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A neuronal-specific differentiation protein that directly modulates retinoid receptor transcriptional activation.

Henry II KW, Spencer ML, Theodosiou M, Lou D, Noonan DJ - Nucl. Recept. (2003)

Bottom Line: Expression of NPDC-1 was also observed to repress transcription mediated by retinoid receptors as well as by several other nuclear receptor family members, although not in a universal manner.CONCLUSIONS: These results suggest that NPDC-1, through direct interaction with retinoid receptors, functions to enhance the transcription complex formation and DNA binding function of retinoid receptors, but ultimately repress retinoid receptor-mediated gene expression.As with NPDC-1, retinoids and their receptors have been implicated in brain development and these data provide a point of convergence for NPDC-1 and retinoid mediation of neuronal differentiation.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Molecular and Cellular Biochemistry, University of Kentucky, 800 Rose Street, Lexington, KY 40536, USA. dnoonan@pop.uky.edu

ABSTRACT
BACKGROUND: The specificity of a nuclear receptor's ability to modulate gene expression resides in its ability to bind a specific lipophilic ligand, associate with specific dimerization partners and bind specific DNA sequences in the promoter regions of genes. This sequence of events appears to be the basis for targeting an additional regulatory complex composed of a variety of protein and RNA components that deliver signals for facilitation or inhibition of the RNA polymerase complex. Characterization of the tissue and cell-specific components of these coregulatory complexes appear to be integral to our understanding of nuclear receptor regulation of transcription. RESULTS: A novel yeast screen sensitive to retinoid-X receptor (RXR) transcriptional activation resulted in the isolation of the rat homologue of the mouse NPDC-1 gene. NPDC-1 has been shown to be involved in the control of neural cell proliferation and differentiation, possibly through interactions with the cell cycle promoting transcription factor E2F-1. Although the amino acid sequence of NPDC-1 is highly conserved between mouse, rat and human homologues, their tissue specific expression was seen to vary. A potential for direct protein:protein interaction between NPDC-1, RXR and retinoic acid receptor beta (RARbeta) was observed in vitro and NPDC-1 facilitated RXR homodimer and RAR-RXR heterodimer DNA binding in vitro. Expression of NPDC-1 was also observed to repress transcription mediated by retinoid receptors as well as by several other nuclear receptor family members, although not in a universal manner. CONCLUSIONS: These results suggest that NPDC-1, through direct interaction with retinoid receptors, functions to enhance the transcription complex formation and DNA binding function of retinoid receptors, but ultimately repress retinoid receptor-mediated gene expression. As with NPDC-1, retinoids and their receptors have been implicated in brain development and these data provide a point of convergence for NPDC-1 and retinoid mediation of neuronal differentiation.

No MeSH data available.


Related in: MedlinePlus

NPDC-1 Alters RARβ••••α and RXRα DNA binding properties. Oligonucleotides corresponding to the consensus RARβ•RARE (DR-5: A) or the consensus RXRα RARE (DR-1: B & C) were end-labeled with 32-P by T4 polynucleotide kinase according to the manufacture's instructions. The resultant DNA binding probes were incubated with PC12 lysate (A), recombinant NPDC-1 and recombinant RXRα (B & C) as indicated. DNA:protein complexes were resolved on non-denaturing PAGE gels which were subsequently exposed to Kodak Xar-5 film. In C, the presence of NPDC-1 within the RXR gel-shift complex was assayed by supershift. Antibodies specific for either NPDC-1 or RXR were used to identify proteins present in the complex.
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Figure 6: NPDC-1 Alters RARβ••••α and RXRα DNA binding properties. Oligonucleotides corresponding to the consensus RARβ•RARE (DR-5: A) or the consensus RXRα RARE (DR-1: B & C) were end-labeled with 32-P by T4 polynucleotide kinase according to the manufacture's instructions. The resultant DNA binding probes were incubated with PC12 lysate (A), recombinant NPDC-1 and recombinant RXRα (B & C) as indicated. DNA:protein complexes were resolved on non-denaturing PAGE gels which were subsequently exposed to Kodak Xar-5 film. In C, the presence of NPDC-1 within the RXR gel-shift complex was assayed by supershift. Antibodies specific for either NPDC-1 or RXR were used to identify proteins present in the complex.

Mentions: Ligand induced RXR homodimers and RAR/RXR heterodimers will form in solution and are capable of binding to specific HRE DNA sequences [33-35]. To investigate the impact NPDC-1 has on nuclear receptor DNA binding, recombinant human NPDC-1 was analyzed for its ability to modulate gel mobility shift complexes generated using PC12 cell nuclear extracts and a radiolabeled DR-5 βRARE DNA response element (Fig. 6A). The βRARE generated two major shift complexes in the presence of PC12 extracts. The addition of recombinant hNPDC was observed to substantially enhance the formation of the lower shift complex at the expense of the higher shift complex. NPDC-1 did not form a measurable unique complex with the radiolabeled DNA. Curiously, the enhanced protein:DNA complex formed without a significant change in mobility indicating that NPDC-1 facilitates DNA binding without remaining in strong association with either proteins or DNA.


