Base editing in bovine embryos reveals a species-specific role of SOX2 in regulation of pluripotency

PLoS Genet. 2022 Jul 5;18(7):e1010307. doi: 10.1371/journal.pgen.1010307. eCollection 2022 Jul.

Abstract

The emergence of the first three lineages during development is orchestrated by a network of transcription factors, which are best characterized in mice. However, the role and regulation of these factors are not completely conserved in other mammals, including human and cattle. Here, we establish a gene inactivation system with a robust efficiency by introducing premature codon with cytosine base editors in bovine early embryos. By using this approach, we have determined the functional consequences of three critical lineage-specific genes (SOX2, OCT4 and CDX2) in bovine embryos. In particular, SOX2 knockout results in a failure of the establishment of pluripotency in blastocysts. Indeed, OCT4 level is significantly reduced and NANOG barely detectable. Furthermore, the formation of primitive endoderm is compromised with few SOX17 positive cells. RNA-seq analysis of single blastocysts (day 7.5) reveals dysregulation of 2074 genes, among which 90% are up-regulated in SOX2-null blastocysts. Intriguingly, more than a dozen lineage-specific genes, including OCT4 and NANOG, are down-regulated. Moreover, SOX2 level is sustained in the trophectoderm in absence of CDX2. However, OCT4 knockout does not affect the expression of SOX2. Overall, we propose that SOX2 is indispensable for OCT4 and NANOG expression and CDX2 represses the expression of SOX2 in the trophectoderm in cattle, which are all in sharp contrast with results in mice.

MeSH terms

  • Animals
  • Blastocyst / metabolism
  • Cattle
  • Endoderm / metabolism
  • Gene Expression Regulation, Developmental*
  • Homeodomain Proteins / genetics
  • Humans
  • Mammals / genetics
  • Mice
  • Nanog Homeobox Protein / genetics
  • Nanog Homeobox Protein / metabolism
  • Octamer Transcription Factor-3* / genetics
  • Octamer Transcription Factor-3* / metabolism
  • SOXB1 Transcription Factors / genetics
  • SOXB1 Transcription Factors / metabolism
  • Species Specificity
  • Transcription Factors / genetics

Substances

  • Homeodomain Proteins
  • Nanog Homeobox Protein
  • Octamer Transcription Factor-3
  • SOX2 protein, human
  • SOXB1 Transcription Factors
  • Sox2 protein, mouse
  • Transcription Factors

Grant support

KZ is supported by the National Natural Science Foundation of China (No. 31872348, No. 32072731 and No. 32161143032)(https://www.nsfc.gov.cn/). HW is funded by the National Natural Science Foundation of China (No. 32072939)(https://www.nsfc.gov.cn/). LL and SW are both supported by the National Natural Science Foundation of China (No.31941007)(https://www.nsfc.gov.cn/). KZ is also funded by Zhejiang Provincial Natural Science Foundation (LZ21C170001) (http://zjnsf.kjt.zj.gov.cn/portal/index.html). HW is also supported by Zhejiang Provincial Natural Science Foundation (No. LY19C180002)(http://zjnsf.kjt.zj.gov.cn/portal/index.html). LL is funded by China Postdoctoral Science Foundation (No. 2020M671742)(http://jj.chinapostdoctor.org.cn/website/index.html). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.