Characterization of the 5′-flanking region and regulation of expression of human anion exchanger SLC26A6

Seema Saksena, Alka Dwivedi, Amika Singla, Ravinder K. Gill, Sangeeta Tyagi, Alip Borthakur, Waddah A. Alrefai, Krishnamurthy Ramaswamy, Pradeep K. Dudeja

Research output: Contribution to journalArticle

14 Citations (Scopus)

Abstract

SLC26A6 (putative anion transporter 1, PAT1) has been shown to play an important role in mediating the luminal Cl-/OH-(HCO 3-) exchange process in the intestine. Very little is known about the molecular mechanisms involved in the transcriptional regulation of intestinal SLC26A6 gene expression in the intestine. Current studies were, therefore, designed to clone and characterize the 5′-regulatory region of the human SLC26A6 gene and determine the mechanisms involved in its regulation. A 1,120 bp (p-964/+156) SLC26A6 promoter fragment cloned upstream to the luciferase reporter gene in pGL2-basic exhibited high promoter activity when transfected in Caco2 cells. Progressive deletions of the 5′-flanking region demonstrated that -214/-44 region of the promoter harbors cis-acting elements important for maximal SLC26A6 promoter activity. Since, diarrhea associated with inflammatory bowel diseases is attributed to increased secretion of pro-inflammatory cytokines, we examined the effects of IFNγ (30 ng/ml, 24 h) on SLC26A6 function, expression and promoter activity. IFNγ decreased both SLC26A6 mRNA and function and repressed SLC26A6 promoter activity. Deletion analysis indicated that IFNγ response element is located between -414/-214 region and sequence analysis of this region revealed the presence of potential Interferon Stimulated Responsive Element (ISRE), a binding site (-318/-300 bp) for interferon regulatory factor-1 transcription factor (IRF-1). Mutations in the potential ISRE site abrogated the inhibitory effects of IFNγ. These studies provided novel evidence for the involvement of IRF-1 in the regulation of SLC26A6 gene expression by IFNg in the human intestine.

Original languageEnglish (US)
Pages (from-to)454-466
Number of pages13
JournalJournal of Cellular Biochemistry
Volume105
Issue number2
DOIs
StatePublished - Oct 1 2008

Fingerprint

Interferon Regulatory Factor-1
5' Flanking Region
Gene expression
Interferons
Intestines
Anions
Transcription Factors
Genes
Nucleic Acid Regulatory Sequences
Response Elements
Ports and harbors
Luciferases
Genetic Promoter Regions
Binding Sites
Gene Expression Regulation
Cytokines
Reporter Genes
Inflammatory Bowel Diseases
Messenger RNA
Sequence Analysis

Keywords

  • Human intestine and chloride absorption
  • IFNγ
  • IRF-1
  • Putative Anion Transporter 1 (PAT1) promoter

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Characterization of the 5′-flanking region and regulation of expression of human anion exchanger SLC26A6. / Saksena, Seema; Dwivedi, Alka; Singla, Amika; Gill, Ravinder K.; Tyagi, Sangeeta; Borthakur, Alip; Alrefai, Waddah A.; Ramaswamy, Krishnamurthy; Dudeja, Pradeep K.

In: Journal of Cellular Biochemistry, Vol. 105, No. 2, 01.10.2008, p. 454-466.

Research output: Contribution to journalArticle

Saksena, S, Dwivedi, A, Singla, A, Gill, RK, Tyagi, S, Borthakur, A, Alrefai, WA, Ramaswamy, K & Dudeja, PK 2008, 'Characterization of the 5′-flanking region and regulation of expression of human anion exchanger SLC26A6', Journal of Cellular Biochemistry, vol. 105, no. 2, pp. 454-466. https://doi.org/10.1002/jcb.21842
Saksena, Seema ; Dwivedi, Alka ; Singla, Amika ; Gill, Ravinder K. ; Tyagi, Sangeeta ; Borthakur, Alip ; Alrefai, Waddah A. ; Ramaswamy, Krishnamurthy ; Dudeja, Pradeep K. / Characterization of the 5′-flanking region and regulation of expression of human anion exchanger SLC26A6. In: Journal of Cellular Biochemistry. 2008 ; Vol. 105, No. 2. pp. 454-466.
@article{c4dbee88cd664c49923978bca395836f,
title = "Characterization of the 5′-flanking region and regulation of expression of human anion exchanger SLC26A6",
abstract = "SLC26A6 (putative anion transporter 1, PAT1) has been shown to play an important role in mediating the luminal Cl-/OH-(HCO 3-) exchange process in the intestine. Very little is known about the molecular mechanisms involved in the transcriptional regulation of intestinal SLC26A6 gene expression in the intestine. Current studies were, therefore, designed to clone and characterize the 5′-regulatory region of the human SLC26A6 gene and determine the mechanisms involved in its regulation. A 1,120 bp (p-964/+156) SLC26A6 promoter fragment cloned upstream to the luciferase reporter gene in pGL2-basic exhibited high promoter activity when transfected in Caco2 cells. Progressive deletions of the 5′-flanking region demonstrated that -214/-44 region of the promoter harbors cis-acting elements important for maximal SLC26A6 promoter activity. Since, diarrhea associated with inflammatory bowel diseases is attributed to increased secretion of pro-inflammatory cytokines, we examined the effects of IFNγ (30 ng/ml, 24 h) on SLC26A6 function, expression and promoter activity. IFNγ decreased both SLC26A6 mRNA and function and repressed SLC26A6 promoter activity. Deletion analysis indicated that IFNγ response element is located between -414/-214 region and sequence analysis of this region revealed the presence of potential Interferon Stimulated Responsive Element (ISRE), a binding site (-318/-300 bp) for interferon regulatory factor-1 transcription factor (IRF-1). Mutations in the potential ISRE site abrogated the inhibitory effects of IFNγ. These studies provided novel evidence for the involvement of IRF-1 in the regulation of SLC26A6 gene expression by IFNg in the human intestine.",
keywords = "Human intestine and chloride absorption, IFNγ, IRF-1, Putative Anion Transporter 1 (PAT1) promoter",
author = "Seema Saksena and Alka Dwivedi and Amika Singla and Gill, {Ravinder K.} and Sangeeta Tyagi and Alip Borthakur and Alrefai, {Waddah A.} and Krishnamurthy Ramaswamy and Dudeja, {Pradeep K.}",
year = "2008",
month = "10",
day = "1",
doi = "10.1002/jcb.21842",
language = "English (US)",
volume = "105",
pages = "454--466",
journal = "Journal of Cellular Biochemistry",
issn = "0730-2312",
publisher = "Wiley-Liss Inc.",
number = "2",

