The locus harbors eGenes in the HLC cohort (CPNE1), the iPSC cohort (CPNE1), and the GTEx liver cohort (ERGIC3); CPNE1 exhibited ASE in HLCs and iPSCs (lowest CPNE1 HLC P = 3 × 10−8; lowest CPNE1 iPSC P = 6 × 10−9). The functions of these genes are poorly understood. We reasoned that rs2277862 might modulate transcription of CPNE1 in both HLCs and undifferentiated human pluripotent stem cells (hPSCs). To test if rs2277862 regulates CPNE1 and/or ERGIC3 expression, we performed four types of experiments, two based in hPSCs. First, in an hPSC line, HUES 8 (C/C, major/major at rs2277862), we used CRISPR-Cas9 to knock in one minor allele (Figure 3B). Using a single-strand DNA oligonucleotide as a repair template, we obtained only a single recombinant heterozygous (C/T) clone at a frequency of 0.15% (1 out of 672 clones screened), highlighting the inefficiency of the procedure. We observed significantly decreased CPNE1 expression in both undifferentiated knock-in hPSCs (down 19%) and differentiated knock-in HLCs (down 10%), with non-significant effects on ERGIC3 (Figure 3C).
