Abstract
RET gene expression is characterized by high tissue and stage specificity during the development of neural crest derivatives and in the pathogenesis of inherited cancer syndromes and Hirschsprung disease. Identifying all elements contributing to its transcriptional regulation might provide new clues to clarify both developmental and pathogenic mechanisms. We previously demonstrated that chromatin acetylation affects RET transcription; therefore, we have set up a strategy based on analysis of sequences conserved among species at the RET locus, combined with the characterization of their chromatin structure, to identify new potential regulatory elements. The histone acetylation level was evaluated by the chromatin immunoprecipitation method applied to cells displaying different degrees of endogenous RET expression. Real-time quantitative PCR of immunoprecipitated DNA-protein complexes and transfection experiments, with constructs expressing a reporter gene in which the putative regulatory regions are inserted, indicate a correlation between histone acetylation and endogenous RET expression and highlight conserved sequences with potential regulatory roles. This paper presents a reliable screening procedure to unearth elements able to affect gene regulation at the transcriptional level in a large genomic region.
| Original language | English |
|---|---|
| Pages (from-to) | 269-274 |
| Number of pages | 6 |
| Journal | Physiol Genomics |
| Volume | 23 |
| Issue number | 3 |
| DOIs | |
| Publication status | Published - Oct 17 2005 |
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Keywords
- Comparative sequence analysis
- Hirschsprung disease
- Histone acetylation
- Real-time quantitative PCR
- Transcriptional regulation
ASJC Scopus subject areas
- Physiology
- Genetics
Cite this
Comparative genomic sequence analysis coupled to chromatin immunoprecipitation : A screening procedure applied to search for regulatory elements at the RET locus. / Puppo, Francesca; Musso, Marco; Pirulli, Doroti; Griseri, Paola; Bachetti, Tiziana; Crovella, Sergio; Patrone, Giovanna; Ceccherini, Isabella; Ravazzolo, Roberto.
In: Physiol Genomics, Vol. 23, No. 3, 17.10.2005, p. 269-274.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Comparative genomic sequence analysis coupled to chromatin immunoprecipitation
T2 - A screening procedure applied to search for regulatory elements at the RET locus
AU - Puppo, Francesca
AU - Musso, Marco
AU - Pirulli, Doroti
AU - Griseri, Paola
AU - Bachetti, Tiziana
AU - Crovella, Sergio
AU - Patrone, Giovanna
AU - Ceccherini, Isabella
AU - Ravazzolo, Roberto
PY - 2005/10/17
Y1 - 2005/10/17
N2 - RET gene expression is characterized by high tissue and stage specificity during the development of neural crest derivatives and in the pathogenesis of inherited cancer syndromes and Hirschsprung disease. Identifying all elements contributing to its transcriptional regulation might provide new clues to clarify both developmental and pathogenic mechanisms. We previously demonstrated that chromatin acetylation affects RET transcription; therefore, we have set up a strategy based on analysis of sequences conserved among species at the RET locus, combined with the characterization of their chromatin structure, to identify new potential regulatory elements. The histone acetylation level was evaluated by the chromatin immunoprecipitation method applied to cells displaying different degrees of endogenous RET expression. Real-time quantitative PCR of immunoprecipitated DNA-protein complexes and transfection experiments, with constructs expressing a reporter gene in which the putative regulatory regions are inserted, indicate a correlation between histone acetylation and endogenous RET expression and highlight conserved sequences with potential regulatory roles. This paper presents a reliable screening procedure to unearth elements able to affect gene regulation at the transcriptional level in a large genomic region.
AB - RET gene expression is characterized by high tissue and stage specificity during the development of neural crest derivatives and in the pathogenesis of inherited cancer syndromes and Hirschsprung disease. Identifying all elements contributing to its transcriptional regulation might provide new clues to clarify both developmental and pathogenic mechanisms. We previously demonstrated that chromatin acetylation affects RET transcription; therefore, we have set up a strategy based on analysis of sequences conserved among species at the RET locus, combined with the characterization of their chromatin structure, to identify new potential regulatory elements. The histone acetylation level was evaluated by the chromatin immunoprecipitation method applied to cells displaying different degrees of endogenous RET expression. Real-time quantitative PCR of immunoprecipitated DNA-protein complexes and transfection experiments, with constructs expressing a reporter gene in which the putative regulatory regions are inserted, indicate a correlation between histone acetylation and endogenous RET expression and highlight conserved sequences with potential regulatory roles. This paper presents a reliable screening procedure to unearth elements able to affect gene regulation at the transcriptional level in a large genomic region.
KW - Comparative sequence analysis
KW - Hirschsprung disease
KW - Histone acetylation
KW - Real-time quantitative PCR
KW - Transcriptional regulation
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UR - http://www.scopus.com/inward/citedby.url?scp=31744441602&partnerID=8YFLogxK
U2 - 10.1152/physiolgenomics.00036.2005
DO - 10.1152/physiolgenomics.00036.2005
M3 - Article
C2 - 16144862
AN - SCOPUS:31744441602
VL - 23
SP - 269
EP - 274
JO - Physiological Genomics
JF - Physiological Genomics
SN - 1094-8341
IS - 3
ER -