Figure 2.

Deficiency of LIN28 causes activation of ERV and 2C marker genes. (A) Scatter plot of RNA-sequencing data comparing gene expression of wild-type and Lin28a knockout ES cells. (B) Embryo stage specific genes were classified into five clusters based on their expression pattern. (C) Log₂(fold change) of gene expression abundance between Lin28a knockout ES cells and wild-type cells for each gene cluster. ***P < 0.001, Mann-Whitney U test. (D) Different classes of Long Terminal Repeat or LTR expression in Lin28a and Lin28b knockout ES cells. (E) Two sub-classes of ERVL genes MERVL-int and MT2_Mm are significantly up-regulated in Lin28a knockout ES cells. For (D and E), the TPM values of LTR expression were linearly normalized to a range from 0 to 1 across all samples. ***P < 0.001, Wilcox signed rank test. (F and G) qPCR validation of selected 2C marker genes and Mervl-polymerase gene (Mervl-pol) in wild-type and Lin28a/b knockout ES cells. (H and I) LIN28A overexpression in knockout cells reversed the activation of 2C genes and ERV genes. (J) Microscopy pictures and flow cytometry showing the percentage of 2C-like cells using ES cells transduced with a 2C::tdTomato reporter construct. (K) Proliferation curve of wild-type and Lin28a knockout ES cells. *P < 0.05, **P < 0.01, ***P < 0.001, two-way ANOVA, n = 3, error bar: standard error of the mean