Functional dissection of the promoter of the interphotoreceptor retinoid-binding protein gene

The cone-rod-homeobox element is essential for photoreceptor-specific expression in vivo

Yijian Fei, Suraporn Matragoon, Sylvia B Smith, Paul A. Overbeek, Shiming Chen, Donald J. Zack, Gregory I Liou

Research output: Contribution to journalArticle

20 Citations (Scopus)

Abstract

The essential control elements in the interphotoreceptor retinoid-binding protein gene (IRBP) promoter are located between -156 and +19. The -156/-109 sequence contains a retina-specific DNAse I footprint and shows a positive regulatory activity in transiently transfected retinoblastoma cells. The -105/-85 sequence is G/C rich, shows a non-tissue specific DNAse I hypersensitivity, and a negative regulatory activity in retinoblastoma cells. The -76/-42 sequence shows a retinal-specific footprint and contains a 'cone-rod-homeobox element' (CRXE) and a 'photoreceptor conserved element'' (PCE). IRBP promoter fragments with mutations in either CRXE, PCE or in both were linked to reporter genes and analyzed both by transient transfection and in transgenic mice. In retinoblastoma cells, the mutated CRXE-containing promoter shows a 60% repression of the CAT activity whereas the mutated PCE-containing promoter shows a 30% repression. In HeLa cells transfected with these promoters, co-transfection of a Crx expression vector with wild-type, but not with CRXE mutant promoter, activates CAT activity 20-fold over the background activity. Mutation of PCE alone or conversion of CRXE to PCE reduces this Crx-activated CAT activity to only 4-fold over the background activity. In the transgenic mouse experiments, none of the 12 lines with CRXE mutant promoter show significant expression of lacZ in the retina. In contrast, 9 of the 17 transgenic lines with PCE mutant promoter show photoreceptor-specific lacZ expression. Thus the Crx interaction with CRXE is essential for the photoreceptor-specific activity of the IRBP promoter in vivo. This interaction does not appear to require PCE, but is enhanced when PCE is present.

Original languageEnglish (US)
Pages (from-to)1189-1199
Number of pages11
JournalJournal of Biochemistry
Volume125
Issue number6
DOIs
StatePublished - Jan 1 1999

Fingerprint

Dissection
Vertebrate Photoreceptor Cells
Homeobox Genes
Cones
Genes
Retinoblastoma
Transgenic Mice
Transfection
Rod-Cone Interaction
Retina
Mutation
interstitial retinol-binding protein
Reporter Genes
HeLa Cells
Hypersensitivity

Keywords

  • CRXE
  • IRBP gene regulation
  • PCE
  • Photoreceptor-specific

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology

Cite this

Functional dissection of the promoter of the interphotoreceptor retinoid-binding protein gene : The cone-rod-homeobox element is essential for photoreceptor-specific expression in vivo. / Fei, Yijian; Matragoon, Suraporn; Smith, Sylvia B; Overbeek, Paul A.; Chen, Shiming; Zack, Donald J.; Liou, Gregory I.

In: Journal of Biochemistry, Vol. 125, No. 6, 01.01.1999, p. 1189-1199.

Research output: Contribution to journalArticle

Fei, Yijian ; Matragoon, Suraporn ; Smith, Sylvia B ; Overbeek, Paul A. ; Chen, Shiming ; Zack, Donald J. ; Liou, Gregory I. / Functional dissection of the promoter of the interphotoreceptor retinoid-binding protein gene : The cone-rod-homeobox element is essential for photoreceptor-specific expression in vivo. In: Journal of Biochemistry. 1999 ; Vol. 125, No. 6. pp. 1189-1199.
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abstract = "The essential control elements in the interphotoreceptor retinoid-binding protein gene (IRBP) promoter are located between -156 and +19. The -156/-109 sequence contains a retina-specific DNAse I footprint and shows a positive regulatory activity in transiently transfected retinoblastoma cells. The -105/-85 sequence is G/C rich, shows a non-tissue specific DNAse I hypersensitivity, and a negative regulatory activity in retinoblastoma cells. The -76/-42 sequence shows a retinal-specific footprint and contains a 'cone-rod-homeobox element' (CRXE) and a 'photoreceptor conserved element'' (PCE). IRBP promoter fragments with mutations in either CRXE, PCE or in both were linked to reporter genes and analyzed both by transient transfection and in transgenic mice. In retinoblastoma cells, the mutated CRXE-containing promoter shows a 60{\%} repression of the CAT activity whereas the mutated PCE-containing promoter shows a 30{\%} repression. In HeLa cells transfected with these promoters, co-transfection of a Crx expression vector with wild-type, but not with CRXE mutant promoter, activates CAT activity 20-fold over the background activity. Mutation of PCE alone or conversion of CRXE to PCE reduces this Crx-activated CAT activity to only 4-fold over the background activity. In the transgenic mouse experiments, none of the 12 lines with CRXE mutant promoter show significant expression of lacZ in the retina. In contrast, 9 of the 17 transgenic lines with PCE mutant promoter show photoreceptor-specific lacZ expression. Thus the Crx interaction with CRXE is essential for the photoreceptor-specific activity of the IRBP promoter in vivo. This interaction does not appear to require PCE, but is enhanced when PCE is present.",
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