Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.3.1.28 (chloramphenicol acetyltransferase)
 5,100 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Macrophages respond to lipopolysaccharide (LPS) with the activation of various genes, including the lysozyme gene. Here, we show that the level of lysozyme mRNA increases following treatment of chicken myelomonocytic HD11 cells with LPS. By transient and stable transfection of the chloramphenicol acetyltransferase (CAT) gene controlled by regulatory elements of the lysozyme gene, we identified a subfragment of the -6.1 kilobase (kb) lysozyme enhancer that mediates the LPS-induced lysozyme expression. This subfragment contains two elements (D and E), each of which matches the highly degenerate consensus sequence of binding sites for C/EBP-like transcription factors. Furthermore, we found protein complexes to interact with elements D and E whose binding activity to elements D and E is LPS-inducible in myelomonocytic HD11 cells. Immunomobility shift assays show that NF-M, a myeloid-specific C/EBP beta-related transcription factor is an essential component of these protein complexes. Mutations of the C/EBP binding sites within D and E cause a reduction of basal activity and abolish LPS responsiveness of the -6.1 kb lysozyme enhancer. These results show that the -6.1 kb lysozyme enhancer, in addition to its role in cell type-specific expression, can mediate, by interacting with NF-M, LPS-induced expression of the lysozyme gene in chicken myelomonocytic cells.
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PMID:The far upstream chicken lysozyme enhancer at -6.1 kilobase, by interacting with NF-M, mediates lipopolysaccharide-induced expression of the chicken lysozyme gene in chicken myelomonocytic cells. 798 75

Chicken NF-M transcription factor, in cooperation with either c-Myb or v-Myb, is active in the combinatorial activation of myeloid-cell-specific genes in heterologous cell types, such as embryonic fibroblasts. In humans, similar effects were observed with homologous members of the CCAAT/enhancer-binding protein (C/EBP) family of transcriptional regulators, especially the human homolog of chicken NF-M, C/EBP-beta (NF-IL6). However, the NF-IL6 gene is expressed in a variety of nonmyeloid cell types and is strongly inducible in response to inflammatory stimuli, making it an unlikely candidate to have an exclusive role as a combinatorial differentiation switch during myelopoiesis in human cells. By using a reverse transcription-PCR-based approach and a set of primers specific for the DNA-binding domains of highly homologous members of the C/EBP family of transcriptional regulators, we have cloned a novel human gene encoding a member of the C/EBP gene family, identified as the human homolog of CRP1, C/EBP-epsilon. A 1.2-kb cDNA encoding full-length human C/EBP-epsilon was cloned from a promyelocyte-late myeloblast-derived lambda gt11 library. Molecular analysis of the cDNA and genomic clones indicated the presence of two exons encoding a protein with an apparent molecular mass of 32 kDa and a pI of 9.5. Primer extension analysis of C/EBP-epsilon mRNA detected a single major transcription start site approximately 200 bp upstream of the start codon. The putative promoter area is similar to those of several other myeloid-cell-specific genes in that it contains no TATAAA box but has a number of purine-rich stretches with multiple sites for the factors of the Ets family of transcriptional regulators. Northern blot analyses indicated a highly restricted mRNA expression pattern, with the strongest expression occurring in promyelocyte and late-myeloblast-like cell lines. Western blot and immunoprecipitation studies using rabbit anti-C/EBP-epsilon antibodies raised against the N-terminal portion of C/EBP-epsilon (amino acids 1 to 115) showed that C/EBP-epsilon is a 32-kDa nuclear phosphoprotein. The human C/EBP-epsilon protein exhibited strong and specific binding to double-stranded DNA containing consensus C/EBP sites. Cotransfection of the C/EBP-epsilon sense and antisense expression constructs together with chloramphenicol acetyltransferase reporter vectors containing myeloid-cell-specific c-mim and human myeloperoxidase promoters suggested a role for C/EBP-epsilon transcription factor in the regulation of a subset of myeloid-cell-specific genes. Transient tranfection of a promyelocyte cell line (NB4) with a C/EBP-epsilon expression plasmid increased cell growth by sevenfold, while antisense C/EBP-epsilon caused a fivefold decrease in clonal growth of these cells.
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PMID:Cloning of the novel human myeloid-cell-specific C/EBP-epsilon transcription factor. 903 64