RNA sequencing data of FACS sorted SVF cells derived from euthyroid and hypothyroid Zfp423GFP mice in the preadipocyte stage and after adipogenic differentiation

Stromal vascular fraction from inguinal adipose tissue of hypothyroid and euthyroid Zfp423GFP mice was isolated and sorted into GFP+ and GFP- cells using flow cytometry. Cell populations were divided into two parts. The first part was harvested after cells reached 80 perc. confluence (approximately after 3-4 days), reflecting cells in the preadipocyte stage. The second part was incubated two times for 48 hours with adipogenic differentiation medium (culture medium supplemented with 30 nM insulin, 1 micrometer rosiglitazone and 0.4 microgram/ml dexamethasone) followed by two times for 48 hours in culture medium, reflecting cells in the differentiated stage. Cells derived from hypothyroid or euthyroid mice were cultured using thyroid hormone (TH) depleted FBS or TH-depleted FBS supplemented with 10nM T3, respectively, to maintain the hypothyroid or euthyroid state in vitro.50ng of total RNA were depleted of ribosomal RNA using the NEBNext rRNA Depletion Kit v2 (NEB) according to the instructions of the manufacturer. Depleted RNA was transcribed using SuperScript IV reverse transcriptase (ThermoFisher) for 2 h at 55C. After second strand synthesis (TargetAmp kit (Epicentre)) the DNA was tagmented using the Illumina Tagment DNA TDE1 Enzyme and Buffer Kits, which fragments DNA and inserts partial sequencing adapter sequences. Final PCR amplification of the libraries was done using KAPA HiFi HotStart Library Amplification Kit with unique dual indexing by IDT for Illumina Nextera DNA Unique Dual Indexes Sets. The barcoded libraries were purified and quantified using Qubit Fluorometric Quantification (ThermoFischer Scientific). Size distribution of the library DNA was analyzed using the FragmentAnalyzer (Agilent). Sequencing of 2x150 bp was performed with an NovaSeq sequencer (Illumina) according to the instructions of the manufacturer.We observed distinct transcriptional profiles during adipogenesis dependent on ZFP423 expression and TH state. Particularly, ZFP423-dependent genes exclusively regulated in the hypothyroid state were related to mitochondrial function and metabolic pathways. Thus, this data give first evidence that transcriptional upregulation of Zfp423 in TH-deficient mice might modulate adipocyte function in regard of lipid storing and thermogenesis.