Evolutionary-conserved telomere-linked helicase genes of fission yeast are repressed by silencing factors, RNAi components and the telomere-binding protein Taz1

In Schizosaccharomyces pombe the RNAi machinery and proteins mediating heterochromatin formation regulate the transcription of non-coding centromeric repeats. These repeats share a high sequence similarity with telomere-linked helicase (tlh) genes, implying an ancestral relationship between the two types of elements and suggesting that transcription of the tlh genes might be regulated by the same factors as centromeric repeats. Indeed, we found that mutants lacking the histone methyltransferase Clr4, the Pcu4 cullin, Clr7 or Clr8, accumulate high levels of tlh forward and reverse transcripts. Mutations and conditions perturbing histone acetylation had similar effects further demonstrating that the tlh genes are normally repressed by heterochromatin. In contrast, mutations in the RNAi factors Dcr1, Ago1 or Rdp1 led only to a modest derepression of the tlh genes indicating an alternate pathway recruits heterochromatin components to telomeres. The telomere-binding protein Taz1 might be part of such a redundant pathway, tlh transcripts being present at low levels in Δtaz1 mutants and at higher levels in Δtaz1 Δdcr1 double mutants. Surprisingly, the chromodomain protein Chp1, a component of the Ago1-containing RITS complex, contributes more to tlh repression than Ago1, indicating the repressive effects of Chp1 are partially independent of RITS. The tlh genes are found in the subtelomeric regions of several other fungi raising the intriguing possibility of conserved regulation and function

H3K9me-Independent Gene Silencing in Fission Yeast Heterochromatin by Clr5 and Histone Deacetylases

Nucleosomes in heterochromatic regions bear histone modifications that distinguish them from euchromatic nucleosomes. Among those, histone H3 lysine 9 methylation (H3K9me) and hypoacetylation have been evolutionarily conserved and are found in both multicellular eukaryotes and single-cell model organisms such as fission yeast. In spite of numerous studies, the relative contributions of the various heterochromatic histone marks to the properties of heterochromatin remain largely undefined. Here, we report that silencing of the fission yeast mating-type cassettes, which are located in a well-characterized heterochromatic region, is hardly affected in cells lacking the H3K9 methyltransferase Clr4. We document the existence of a pathway parallel to H3K9me ensuring gene repression in the absence of Clr4 and identify a silencing factor central to this pathway, Clr5. We find that Clr5 controls gene expression at multiple chromosomal locations in addition to affecting the mating-type region. The histone deacetylase Clr6 acts in the same pathway as Clr5, at least for its effects in the mating-type region, and on a subset of other targets, notably a region recently found to be prone to neo-centromere formation. The genomic targets of Clr5 also include Ste11, a master regulator of sexual differentiation. Hence Clr5, like the multi-functional Atf1 transcription factor which also modulates chromatin structure in the mating-type region, controls sexual differentiation and genome integrity at several levels. Globally, our results point to histone deacetylases as prominent repressors of gene expression in fission yeast heterochromatin. These deacetylases can act in concert with, or independently of, the widely studied H3K9me mark to influence gene silencing at heterochromatic loci

The fork protection complex promotes parental histone recycling and epigenetic memory

The inheritance of parental histones across the replication fork is thought to mediate epigenetic memory. Here, we reveal that fission yeast Mrc1 (CLASPIN in humans) binds H3-H4 tetramers and operates as a central coordinator of symmetric parental histone inheritance. Mrc1 mutants in a key connector domain disrupted segregation of parental histones to the lagging strand comparable to Mcm2 histone-binding mutants. Both mutants showed clonal and asymmetric loss of H3K9me-mediated gene silencing. AlphaFold predicted co-chaperoning of H3-H4 tetramers by Mrc1 and Mcm2, with the Mrc1 connector domain bridging histone and Mcm2 binding. Biochemical and functional analysis validated this model and revealed a duality in Mrc1 function: disabling histone binding in the connector domain disrupted lagging-strand recycling while another histone-binding mutation impaired leading strand recycling. We propose that Mrc1 toggles histones between the lagging and leading strand recycling pathways, in part by intra-replisome co-chaperoning, to ensure epigenetic transmission to both daughter cells

Expression-state boundaries in the mating-type region of fission yeast.

A transcriptionally silent chromosomal domain is found in the mating-type region of fission yeast. Here we show that this domain is delimited by 2-kb inverted repeats, IR-L and IR-R. IR-L and IR-R prevent the expansion of transcription-permissive chromatin into the silenced region and that of silenced chromatin into the expressed region. Their insulator activity is partially orientation dependent. The silencing defects that follow deletion or inversion of IR-R are suppressed by high dosage of the chromodomain protein Swi6. Combining chromosomal deletions and Swi6 overexpression shows that IR-L and IR-R provide firm borders in a region where competition between silencing and transcriptional competence occurs. IR-R possesses autonomously replicating sequence (ARS) activity, leading to a model where replication factors, or replication itself, participate in boundary formation