Document Detail


Two polypyrimidine tracts in the nitric oxide synthase 2 gene: similar regulatory sequences with different properties.
MedLine Citation:
PMID:  19669598     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
We reported previously that the polymorphic polypyrimidine CCTTT-microsatellite in the regulatory region of nitric oxide synthase 2 (NOS2) bound nuclear proteins in vitro. In the present work, we aimed to characterize and investigate a potential regulatory role of the CCTTT-microsatellite in NOS2 expression. Therefore, we performed gel-shift, S1-nuclease, and chromatin immunoprecipitation (ChIP) assays. In vitro experiments showed that the microsatellite formed triplex-DNA both with and without superhelical constraint. We also found that the CCTTT-microsatellite and an apparently similar CT-repeat in the first intron of NOS2 were specifically cleaved by S1-nuclease, when cloned into a supercoiled plasmid. In vitro data suggested that the CCTTT-microsatellite bound both polypyrimidine tract-binding protein (PTBP1) and heterogeneous nuclear ribonucleoprotein K (hnRNPK). On the contrary, ChIP revealed binding of PTBP1 and hnRNPK rather to the CT-repeat in the first intron than to the CCTTT-microsatellite. Enrichment for RNA polymerase II and acetylated histones H3 and H4 was also detected at the intronic site. We suggest that both PTBP1 and hnRNPK binds the single strand of the triplex-DNA formed at the CT-repeat in the first intron and that this interaction could be involved in the regulation of NOS2 expression.
Authors:
Mehdi Motallebipour; Alvaro Rada-Iglesias; Gunnar Westin; Claes Wadelius
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't     Date:  2009-08-08
Journal Detail:
Title:  Molecular biology reports     Volume:  37     ISSN:  1573-4978     ISO Abbreviation:  Mol. Biol. Rep.     Publication Date:  2010 Apr 
Date Detail:
Created Date:  2010-03-03     Completed Date:  2010-06-08     Revised Date:  2011-10-27    
Medline Journal Info:
Nlm Unique ID:  0403234     Medline TA:  Mol Biol Rep     Country:  Netherlands    
Other Details:
Languages:  eng     Pagination:  2021-30     Citation Subset:  IM    
Affiliation:
Department of Genetics and Pathology, Rudbeck Laboratory, Uppsala University, 751 85, Uppsala, Sweden.
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MeSH Terms
Descriptor/Qualifier:
Base Sequence
Hep G2 Cells
Heterogeneous-Nuclear Ribonucleoproteins / metabolism
Histones / metabolism
Humans
Microsatellite Repeats / genetics
Molecular Sequence Data
Nitric Oxide Synthase Type II / genetics*,  metabolism
Nucleic Acid Conformation
Polymerase Chain Reaction
Protein Binding
Regulatory Sequences, Nucleic Acid / genetics*
Repetitive Sequences, Nucleic Acid / genetics
Chemical
Reg. No./Substance:
0/Heterogeneous-Nuclear Ribonucleoproteins; 0/Histones; EC 1.14.13.39/NOS2 protein, human; EC 1.14.13.39/Nitric Oxide Synthase Type II

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