Comprehensive Cell Surface Protein Profiling Identifies Specific Markers of Human Naive and Primed Pluripotent States

Abstract

SummaryHuman pluripotent stem cells (PSCs) exist in naive and primed states and provide important models to investigate the earliest stages of human development. Naive cells can be obtained through primed-to-naive resetting, but there are no reliable methods to prospectively isolate unmodified naive cells during this process. Here we report comprehensive profiling of cell surface proteins by flow cytometry in naive and primed human PSCs. Several naive-specific, but not primed-specific, proteins were also expressed by pluripotent cells in the human preimplantation embryo. The upregulation of naive-specific cell surface proteins during primed-to-naive resetting enabled the isolation and characterization of live naive cells and intermediate cell populations. This analysis revealed distinct transcriptional and X chromosome inactivation changes associated with the early and late stages of naive cell formation. Thus, identification of state-specific proteins provides a robust set of molecular markers to define the human PSC state and allows new insights into the molecular events leading to naive cell resetting.

Single-cell transcriptome analysis of human, marmoset and mouse embryos reveals common and divergent features of preimplantation development

“…S5F). We also examined families associated with naïve versus primed pluripotency in cultured human pluripotent stem cells (PSC) (Theunissen et al, 2016;Collier et al, 2017), and found robust expression of SINE/VNTR/Alu (SVA) elements, specifically SVA_B, SVA_C, SVA_D, SVA_E, SVA_F, as well as HERVH-int and LTR5/7 in human EPI (Fig. S5G).…”

New Advances in Human X Chromosome Status from a Developmental and Stem Cell Biology

“…During this time, changes in XACT expression within the ICM lead to loss of XACT:XIST competition at one X chromosome, which is subsequently inactivated. In naïve hESCs, extended passaging drives the complete regression into the naïve state (62), which exhibits increased biallelic expression from both chromosomes in a manner corresponding to the pre-implantation ICM (7, 12, 62), in which the cells with the most biallelic sites on the X are those that also have the highest expression of XACT (7). At the very least, there seems to be a temporal link between XACT, the naïve-to-primed transition, and X chromosome inactivation.…”

Chemical Reversion of Conventional Human Pluripotent Stem Cells to a Naïve-like State with Improved Multilineage Differentiation Potency

“…Although these markers do not discriminate between conventional and LIF-3i states, their levels inversely correlate with the frequency of spontaneous differentiation that may occur in hPSC when transitioning from conventional hPSC to LIF-3i conditions. Additional surface antigens that may more specifically mark human naïve-like states in vitro 2,12,13 can also be employed to detect effective LIF-3i hPSC reversion.…”