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Analysis of Response Elements Involved in the Regulation of the Human Neonatal Fc Receptor Gene (FCGRT).

Mikulska JE - PLoS ONE (2015)

Bottom Line: Moreover, the CF1/YY1 site at -586 in differentiated THP-1 cells, plays an essential role in the transcriptional activity of the promoter.In addition, the C/EBPbeta binding site at -497 of the hFCGRT promoter in epithelial and endothelial cells, and the C/EBPbeta motif located at -497 and -233 within the hFCGRT promoter in differentiated THP-1 cells may function as positive regulatory sequences in response to LPS or PMA stimulation.EMSA and supershift analyses showed that the functionally identified binding motifs in the hFCGRT promoter were able to specifically interact with their corresponding (Sp1, Sp2, Sp3, c-Fos, c-Jun, YY1, and C/EBPbeta or C/EBPdelta) transcription factors (TFs), suggesting their possible involvement in the regulation of the human FCGRT gene expression.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunochemistry, Ludwik Hirszfeld Institute of Immunology and Experimental Therapy, Polish Academy of Sciences, Wroclaw, Poland.

ABSTRACT
Human epithelial, endothelial and PMA-differentiated THP-1 cell lines were used as model systems to study the transcriptional regulation of the human FCGRT gene encoding the alpha chain of hFcRn. The data obtained from site-directed mutagenesis in transient transfection experiments indicate that the Sp1 sites at positions -641, -635, and -313, CF1/YY1 elements at positions -586 and -357, and the AP-1 motif at -276 within the-660/-233 fragment of the human FCGRT promoter (hFCGRT) participate in the regulation of human FCGRT in all selected cell lines. However, their individual contribution to promoter activity is not equivalent. The Sp1 binding site at -313 and the AP-1 site at -276 are critical for the activity of the hFCGRT promoter in epithelial and endothelial cells. Moreover, the CF1/YY1 site at -586 in differentiated THP-1 cells, plays an essential role in the transcriptional activity of the promoter. In addition, the C/EBPbeta binding site at -497 of the hFCGRT promoter in epithelial and endothelial cells, and the C/EBPbeta motif located at -497 and -233 within the hFCGRT promoter in differentiated THP-1 cells may function as positive regulatory sequences in response to LPS or PMA stimulation. EMSA and supershift analyses showed that the functionally identified binding motifs in the hFCGRT promoter were able to specifically interact with their corresponding (Sp1, Sp2, Sp3, c-Fos, c-Jun, YY1, and C/EBPbeta or C/EBPdelta) transcription factors (TFs), suggesting their possible involvement in the regulation of the human FCGRT gene expression.

No MeSH data available.


Related in: MedlinePlus

Functional analysis of the CF1 binding sites within the hFCGRT promoter.(A) Schematic diagram representing the wild-type promoter construct and the mutant promoter constructs. Mutations are marked with crosses. (B) Summarized results of luciferase activity tests. The mutant constructs pGL3-711(mCF1-1), pGL3-711(mCF1-2), and pGL3-711(mCF1-1+2) are derivatives of the wild-type pGL3-711(WT) with mutation in the CF1 site at positions-586, -357, -586 plus -357, respectively. Transfection and normalization were performed as described in the legend to Fig 8. All results represent means ± SD of three to six independent experiments performed in duplicate and plotted as a percentage of the normalized activity of the wild-type pGL3-711(WT) construct, which is defined as 100%. The promoter activity of the labeled construct is significantly decreased compared to the wild-type construct; *P < 0.01 vs. pGL3-711(WT); **P < 0.01 compared with pGL3-711(mCF1-2) and pGL3-711(mCF1-1+2) in THP-1.
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pone.0135141.g010: Functional analysis of the CF1 binding sites within the hFCGRT promoter.(A) Schematic diagram representing the wild-type promoter construct and the mutant promoter constructs. Mutations are marked with crosses. (B) Summarized results of luciferase activity tests. The mutant constructs pGL3-711(mCF1-1), pGL3-711(mCF1-2), and pGL3-711(mCF1-1+2) are derivatives of the wild-type pGL3-711(WT) with mutation in the CF1 site at positions-586, -357, -586 plus -357, respectively. Transfection and normalization were performed as described in the legend to Fig 8. All results represent means ± SD of three to six independent experiments performed in duplicate and plotted as a percentage of the normalized activity of the wild-type pGL3-711(WT) construct, which is defined as 100%. The promoter activity of the labeled construct is significantly decreased compared to the wild-type construct; *P < 0.01 vs. pGL3-711(WT); **P < 0.01 compared with pGL3-711(mCF1-2) and pGL3-711(mCF1-1+2) in THP-1.

Mentions: Functional analysis of the CF1/YY1 binding sites within the hFCGRT promoter is presented in Fig 10.


