Sitong Liu1, Rui Wei2, Hongyang Liu2, Ruiqi Liu3, Pengxiao Li3, Xiaoyu Zhang2, Wei Wei4, Xiaodong Zhao3, Xiaomeng Li4, Yang Yang5, Xueqi Fu1, Kang Zou2. 1. College of Life Sciences, Jilin University, Changchun, China. 2. Germline Stem Cells and Microenvironment Lab, College of Animal Science and Technology, Nanjing Agricultural University, Nanjing, China. 3. Key Laboratory of Systems Biomedicine (Ministry of Education), Shanghai Center for Systems Bio-medicine, Shanghai Jiao Tong University, Shanghai, China. 4. The Key Laboratory of Molecular Epigenetics of MOE, Institute of Genetics and Cytology, Northeast Normal University, Changchun, China. 5. State Key Laboratory of Reproductive Medicine, Nanjing Medical University, Nanjing, China.
Abstract
OBJECTIVES: Spermatogonial stem cells (SSCs), the germline stem cells (GSCs) committed to spermatogenesis in niche, can transform into pluripotent state in long-term culture without introduction of exogenous factors, typically in p53 deficiency condition. As the guardian for genomic stability, p53 is associated with epigenetic alterations during SSCs transformation. However, the mechanism is still unknown, since complicated roles of p53 baffle our understanding of the regulating process. MATERIALS AND METHODS: The chromatin accessibility and differentially expressed genes (DEGs) were analysed in p53+/+ and p53-/- SSCs using the Assay for Transposase-Accessible Chromatin with high-throughput Sequencing (ATAC-seq) and RNA-sequencing (RNA-seq), to explore the connection of p53 and cell fate at chromosomal level. RESULTS: Several transcription factors (TFs), such as CTCF, SMAD3 and SOX2, were predicted as important factors mediating the transformation. Molecular evidence suggested that SMAD3 efficiently promoted pluripotency-associated gene expression both in fresh and long-term cultured SSCs. However, p53 knockout (KO) is insufficient to induce SMAD3 expression in SSCs. CONCLUSIONS: These observations indicate that SMAD3 is a key factor for SSCs transformation, and an unknown event is required to activate SMAD3 as the prerequisite for SSCs reprogramming, which may occur in the long-term culture of SSCs. This study demonstrates the connection of p53 and pluripotency-associated factors, providing new insight for understanding the mechanisms of SSCs reprogramming and germline tumorigenesis.
OBJECTIVES: Spermatogonial stem cells (SSCs), the germline stem cells (GSCs) committed to spermatogenesis in niche, can transform into pluripotent state in long-term culture without introduction of exogenous factors, typically in p53 deficiency condition. As the guardian for genomic stability, p53 is associated with epigenetic alterations during SSCs transformation. However, the mechanism is still unknown, since complicated roles of p53 baffle our understanding of the regulating process. MATERIALS AND METHODS: The chromatin accessibility and differentially expressed genes (DEGs) were analysed in p53+/+ and p53-/- SSCs using the Assay for Transposase-Accessible Chromatin with high-throughput Sequencing (ATAC-seq) and RNA-sequencing (RNA-seq), to explore the connection of p53 and cell fate at chromosomal level. RESULTS: Several transcription factors (TFs), such as CTCF, SMAD3 and SOX2, were predicted as important factors mediating the transformation. Molecular evidence suggested that SMAD3 efficiently promoted pluripotency-associated gene expression both in fresh and long-term cultured SSCs. However, p53 knockout (KO) is insufficient to induce SMAD3 expression in SSCs. CONCLUSIONS: These observations indicate that SMAD3 is a key factor for SSCs transformation, and an unknown event is required to activate SMAD3 as the prerequisite for SSCs reprogramming, which may occur in the long-term culture of SSCs. This study demonstrates the connection of p53 and pluripotency-associated factors, providing new insight for understanding the mechanisms of SSCs reprogramming and germline tumorigenesis.
Authors: Sven Heinz; Christopher Benner; Nathanael Spann; Eric Bertolino; Yin C Lin; Peter Laslo; Jason X Cheng; Cornelis Murre; Harinder Singh; Christopher K Glass Journal: Mol Cell Date: 2010-05-28 Impact factor: 17.970