Oral Presentation 44th Lorne Genome Conference 2023

Epigenetic memory and differentiation bias in human iPS cells can be removed by a novel transient naive reprogramming strategy (#53)

Daniel Poppe 1 2 3 , Sam Buckberry 1 2 , Xiaodong Lu 4 5 6 , Jia P Tan 4 5 6 , Guizhi Sun 4 5 6 , Joseph Chen 4 5 6 , Trung V Nguyen 1 2 , Alex de Mendoza 1 2 7 , Jahnvi Pflueger 1 2 , Nathan Smits 8 9 , Thomas Frazer 1 2 , Dulce B Vargas-Landin 1 2 , Jacob M Paynter 4 5 6 , Ning Liu 10 , John F Ouyang 11 , Fernando J Rossello 4 5 6 , Hun S Chy 6 12 , Owen JL Rackham 11 , Andrew L Laslett 6 12 , James Breen 10 , Geoffrey J Faulkner 8 9 , Christian M Nefzger 4 5 6 , Jose M Polo 4 5 6 13 14 , Ryan Lister 1 2
  1. Australian Research Council Centre of Excellence in Plant Energy Biology, University of Western Australia, Perth, Western Australia, Australia
  2. Harry Perkins Institute of Medical Research, Perth, Western Australia, Australia
  3. University of Western Australia, Nedlands, WA, Australia
  4. Department of Anatomy and Developmental Biology, Monash University, Melbourne, Victoria, Australia
  5. Development and Stem Cells Program, Monash Biomedicine Discovery Institute, Melbourne, Victoria, Australia
  6. Australian Regenerative Medicine Institute, Monash University, Melbourne, Victoria, Australia
  7. School of Biological and Chemical Sciences, Queen Mary University of London, London, United Kingdom
  8. Mater Research Institute, University of Queensland, Woolloongabba, Queensland, Australia
  9. Queensland Brain Institute, University of Queensland, Brisbane, Queensland, Australia
  10. South Australian Health & Medical Research Institute, Adelaide, South Australia, Australia
  11. Program in Cardiovascular and Metabolic Disorders, Duke-National University of Singapore Medical School, Singapore
  12. Biomedical Manufacturing, Commonwealth Scientific and Industrial Research Organisation Manufacturing, Melbourne, Victoria, Australia
  13. Adelaide Centre for Epigenetics and the South Australian Immunogenomics Cancer Institute, The University of Adelaide, Adelaide, South Australia, Australia
  14. Faculty of Medicine, Nursing and Health Sciences, Monash University, Melbourne, Victoria, Australia

The reprogramming of somatic cells to human induced pluripotent stem cells (hiPSCs) involves major reconfiguration of the epigenome. However, numerous independent studies have found that the epigenomes of hiPSCs and human embryonic stem cells (hESCs) show significant differences, which impact the function and differentiation potential of hiPSCs. These differences include epigenetic memory of the originating cell population as well as epigenetic alterations introduced during reprogramming itself. The mechanisms and dynamics underpinning these processes are still unknown. Here, we used genome-wide temporal DNA methylation profiling throughout human primed and naive reprogramming to characterise the persistence and emergence of such epigenetic differences in systems that resemble different stages of pluripotency and epigenome establishment during embryonic development. Reprogramming-induced epigenetic aberrations emerge midway through primed reprogramming, while somatic cell memory erasure begins early during naive reprogramming. This knowledge was employed to develop a transient-naive-treatment reprogramming strategy that mimics the epigenetic reset during early human embryonic development. Epigenetic memory in conventional primed reprogrammed hiPSCs is concentrated in cell-of-origin large, lamina-associated repressive domains marked by H3K9me3 and aberrant CH methylation of DNA. Transient-naive-treatment reprogramming reconfigures these domains to an hESC-like state while maintaining genomic imprinting similar to early development, thus avoiding the imprinting erasure induced by long-term naive culturing. Using genetically matched hESCs and hiPSCs, a strong lineage–of–origin differentiation bias was found together with increased transposable element expression in conventional primed hiPSCs, which is eliminated with transient-naive-treatment reprogramming. Thus, transient-naive-treatment reprogramming effectively eliminates epigenetic memory and aberrations, resulting in hiPSCs that are molecularly and functionally more similar to hESCs than conventional primed hiPSCs. We anticipate this reprogramming system will become a new standard for biomedical research and therapeutic applications and provides a new model system for studying the mechanisms of epigenetic memory.