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Genome-scale Screening in a Rat Haploid System Identifies Thop1 as a Modulator of Pluripotency Exit II

ABSTRACT: Rat embryonic stem cells (ESCs), which are widely studied, can self-renew and exhibit pluripotency in long-term culture, but the mechanism underlying how they exit pluripotency remains obscure. Rat haploid ESCs (haESCs) enable advances in the discovery of unknown functional genes owing to their homozygous and pluripotent characteristics. Herein, we performed genome-wide mutation using piggyBac transposons in rat haESCs and obtained differentiation-retarded mutants assisted by Rex1-GFP reporter selection. High-throughput sequencing analysis further revealed numerous insertions related to various pathways affecting random differentiation. Thereafter, deletion of Thop1 (one candidate gene in the screened list) arrested the differentiation of rat ESCs by inhibiting the phosphorylation of ERK1/2, whereas overexpression of Thop1 promoted rat ESCs exit from pluripotency. Our findings provide an ideal tool to study functional genomics in rats: a homozygous haploid system carrying a pluripotency reporter that facilitates robust discovery of the mechanisms involved in the self-renewal or pluripotency of rat ESCs.

ORGANISM(S): Rattus norvegicus

PROVIDER: GSE190480 | GEO | 2021/12/15

REPOSITORIES: GEO

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Genome-scale Screening in a Rat Haploid System Identifies Thop1 as a Modulator of Pluripotency Exit

Project description:Rat embryonic stem cells (ESCs), which are widely studied, can self-renew and exhibit pluripotency in long-term culture, but the mechanism underlying how they exit pluripotency remains obscure. Rat haploid ESCs (haESCs) enable advances in the discovery of unknown functional genes owing to their homozygous and pluripotent characteristics. Herein, we performed genome-wide mutation using piggyBac transposons in rat haESCs and obtained differentiation-retarded mutants assisted by Rex1-GFP reporter selection. High-throughput sequencing analysis further revealed numerous insertions related to various pathways affecting random differentiation. Thereafter, deletion of Thop1 (one candidate gene in the screened list) arrested the differentiation of rat ESCs by inhibiting the phosphorylation of ERK1/2, whereas overexpression of Thop1 promoted rat ESCs exit from pluripotency. Our findings provide an ideal tool to study functional genomics in rats: a homozygous haploid system carrying a pluripotency reporter that facilitates robust discovery of the mechanisms involved in the self-renewal or pluripotency of rat ESCs.

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