Chromatin immunoprecipitation with mouse adipocytes using hypotonic buffer to enrich nuclear fraction before fixation.

STAR Protoc

Division for Health Service Promotion, The University of Tokyo, Tokyo 113-0033, Japan; The University of Tokyo Excellent Young Researcher Program, The University of Tokyo, Tokyo 113-8654, Japan; Department of Diabetes and Metabolic Diseases, Graduate School of Medicine, The University of Tokyo, Toky

Published: March 2023


Category Ranking

98%

Total Visits

921

Avg Visit Duration

2 minutes

Citations

20

Article Abstract

Chromatin immunoprecipitation (ChIP) experiments with differentiated adipocytes are challenging because lipid droplets interfere with immunoprecipitation efficiency. Here, the author describes optimized procedures to minimize the burden of lipid droplets by using hypotonic buffer to enrich nuclear fraction before formaldehyde crosslinking, thus increasing the sensitivity and specificity of ChIP experiments with differentiated adipocytes. The author also describes steps after fixation, including sonication, immunoprecipitation, washing, reverse-crosslinking, and purification. This protocol is compatible with ChIP-qPCR and ChIP-seq of various transcription factors and histone modifications. For complete details on the use and execution of this protocol, please refer to Hiraike et al. (2022)..

Download full-text PDF

Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC9937945PMC
http://dx.doi.org/10.1016/j.xpro.2023.102093DOI Listing

Publication Analysis

Top Keywords

chromatin immunoprecipitation
8
hypotonic buffer
8
buffer enrich
8
enrich nuclear
8
nuclear fraction
8
chip experiments
8
experiments differentiated
8
differentiated adipocytes
8
lipid droplets
8
author describes
8

Similar Publications

NFATc3 and PML Synergistically Regulate Tumor-Associated Gene Expression in a SUMOylation-Independent Manner.

Biochimie

September 2025

Department of Oncology, Xin Hua Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai 200092, China. Electronic address:

The nuclear factor of activated T cells 3 (NFATc3) plays a significant role in various cancer-related processes, but its interactions with transcriptional modulators, particularly Promyelocytic Leukemia protein (PML), remain poorly understood. PML, a nuclear scaffold protein, is involved in tumor suppression and transcriptional regulation. This study investigates the interaction between NFATc3 and PML, focusing on the role of SUMOylation and its impact on downstream target genes.

View Article and Find Full Text PDF

A clinical and genotype-phenotype analysis of MACF1 variants.

Am J Hum Genet

September 2025

Department of Clinical Genetics, Erasmus MC, University Medical Center Rotterdam, PO Box 2040, Rotterdam 3000 CA, the Netherlands.

Microtubule-actin cross-linking factor 1 (MACF1) is a large protein of the spectraplakin family, which is essential for brain development. MACF1 interacts with microtubules through the growth arrest-specific 2 (Gas2)-related (GAR) domain. Heterozygous MACF1 missense variants affecting the zinc-binding residues in this domain result in a distinctive cortical and brain stem malformation.

View Article and Find Full Text PDF

Salt stress impairs photosynthetic efficiency and consequently reduces the growth, development, and grain yield of crop plants. The formation of hydrophobic barriers in the root endodermis, including the suberin lamellae and Casparian strips, is a key adaptive strategy for salt stress tolerance. In this study, we identified the role of the rice NAC transcription factor, ONAC005, in salt stress tolerance.

View Article and Find Full Text PDF

Introduction: Epigenetic changes are important modulators of gene expression. The histone acetyltransferase gene non-derepressible 5 (Gcn5) is emerging as a pivotal epigenetic player in metabolism and cancer, yet its role in obesity and cardiovascular disease remains elusive.

Aims: To investigate Gcn5 role in obesity-related endothelial dysfunction.

View Article and Find Full Text PDF

Hepatitis B virus (HBV) precore G1896A mutation is closely associated with poor prognosis of liver disease. We previously revealed that the G1896A mutation could enhance HBV replication and promote hepatocellular carcinoma (HCC) cell growth both in vitro and in vivo. However, the in-depth mechanisms by which this mutation promotes the malignancy of HCC still need to be explored.

View Article and Find Full Text PDF