Transcriptome analysis of IMR90 fibroblasts expressing OSKM or control vector and OSKM-induced senescent IMR90 fibroblasts upon p21CIP or mTOR knockdown

Ectopic expression of the transcription factors Oct4, Sox2, Klf4 and c-Myc (OSKM) can reprogram somatic cells into induced pluripotent stem cells (iPSCs). These iPSCs are highly similar to embryonic stem cells and can be used for regenerative medicine, drug screening and disease modelling. Despite recent advances, reprogramming is a slow and inefficient process. This suggests that there are several safeguarding mechanisms to counteract cell fate conversion. Cellular senescence is one of these barriers, which is mediated through activation of the tumour suppressors p53/p21CIP1, p15INK4b and p16INK4a. In this study, we have screened for shRNAs blunting reprogramming-induced senescence. To investigate the effects of OSKM expression on the transcriptome of human primary IMR90 fibroblasts, we performed RNA-sequencing (RNA-Seq). Cells were transduced with OSKM expression or control vector and RNA was extracted after 14 or 20 days. In addition, to study the transcriptome of cells bypassing OSKM-induced senescence due to p21 or mTOR depletion, we performed RNA-seq of cells transduced with OSKM and shRNA expressing vectors targeting p21 or mTOR for 14 or 20 days. Overall design: 8 samples in triplicate: IMR90 + OSKM + shRNA targeting p21CIP1 for 20 days (O211_1) or 14 days (O211_2), IMR90 + OSKM + different shRNA targeting p21CIP1 for 20 days (O212_1) or 14 days (O212_2), IMR90 + OSKM + shRNA targeting mTOR for 20 days (OmT2_1) or 14 days (OmT2_2), IMR90 + OSKM + different shRNA targeting mTOR for 20 days (OmT3_1) or 14 days (OmT3_2), IMR90 + control vector for 20 days (EV_1), IMR90 + control vector for 14 days (EV_2), IMR90 + OSKM for 20 days (OSKM_1), IMR90 + OSKM for 14 days (OSKM_2)