Pitfalls in global normalization of ChIP-seq data in CD4(+) T cells treated with butyrate: A possible solution strategy.
Regulatory T cells (Treg) play a central role in the suppression of inflammatory and allergic responses. Colonization of certain gut commensal microbes such as Clostridia class IV and XIVa in the gut can induce development of colonic Treg cells contributing to the maintenance of gut immune homeostasis. Clostridia-derived butyrate promotes the differentiation of naïve T cells into Treg cells through upregulation of Foxp3, the master transcription factor of Treg cells. Chromatin immunoprecipitation-sequencing (ChIP-seq) analysis revealed that treatment of naïve T cells with butyrate induces Treg-polarizing conditions by enhanced histone H3 acetylation in the promoter and conserved non-coding sequence regions of the Foxp3 locus. In general, global normalization was utilized for ChIP-seq analysis to compare the data obtained from two or more samples. However, global normalization is not appropriate for the evaluation of ChIP-seq data when treatment can affect the total amount of target protein. Here, we introduce a unique normalization method for ChIP-seq analysis in cells treated with butyrate, a pan-HDAC inhibitor that is likely to affect total acetylation levels of histone H3.
Graduate School of Pharmaceutical Science, Keio University, Tokyo 105-8512, Japan ; The Institute of Medical Science, The University of Tokyo, Tokyo 108 8639, Japan ; RIKEN Center for Integrative Medical Sciences (IMS-RCAI), Kanagawa 230 0045, Japan.
Yukihiro
Y
Furusawa
RIKEN Center for Integrative Medical Sciences (IMS-RCAI), Kanagawa 230 0045, Japan.
Takaho A
TA
Endo
Graduate School of Pharmaceutical Science, Keio University, Tokyo 105-8512, Japan ; The Institute of Medical Science, The University of Tokyo, Tokyo 108 8639, Japan ; RIKEN Center for Integrative Medical Sciences (IMS-RCAI), Kanagawa 230 0045, Japan ; Graduate School of Medicine, Chiba University, Chiba 260 8670, Japan.
Yuuki
Y
Obata
RIKEN Center for Integrative Medical Sciences (IMS-RCAI), Kanagawa 230 0045, Japan.
Osamu
O
Ohara
RIKEN Center for Integrative Medical Sciences (IMS-RCAI), Kanagawa 230 0045, Japan ; Graduate School of Medicine, Chiba University, Chiba 260 8670, Japan ; Graduate School of Medical Life Science, Yokohama City University, Kanagawa 230 0045, Japan.
Hiroshi
H
Ohno
Graduate School of Pharmaceutical Science, Keio University, Tokyo 105-8512, Japan ; The Institute of Medical Science, The University of Tokyo, Tokyo 108 8639, Japan ; RIKEN Center for Integrative Medical Sciences (IMS-RCAI), Kanagawa 230 0045, Japan.
Koji
K
Hase
Butyrate; ChIP-seq; HDAC; Microarray; Normalization; Regulatory T cells