Analysis of the Clustered Protocadherin (cPcdh) Locus in Human Pluripotent Stem and Derived Cells (RNA-seq I out of II)

In the mammalian genome, the clustered protocadherin (cPcdh) locus is a paradigm of stochastic gene expression with the potential to express a different cPcdh combination in every neuron. Here, we report a limited version established during the transition from the naive to the primed states of human cell pluripotency that reduces by orders of magnitude the combinatorial potential in the cPcdh locus. It is a chromatin-based mechanism that increases the frequency of stochastic selection of a subset of cPcdh promoters upon differentiation to fetal-like neurons in monolayers, organoids, or the rat spinal cord. Signs of similar preferential selections can be observed in the brain throughout fetal development, disappearing after birth, but not in a condition of delayed maturation such as Down Syndrome. We therefore propose that pluripotent cells impose a pattern of limited cPcdh diversity on neurons that increases the likelihood of cPcdh repetition until these cells acquire adult-like maturation. Lines/sublines: The hiPSC1-8 sublines were generated from single cells of the parent CVB hiPSC line. Six of these sublines were genetically modified (hiPSC2-7) and two were cases of failed genome editing (hiPSC1 and 8); please see Supplementary Table S1 in the associated publication for more details. Initially, these modifications should be irrelevant to the study of the cPcdh locus. The hESC 1.8 subline was generated from a single cell of the parent HUES9 hESC line and it was profiled in the primed state, naive state (conversion from the primed state, 5iLA protocol), and re-primed state (which are cells converted into the naive state and returned to the primed state). RNA-seq data: triplicate differentiation replicates of neurons generated from NPCs derived from hiPSC1-8 (n=24); cortical neurons generated from hiPSC1/3/5 (n=3); 10-month-old cortical organoids generated from the CVB line (n=1, pool of organoids); and primed, naive, and re-primed HUES9 1.8 subline (n=1 each).