The totipotent 2CLCs have also been reported to exhibit increased histone modifications in H3K27ac, H3K4me1, and H3K4me3, as compared with ESCs
[38]. Although these histone modifications are associated with transcriptional activation, no evidence exists to show that Dux expression will be directly regulated by these modifications. The downregulations of chromatin modifiers such as LSD1 and chromatin assembly factor 1 (CAF-1) facilitate MERVL activation
[15][39]. Furthermore, MERVL requires lysine (K)-specific demethylase 1A (KDM1A, also as LSD1)—a histone lysine-specific demethylase, a KRAP-associated transcriptional repressor (KAP1), and G9A—a H3K9 histone methyltransferase—for epigenetic repression in normal mESCs
[27][40][41]. Likewise, there is no direct evidence showing that the expression of Dux can be regulated by these chromatin modifiers. Recently, LIN28, an RNA-binding protein, was identified as able to repress Dux by an epigenetic program (
Figure 2A). H3K9me3 levels were decreased at Dux and its downstream targets, and thus de-repressed Dux expression in Lin28 knockout cells
[11]. However, the mechanisms underlying how Lin28 regulates H3K9me3 remain elusive. It is worth noting that Lin28a depletion releases Dux repression by reducing the occupancy of Nucleolin/tripartite motif-containing protein 28 (NCL/TRIM28) in the Dux region. TRIM28 is also known as KAP1, which was demonstrated to repress Dux expression in a long interspersed nuclear element-1 (LINE1)-dependent manner in mESCs
[27][42]. LINE1 are Class I transposable elements, which can repress Dux expression by interacting with NCL and KAP1 in mESCs
[42]. Mechanistically, after LINE1 RNA is methylated by METTLE3, the m6A-modified LINE1 RNA then works as a scaffold recognized by the YTH domain containing 1 (YTHDC1), which further recruits H3K9me3 regulators, including SET domain bifurcated histone lysine methyltransferase 1 (SETDB1) and KAP1, to the locus of Dux, inhibiting its expression
[43][44]. Furthermore, polycomb-repressive complexes (PRCs) bind LINE1 RNA and act as an essential partner for
Dux gene repression
[45]. SUMO modification enhances the H3K9me3 levels on a genome-wide scale, including the Dux locus, and facilitates the recruitment of PRC1.6 and KAP/SETDB1 complexes to the locus to repress
Dux gene expression (
Figure 2A)
[46]. In fact, Ythdc1 depletion results in a global decrease in the SETDB1-mediated H3K9me3 enrichment, which is accompanied by the re-activation of MERVL and Dux
[43]. However, Ythdc1-depleted cells still retain the ability to re-activate many retrotransposons upon Dux removal, indicating a parallel regulation pattern between Ythdc1 and Dux with regard to retrotransposon regulation
[44]. Due to the sequence differences in LINE1 among species, it is not clear whether LINE1 has similar effects in other mammals, including
Homo sapiens [47].