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Determining a Fundamental Role for APC in hESC Self-renewal

Review of “APC as a major regulator of hESCs self‐renewal” from STEM CELLS by Stuart P. Atkinson

Adenomatous polyposis coli (APC), a negative regulator of Wnt/β‐catenin signaling [1], is implicated in sporadic colorectal cancers and familial adenomatous polyposis (FAP), an autosomal dominant inherited syndrome. Researchers led by Dalit Ben-Yosef (Tel Aviv Sourasky Medical Center, Tel Aviv, Israel) previously derived a human embryonic stem cell (hESC) line from blastocyst embryos following preimplantation genetic diagnosis for FAP that carries a germline mutation in the APC gene [2, 3]. As studies into the role of Wnt/β‐catenin signaling in hESCs have generated conflicting results, the authors aimed to directly target the wild type APC allele in APC mutation-carrying hESCs (FAP-hESCs) to specifically investigate any role for APC in pluripotency and differentiation.

In their new STEM CELLS article [4], Preisler et al. now provide direct evidence for the strict requirement for constant β‐catenin degradation via APC activity to maintain pluripotency, thereby suggesting a fundamental role for APC in hESC self‐renewal.

FAP‐hESCs in culture existed as a homogenous population of cells lacking an elevated rate of spontaneous differentiation, suggesting that monoallelic APC expression maintains hESC self‐renewal. The authors employed CRISPR/Cas9-gene-editing technology to generate “second-hit” APC heterozygous frameshift mutations in FAP‐hESCs in two different loci; however, despite high CRISPR/Cas9 targeting efficiency and the successful isolation of clones, the authors failed to isolate any clone that carried a loss of function mutation in the wild type APC allele, in a similar result to a previous report [5]. The authors detected the expression of full-length APC protein in generated clones, suggesting that mutations occurred in the germline‐mutant allele rather than the wild type allele.

The team then turned to the application of fluorescent β‐catenin reporters to track APC double-mutant cells immediately following CRISPR/Cas9 mutagenesis. This approach established that APC double-mutant hESCs display dramatically elevated levels of β‐catenin and the activation of Wnt/β‐catenin signaling to induce rapid differentiation to endodermal and mesodermal lineages, thus revealing a strict requirement for constant β‐catenin degradation through the APC destruction complex to maintain pluripotency.

Overall, this compelling new study underscores the vital importance of APC and hence β‐catenin destruction and basal Wnt/β‐catenin signaling to the maintenance of hESC self-renewal.

For more on the fundamental factors controlling the self-renewal of human embryonic stem cells, stay tuned to the Stem Cells Portal!

 

References 

  1. Barker N, The canonical Wnt/β-catenin signalling pathway, in Wnt Signaling. 2008, Springer. p. 5-15.
  2. Ben-Yosef D, Amit A, Malcov M, et al., Female sex bias in human embryonic stem cell lines. Stem cells and development 2011;21:363-372.
  3. Yedid N, Kalma Y, Malcov M, et al., The effect of a germline mutation in the APC gene on β-catenin in human embryonic stem cells. BMC cancer 2016;16:952.
  4. Preisler L, Ben-Yosef D, and Mayshar Y, Adenomatous Polyposis Coli as a Major Regulator of Human Embryonic Stem Cells Self-Renewal. STEM CELLS 2019;37:1505-1515.
  5. Sommer CA, Capilla A, Molina-Estevez FJ, et al., Modeling APC mutagenesis and familial adenomatous polyposis using human iPS cells. PLOS ONE 2018;13:e0200657.