Analysis of Response Elements Involved in the Regulation of the Human Neonatal Fc Receptor Gene (FCGRT).

Mikulska JE - PLoS ONE (2015)

Functional analysis of the CF1 binding sites within the hFCGRT promoter.(A) Schematic diagram representing the wild-type promoter construct and the mutant promoter constructs. Mutations are marked with crosses. (B) Summarized results of luciferase activity tests. The mutant constructs pGL3-711(mCF1-1), pGL3-711(mCF1-2), and pGL3-711(mCF1-1+2) are derivatives of the wild-type pGL3-711(WT) with mutation in the CF1 site at positions-586, -357, -586 plus -357, respectively. Transfection and normalization were performed as described in the legend to Fig 8. All results represent means ± SD of three to six independent experiments performed in duplicate and plotted as a percentage of the normalized activity of the wild-type pGL3-711(WT) construct, which is defined as 100%. The promoter activity of the labeled construct is significantly decreased compared to the wild-type construct; *P < 0.01 vs. pGL3-711(WT); **P < 0.01 compared with pGL3-711(mCF1-2) and pGL3-711(mCF1-1+2) in THP-1.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4529178&req=5

pone.0135141.g010: Functional analysis of the CF1 binding sites within the hFCGRT promoter.(A) Schematic diagram representing the wild-type promoter construct and the mutant promoter constructs. Mutations are marked with crosses. (B) Summarized results of luciferase activity tests. The mutant constructs pGL3-711(mCF1-1), pGL3-711(mCF1-2), and pGL3-711(mCF1-1+2) are derivatives of the wild-type pGL3-711(WT) with mutation in the CF1 site at positions-586, -357, -586 plus -357, respectively. Transfection and normalization were performed as described in the legend to Fig 8. All results represent means ± SD of three to six independent experiments performed in duplicate and plotted as a percentage of the normalized activity of the wild-type pGL3-711(WT) construct, which is defined as 100%. The promoter activity of the labeled construct is significantly decreased compared to the wild-type construct; *P < 0.01 vs. pGL3-711(WT); **P < 0.01 compared with pGL3-711(mCF1-2) and pGL3-711(mCF1-1+2) in THP-1.
Mentions: Functional analysis of the CF1/YY1 binding sites within the hFCGRT promoter is presented in Fig 10.

Bottom Line: Moreover, the CF1/YY1 site at -586 in differentiated THP-1 cells, plays an essential role in the transcriptional activity of the promoter.In addition, the C/EBPbeta binding site at -497 of the hFCGRT promoter in epithelial and endothelial cells, and the C/EBPbeta motif located at -497 and -233 within the hFCGRT promoter in differentiated THP-1 cells may function as positive regulatory sequences in response to LPS or PMA stimulation.EMSA and supershift analyses showed that the functionally identified binding motifs in the hFCGRT promoter were able to specifically interact with their corresponding (Sp1, Sp2, Sp3, c-Fos, c-Jun, YY1, and C/EBPbeta or C/EBPdelta) transcription factors (TFs), suggesting their possible involvement in the regulation of the human FCGRT gene expression.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunochemistry, Ludwik Hirszfeld Institute of Immunology and Experimental Therapy, Polish Academy of Sciences, Wroclaw, Poland.

ABSTRACT
Human epithelial, endothelial and PMA-differentiated THP-1 cell lines were used as model systems to study the transcriptional regulation of the human FCGRT gene encoding the alpha chain of hFcRn. The data obtained from site-directed mutagenesis in transient transfection experiments indicate that the Sp1 sites at positions -641, -635, and -313, CF1/YY1 elements at positions -586 and -357, and the AP-1 motif at -276 within the-660/-233 fragment of the human FCGRT promoter (hFCGRT) participate in the regulation of human FCGRT in all selected cell lines. However, their individual contribution to promoter activity is not equivalent. The Sp1 binding site at -313 and the AP-1 site at -276 are critical for the activity of the hFCGRT promoter in epithelial and endothelial cells. Moreover, the CF1/YY1 site at -586 in differentiated THP-1 cells, plays an essential role in the transcriptional activity of the promoter. In addition, the C/EBPbeta binding site at -497 of the hFCGRT promoter in epithelial and endothelial cells, and the C/EBPbeta motif located at -497 and -233 within the hFCGRT promoter in differentiated THP-1 cells may function as positive regulatory sequences in response to LPS or PMA stimulation. EMSA and supershift analyses showed that the functionally identified binding motifs in the hFCGRT promoter were able to specifically interact with their corresponding (Sp1, Sp2, Sp3, c-Fos, c-Jun, YY1, and C/EBPbeta or C/EBPdelta) transcription factors (TFs), suggesting their possible involvement in the regulation of the human FCGRT gene expression.

No MeSH data available.


Related in: MedlinePlus