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Chromatin interaction of Cebpa during adipocyte differentiation (4C-Seq)

ABSTRACT: Cebpa is a critical transcription factor gene for adipocyte differentiation and adipose tissue development. However, mechanisms controlling Cebpa expression during adipogenic differentiation remain largely unknown. Here, we generated the high-resolution chromatin interaction maps of Cebpa in 3T3-L1 preadipocytes (3T3-L1) and 3T3-L1 adipocytes (3T3-L1-AD) using circularized chromosome conformation capture coupled with next-generation sequencing (4C-seq), and characterized differences in their chromatin interactomes and chromatin status of the interaction sites during adipogenic differentiation. We performed a 4C-seq experiment on inguinal white adipose tissue (iWAT) to evaluate whether chromatin interaction between Cebpa-L1-AD-En2 and Cebpa promoters in 3T3-L1 adipocytes also exists in mouse adipose tissue.

ORGANISM(S): Mus musculus

PROVIDER: GSE222688 | GEO | 2024/03/07

REPOSITORIES: GEO

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Project description:Adipose tissues are closely related to physiological functions and pathological conditions in most organs. Although differentiated 3T3-L1 preadipocytes have been used for in vitro adipose studies, the difference in cellular characteristics of adipogenic differentiation in two-dimensional (2D) culture and three-dimensional (3D) culture remain unclear. In this study, we evaluated gene expression patterns using RNA sequencing and metabolic functions using an extracellular flux analyzer in 3T3-L1 preadipocytes with and without adipogenic induction in 2D culture and 3D culture. In 2D culture, 565 up-regulated genes and 391 down-regulated genes were identified as differentially expressed genes (DEGs) by adipogenic induction of 3T3-L1 preadipocytes, whereas only 69 up-regulated genes and 59 down-regulated genes were identified as DEGs in 3D culture. Ingenuity Pathway Analysis (IPA) revealed that genes associated with lipid metabolism were identified as 2 out of the top 3 causal networks related to diseases and function in 3D spheroids, whereas only one network related to lipid metabolism was identified within the top 9 of these causal networks in the 2D planar cells, suggesting that adipogenic induction in the 3D culture condition exhibits a more adipocyte-specific gene expression pattern in 3T3-L1 preadipocytes. Real-time metabolic analysis revealed that the metabolic capacity shifted from glycolysis to mitochondrial respiration in differentiated 3T3-L1 cells in the 3D culture condition but not in those in the 2D cultured condition, suggesting that adipogenic differentiation in 3D culture induces a metabolic phenotype of well-differentiated adipocytes. Consistently, expression levels of mitochondria-encoded genes including mt-Nd6, mt-Cytb, and mt-Co1 were significantly increased by adipogenic induction of 3T3-L1 preadipocytes in 3D culture compared with those in 2D culture. Taken together, the findings suggest that induction of adipogenesis in 3D culture provides a more adipocyte-specific gene expression pattern and enhances mitochondrial respiration, resulting in more adipocyte-like cellular properties.