Transient Naive Reprogramming Corrects Human iPS Cells Functionally and Epigenetically

Cells undergo significant epigenome reconfiguration during reprogram- ming to human induced pluripotent stem cells (hiPS cells). However, notable differences exist between the epigenomes of hiPS cells and human embryonic stem (hES) cells, impacting hiP memory of the originating cell population and aberrations that arise during reprogramming itself. The mechanisms and dynamics under- pinning these processes are not fully understood. We characterized
the persistence and emergence of these differences by performing genome-wide DNA methylation profiling throughout primed and naive reprogramming of human somatic cells to hiPS cells. We find that repro- gramming-induced epigenetic aberrations manifest midway through primed reprogramming, while DNA demethylation initiates early in naive reprogramming. Utilizing this knowledge, we devised a transient-na- ive-treatment (TNT) reprogramming strategy to emulate the epigenetic reset that occurs during early human embryogenesis. We show that the epigenetic memory in hiPS cells is concentrated in cell of origin-depen- dent repressive chromatin marked by H3K9me3, lamin-B1 and aberrant CpH methylation. TNT reprogramming effectively reconfigures such domains to a hES-cell-like state but does not disrupt genomic imprinting, thereby avoiding problems associated with long-term naive culturing. Using an isogenic system, we demonstrate that TNT reprogramming
can correct transposable element overexpression and differential gene expression seen in conventional hiPS cells, and that TNT-reprogrammed hiPS and hES cells show similar differentiation efficiencies. Moreover, TNT reprogramming enhances the differentiation of hiPS cells derived from multiple cell types and different germ layers. Thus, TNT repro- gramming corrects epigenetic memory and aberrations, producing hiPS cells that are molecularly and functionally more similar to hES cells than conventional hiPS cells. We foresee TNT reprogramming becoming a new standard for biomedical and therapeutic applications and providing a novel system for studying epigenetic memory.S cell function and rendering them not equally capable. Differences include epigenetic