LINE-1 (L1) retrotransposons are the only active and autonomous transposable element in the human genome. We recently analyzed the DNA methylation levels of their promoter region at individual loci, in a varied panel of human cell lines. We found that embryonic cells exhibit a unique profile with the youngest primate-specific L1 families being hypomethylated but older ones being hypermethylated. Notably, the transition seems to occur within the L1PA3 family, which possesses two versions of the promoter. In an older subgroup, the L1 promoter contains a binding site for ZNF93, a Krüppel-associated box (KRAB) domain-containing zinc-finger protein, which can recruit Kap1/TRIM28, leading to the formation of repressive chromatin. By contrast, a younger L1PA3 subgroup shows a deletion encompassing the ZNF93 binding site. We confirmed that these two L1PA3 subgroups recapitulate the differential methylation observed between young vs old primate-specific L1 elements in embryonic cells, the loss of methylation being associated with the deletion of the ZNF93 binding site. To test whether ZNF93 plays a direct role in L1 promoter methylation, we generated ZNF93 knock-out clones and assessed DNA methylation of each individual primate-specific L1 promoter using our recently developed strategy, bs-ATLAS-seq. We did not observe significant alterations of L1 DNA methylation profiles, indicating that ZNF93 is not a necessary factor to maintain DNA methylation of L1 promoters from L1PA3 and older primate-specific families in these cells. However, it is still possible that ZNF93 is involved in the establishment of their methylation profiles.

AJD, CP and SL contributed equally. This work was supported by Agence Nationale de la Recherche (ANR-11-LABX-0028; ANR-11-LABX-0071; ANR-15-IDEX-0001; ANR-16-CE12-0020; ANR-18-IDEX-0001; ANR-19-CE12-0032; ANR-21-CE12-0001), Fondation pour la Recherche Médicale (DEQ20180339170), Institut National Du Cancer (INCa PLBIO 2020-095), Fondation ARC (PGA1/RF20180206807), grants from the Canceropôle PACA, INCa, Region Sud (Projet Emergence), INSERM (GOLD Cross-cutting Program on Genomic Variability), CNRS (GDR 3546).