Highlights d Exquisitely specific maintenance methylase enzyme drives all 5mC in C. neoformans d Once lost, methylation is not efficiently restored mitotically or meiotically d The de novo enzyme DnmtX was lost in an ancestral species 50-150 mya d Persistence of 5mC for millions of years explained by Darwinian epigenome evolution

After the initial establishment of symmetric cytosine methylation patterns by de novo DNA methyltransferases (DNMTs), maintenance DNMTs mediate epigenetic memory by propagating the initial signal. We find that CG methylation in the yeast Cryptococcus neoformans is dependent on a purely epigenetic mechanism mediated by the single DNMT encoded by the genome, Dnmt5. Purified Dnmt5 is a maintenance methyltransferase that strictly requires a hemimethylated substrate, and methylation lost by removal of Dnmt5 in vivo is not restored by its mitotic or meiotic reintroduction. Phylogenetic analysis reveals that the ancestral species had a second methyltransferase, DnmtX, whose gene was lost between 50 and 150 Mya.Expression of extant DnmtXs in C. neoformans triggers de novo methylation. These data indicate that DNA methylation has been maintained epigenetically by the Dnmt5 system since the ancient loss of the DnmtX de novo enzyme, implying remarkably long-lived epigenetic memory. Single sentence summary: Epigenetic information can be inherited over geological timescalesAll rights reserved. No reuse allowed without permission.

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Embryonic development is dictated by tight regulation of DNA replication, cell division and differentiation. Mutations in DNA repair and replication genes disrupt this equilibrium, giving rise to neurodevelopmental disease characterized by microcephaly, short stature and chromosomal breakage. Here, we identify biallelic variants in two components of the RAD18-SLF1/2-SMC5/6 genome stability pathway, SLF2 and SMC5, in 11 patients with microcephaly, short stature, cardiac abnormalities and anemia. Patient-derived cells exhibit a unique chromosomal instability phenotype consisting of segmented and dicentric chromosomes with mosaic variegated hyperploidy. To signify the importance of these segmented chromosomes, we have named this disorder Atelís (meaning - incomplete) Syndrome. Analysis of Atelís Syndrome cells reveals elevated levels of replication stress, partly due to a reduced ability to replicate through G-quadruplex DNA structures, and also loss of sister chromatid cohesion. Together, these data strengthen the functional link between SLF2 and the SMC5/6 complex, highlighting a distinct role for this pathway in maintaining genome stability.

Despite being the first homolog of the bacterial RecQ helicase to be identified in humans the function of RECQL1 remains poorly characterised. Furthermore, unlike other members of the human RECQ family of helicases, mutations in RECQL1 have not been associated with a genetic disease. Here we identify two families with a novel genome instability disorder, named RECON (RECql ONe) Syndrome caused by biallelic mutations in the RECQL gene. The affected individuals exhibit short stature, progeroid facial features, a hypoplastic nose, xeroderma and skin photosensitivity. Affected individuals were homozygous for the same missense mutation in RECQL1 (p.Ala459Ser) located within its zinc binding domain. Biochemical analysis of the mutant RECQL1 protein revealed that the p.A459S missense mutation compromised its ATPase, helicase and fork restoration activity, whilst its capacity to promote single-strand DNA annealing was largely unaffected. At the cellular level, this mutation in RECQL1 gave rise to a defect in the ability to repair DNA damage induced by exposure to topoisomerase poisons and a failure of DNA replication to progress efficiently in the presence of abortive topoisomerase lesions. Taken together, RECQL1 is the fourth member of the RecQ family of helicases to be associated with a human genome instability disorder.