

[19]  D. Kulesh,et al.  Activation of an intron enhancer within the keratin 18 gene by expression of c-fos and c-jun in undifferentiated F9 embryonal carcinoma cells. , 1990, Genes & development.

[20]  M. Sorci-Thomas,et al.  Transcriptional regulation of the apolipoprotein A-I gene. Species-specific expression correlates with rates of gene transcription. , 1991, The Journal of biological chemistry.

[21]  B. Paulweber,et al.  Nuclease-hypersensitive sites define a region with enhancer activity in the third intron of the human apolipoprotein B gene. , 1992, The Journal of biological chemistry.

[22]  S. Lamon-Fava,et al.  Evolutionary distinct mechanisms regulate apolipoprotein A-I gene expression: differences between avian and mammalian apoA-I gene transcription control regions. , 1992, Journal of lipid research.

[23]  J. Burch,et al.  Expression of endogenous and transfected apolipoprotein II and vitellogenin II genes in an estrogen responsive chicken liver cell line. , 1990, Molecular endocrinology.

[24]  D. Banerjee,et al.  Transcriptional regulation of the gene encoding apolipoprotein AI in chicken LMH cells. , 1993, Gene.

[25]  J. Taylor,et al.  Identification and characterization of transcriptional regulatory regions associated with expression of the human apolipoprotein E gene. , 1988, The Journal of biological chemistry.

[26]  Y. Hirayama,et al.  Establishment and characterization of a chicken hepatocellular carcinoma cell line, LMH. , 1987, Cancer research.