Adipocyte-to-osteoblast reprogramming in ST2 cells. Single-Cell RNA Sequencing of ST2 Cells Undergoing Transcription Factor Perturbations to Validate Boolean Network Predictions of Adipocyte-to-Osteoblast Reprogramming

This study presents single-cell RNA sequencing (scRNA-seq) data from differentiated ST2 mesenchymal cells subjected to targeted transcription factor perturbations to experimentally validate predictions derived from Boolean network modeling. The modeling suggested specific transcription factors that, when perturbed, could reprogram adipocytes into osteoblast-like cells. ST2 cells were differentiated into adipogenic lineages, with selected transcription factors—CEBPA, SP1, TCF7L2, and TRP63—either knocked down or overexpressed using lentiviral constructs and siRNA-mediated gene repression. This was followed by transfer to osteogenic culture conditions to evaluate reprogramming capacity of different transcription factor combinations. scRNA-seq was conducted using the Chromium Next GEM Single Cell 3’ Kit to capture the transcriptome of individual cells at multiple stages of differentiation. Data were processed to identify differentially expressed genes in reprogrammed cells compared to controls, assessing the fidelity of predicted transcriptional changes. This dataset provides insights into the transcriptional landscape underlying adipocyte-to-osteoblast reprogramming and serves as a resource to assess the effectiveness of computational predictions in guiding cellular differentiation.