CDCA7 targets LSH to DNA maintenance methylation in S phase and transcription regulation in interphase via two distinct DNA-binding modes.

Mutations or pathogenic variants in CDCA7 and lymphoid-specific helicase (LSH) lead to immunodeficiency, centromeric instability, and facial anomalies (ICF) syndrome, and CDCA7 has been recognized as a hemimethylated DNA sensor facilitating DNA methylation. Here, via genome profiling experiments we unravel that CDCA7 is actually highly enriched at the poorly methylated, CG-rich promoter regions and that an ICF syndrome-causing G294V mutation abolishes this activity. In vitro CDCA7 but not the G294V mutant can bind both CG-rich DNA probes and hemimethylated DNA. Consistent with these dual DNA-binding activities, CDCA7 exhibits two distinct subcellular localization patterns along the cell cycle: a diffuse nuclear distribution in interphase and a hemimethylated DNA-dependent pericentromeric heterochromatin domain localization in late S phase. Notably, LSH is recruited by CDCA7 to both the CG-rich promoter regions in interphase and heterochromatin domains in late S phase. Whole genome DNA methylation analysis confirmed a crucial and conserved role of CDCA7 and LSH in DNA maintenance methylation. Transcription analyses revealed that CDCA7 and LSH can interdependently regulate transcription independent of DNA methylation. Altogether, our study reveals that CDCA7 and LSH have a role beyond DNA methylation and can regulate gene expression via binding CG-rich DNA motifs, thus providing new insights into the function of CDCA7/LSH and the complexity of ICF syndrome.