A neuronal-specific differentiation protein that directly modulates retinoid receptor transcriptional activation.

Henry II KW, Spencer ML, Theodosiou M, Lou D, Noonan DJ - Nucl. Recept. (2003)

NPDC-1 Alters RARβ••••α and RXRα DNA binding properties. Oligonucleotides corresponding to the consensus RARβ•RARE (DR-5: A) or the consensus RXRα RARE (DR-1: B & C) were end-labeled with 32-P by T4 polynucleotide kinase according to the manufacture's instructions. The resultant DNA binding probes were incubated with PC12 lysate (A), recombinant NPDC-1 and recombinant RXRα (B & C) as indicated. DNA:protein complexes were resolved on non-denaturing PAGE gels which were subsequently exposed to Kodak Xar-5 film. In C, the presence of NPDC-1 within the RXR gel-shift complex was assayed by supershift. Antibodies specific for either NPDC-1 or RXR were used to identify proteins present in the complex.
© Copyright Policy
Related In: Results  -  Collection

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Figure 6: NPDC-1 Alters RARβ••••α and RXRα DNA binding properties. Oligonucleotides corresponding to the consensus RARβ•RARE (DR-5: A) or the consensus RXRα RARE (DR-1: B & C) were end-labeled with 32-P by T4 polynucleotide kinase according to the manufacture's instructions. The resultant DNA binding probes were incubated with PC12 lysate (A), recombinant NPDC-1 and recombinant RXRα (B & C) as indicated. DNA:protein complexes were resolved on non-denaturing PAGE gels which were subsequently exposed to Kodak Xar-5 film. In C, the presence of NPDC-1 within the RXR gel-shift complex was assayed by supershift. Antibodies specific for either NPDC-1 or RXR were used to identify proteins present in the complex.
Mentions: Ligand induced RXR homodimers and RAR/RXR heterodimers will form in solution and are capable of binding to specific HRE DNA sequences [33-35]. To investigate the impact NPDC-1 has on nuclear receptor DNA binding, recombinant human NPDC-1 was analyzed for its ability to modulate gel mobility shift complexes generated using PC12 cell nuclear extracts and a radiolabeled DR-5 βRARE DNA response element (Fig. 6A). The βRARE generated two major shift complexes in the presence of PC12 extracts. The addition of recombinant hNPDC was observed to substantially enhance the formation of the lower shift complex at the expense of the higher shift complex. NPDC-1 did not form a measurable unique complex with the radiolabeled DNA. Curiously, the enhanced protein:DNA complex formed without a significant change in mobility indicating that NPDC-1 facilitates DNA binding without remaining in strong association with either proteins or DNA.

Bottom Line: Expression of NPDC-1 was also observed to repress transcription mediated by retinoid receptors as well as by several other nuclear receptor family members, although not in a universal manner.CONCLUSIONS: These results suggest that NPDC-1, through direct interaction with retinoid receptors, functions to enhance the transcription complex formation and DNA binding function of retinoid receptors, but ultimately repress retinoid receptor-mediated gene expression.As with NPDC-1, retinoids and their receptors have been implicated in brain development and these data provide a point of convergence for NPDC-1 and retinoid mediation of neuronal differentiation.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Molecular and Cellular Biochemistry, University of Kentucky, 800 Rose Street, Lexington, KY 40536, USA. dnoonan@pop.uky.edu

ABSTRACT
BACKGROUND: The specificity of a nuclear receptor's ability to modulate gene expression resides in its ability to bind a specific lipophilic ligand, associate with specific dimerization partners and bind specific DNA sequences in the promoter regions of genes. This sequence of events appears to be the basis for targeting an additional regulatory complex composed of a variety of protein and RNA components that deliver signals for facilitation or inhibition of the RNA polymerase complex. Characterization of the tissue and cell-specific components of these coregulatory complexes appear to be integral to our understanding of nuclear receptor regulation of transcription. RESULTS: A novel yeast screen sensitive to retinoid-X receptor (RXR) transcriptional activation resulted in the isolation of the rat homologue of the mouse NPDC-1 gene. NPDC-1 has been shown to be involved in the control of neural cell proliferation and differentiation, possibly through interactions with the cell cycle promoting transcription factor E2F-1. Although the amino acid sequence of NPDC-1 is highly conserved between mouse, rat and human homologues, their tissue specific expression was seen to vary. A potential for direct protein:protein interaction between NPDC-1, RXR and retinoic acid receptor beta (RARbeta) was observed in vitro and NPDC-1 facilitated RXR homodimer and RAR-RXR heterodimer DNA binding in vitro. Expression of NPDC-1 was also observed to repress transcription mediated by retinoid receptors as well as by several other nuclear receptor family members, although not in a universal manner. CONCLUSIONS: These results suggest that NPDC-1, through direct interaction with retinoid receptors, functions to enhance the transcription complex formation and DNA binding function of retinoid receptors, but ultimately repress retinoid receptor-mediated gene expression. As with NPDC-1, retinoids and their receptors have been implicated in brain development and these data provide a point of convergence for NPDC-1 and retinoid mediation of neuronal differentiation.

No MeSH data available.


Related in: MedlinePlus