A fast carrier chromatin immunoprecipitation method applicable to microdissected tissue samples.
Transcriptional regulation studies of CNS neurons are complicated by both cellular diversity and plasticity. Microdissection of specific functionally related populations of neurons can greatly reduce these issues, but typically excludes the use of many technologies due to tissue requirements, such as Chromatin Immunoprecipitation (ChIP), a powerful tool for studying in vivo protein-DNA interactions. We have developed a fast carrier ChIP (Fast CChIP) method for analyzing specific in vivo transcription factor-DNA interactions in as little as 0.2 mm(3) brain tissue. Using an antibody against phosphorylated cyclic-AMP response element binding (CREB) protein, we confirmed phospho-CREB (pCREB) binding at the c-fos gene promoter. Then we further demonstrated the applicability of Fast CChIP in determining hypertension-induced pCREB binding at the c-fos gene promoter in the rat nucleus tractus solitarius (NTS), confirming CREB's role in mediating hypertension-induced c-fos expression. This method will be broadly applicable to individual brain nucleus and biopsy/surgical samples.
Hao
Haiping
H
Daniel Baugh Institute for Functional Genomics and Computational Biology, Department of Pathology, Anatomy and Cell Biology, Thomas Jefferson University, Philadelphia, PA 19107, United States.
Liu
Hester
H
Gonye
Gregory
G
Schwaber
James S
JS
D000818:Animals; D001921:Brain / Q000166:cytology / Q000378:metabolism*; D047369:Chromatin Immunoprecipitation / Q000379:methods*; D017362:Cyclic AMP Response Element-Binding Protein / Q000235:genetics / Q000378:metabolism; D004195:Disease Models, Animal; D006973:Hypertension / Q000473:pathology / Q000503:physiopathology; D008297:Male; D042282:Microdissection / Q000379:methods*; D016760:Proto-Oncogene Proteins c-fos / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D015533:Transcriptional Activation / Q000502:physiology
D017362:Cyclic AMP Response Element-Binding Protein; D016760:Proto-Oncogene Proteins c-fos