}

TY - JOUR

T1 - Characterization of the 5′-flanking region and regulation of expression of human anion exchanger SLC26A6

AU - Saksena, Seema

AU - Dwivedi, Alka

AU - Singla, Amika

AU - Gill, Ravinder K.

AU - Tyagi, Sangeeta

AU - Borthakur, Alip

AU - Alrefai, Waddah A.

AU - Ramaswamy, Krishnamurthy

AU - Dudeja, Pradeep K.

PY - 2008/10/1

Y1 - 2008/10/1

N2 - SLC26A6 (putative anion transporter 1, PAT1) has been shown to play an important role in mediating the luminal Cl-/OH-(HCO 3-) exchange process in the intestine. Very little is known about the molecular mechanisms involved in the transcriptional regulation of intestinal SLC26A6 gene expression in the intestine. Current studies were, therefore, designed to clone and characterize the 5′-regulatory region of the human SLC26A6 gene and determine the mechanisms involved in its regulation. A 1,120 bp (p-964/+156) SLC26A6 promoter fragment cloned upstream to the luciferase reporter gene in pGL2-basic exhibited high promoter activity when transfected in Caco2 cells. Progressive deletions of the 5′-flanking region demonstrated that -214/-44 region of the promoter harbors cis-acting elements important for maximal SLC26A6 promoter activity. Since, diarrhea associated with inflammatory bowel diseases is attributed to increased secretion of pro-inflammatory cytokines, we examined the effects of IFNγ (30 ng/ml, 24 h) on SLC26A6 function, expression and promoter activity. IFNγ decreased both SLC26A6 mRNA and function and repressed SLC26A6 promoter activity. Deletion analysis indicated that IFNγ response element is located between -414/-214 region and sequence analysis of this region revealed the presence of potential Interferon Stimulated Responsive Element (ISRE), a binding site (-318/-300 bp) for interferon regulatory factor-1 transcription factor (IRF-1). Mutations in the potential ISRE site abrogated the inhibitory effects of IFNγ. These studies provided novel evidence for the involvement of IRF-1 in the regulation of SLC26A6 gene expression by IFNg in the human intestine.

AB - SLC26A6 (putative anion transporter 1, PAT1) has been shown to play an important role in mediating the luminal Cl-/OH-(HCO 3-) exchange process in the intestine. Very little is known about the molecular mechanisms involved in the transcriptional regulation of intestinal SLC26A6 gene expression in the intestine. Current studies were, therefore, designed to clone and characterize the 5′-regulatory region of the human SLC26A6 gene and determine the mechanisms involved in its regulation. A 1,120 bp (p-964/+156) SLC26A6 promoter fragment cloned upstream to the luciferase reporter gene in pGL2-basic exhibited high promoter activity when transfected in Caco2 cells. Progressive deletions of the 5′-flanking region demonstrated that -214/-44 region of the promoter harbors cis-acting elements important for maximal SLC26A6 promoter activity. Since, diarrhea associated with inflammatory bowel diseases is attributed to increased secretion of pro-inflammatory cytokines, we examined the effects of IFNγ (30 ng/ml, 24 h) on SLC26A6 function, expression and promoter activity. IFNγ decreased both SLC26A6 mRNA and function and repressed SLC26A6 promoter activity. Deletion analysis indicated that IFNγ response element is located between -414/-214 region and sequence analysis of this region revealed the presence of potential Interferon Stimulated Responsive Element (ISRE), a binding site (-318/-300 bp) for interferon regulatory factor-1 transcription factor (IRF-1). Mutations in the potential ISRE site abrogated the inhibitory effects of IFNγ. These studies provided novel evidence for the involvement of IRF-1 in the regulation of SLC26A6 gene expression by IFNg in the human intestine.

KW - Human intestine and chloride absorption

KW - IFNγ

KW - IRF-1

KW - Putative Anion Transporter 1 (PAT1) promoter

UR - http://www.scopus.com/inward/record.url?scp=54849421781&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=54849421781&partnerID=8YFLogxK

U2 - 10.1002/jcb.21842

DO - 10.1002/jcb.21842

M3 - Article

C2 - 18655181

AN - SCOPUS:54849421781

VL - 105

SP - 454

EP - 466

JO - Journal of Cellular Biochemistry

JF - Journal of Cellular Biochemistry

SN - 0730-2312

IS - 2

ER -