The SNF2-Family Member Fun30 Promotes Gene Silencing in Heterochromatic Loci

Abstract

Chromatin regulates many key processes in the nucleus by controlling access to the underlying DNA. SNF2-like factors are ATP-driven enzymes that play key roles in the dynamics of chromatin by remodelling nucleosomes and other nucleoprotein complexes. Even simple eukaryotes such as yeast contain members of several subfamilies of SNF2-like factors. The FUN30/ETL1 subfamily of SNF2 remodellers is conserved from yeasts to humans, but is poorly characterized. We show that the deletion of FUN30 leads to sensitivity to the topoisomerase I poison camptothecin and to severe cell cycle progression defects when the Orc5 subunit is mutated. We demonstrate a role of FUN30 in promoting silencing in the heterochromatin-like mating type locus HMR, telomeres and the rDNA repeats. Chromatin immunoprecipitation experiments demonstrate that Fun30 binds at the boundary element of the silent HMR and within the silent HMR. Mapping of nucleosomes in vivo using micrococcal nuclease demonstrates that deletion of FUN30 leads to changes of the chromatin structure at the boundary element. A point mutation in the ATP-binding site abrogates the silencing function of Fun30 as well as its toxicity upon overexpression, indicating that the ATPase activity is essential for these roles of Fun30. We identify by amino acid sequence analysis a putative CUE motif as a feature of FUN30/ETL1 factors and show that this motif assists Fun30 activity. Our work suggests that Fun30 is directly involved in silencing by regulating the chromatin structure within or around silent loci.

Mediator, TATA-binding Protein, and RNA Polymerase II Contribute to Low Histone Occupancy at Active Gene Promoters in Yeast

“…Little or no change was seen in steady-state-induced expression levels of CHA1 in yeast strains lacking core members of the ISW and INO80 complexes (30,31), or Fun30, which has homology to Snf2 and functions in gene silencing in heterochromatin and DNA repair (Fig. 1B) (32)(33)(34). We also tested the effect on CHA1 induction in the absence of the histone chaperone Asf1, which contributes to chromatin remodeling and transcriptional activation of the paradigmatic PHO5 gene in yeast, particularly under conditions of partial phosphate depletion (35,36), and again found no significant effect (Fig.…”

Roles of Chromatin Remodeling Factors in the Formation and Maintenance of Heterochromatin Structure

“…For example, HMR silencing is more dependent on the nearby telomere than HML silencing, whereas HML silencing is preferentially affected by certain histone mutations (30,31). Although FUN30 and ISW1 are required for HMR silencing, it has not been examined whether they are also required for HML silencing (20,22). To address this question, we deleted FUN30 and ISW1 individually from strain 1 bearing a URA3 reporter in the middle of HML ( Fig.…”

“…Isw1p is required for HMR but not telomeric silencing, whereas Snf2p is required for telomeric but not HM silencing (20,21). Fun30p, on the other hand, is required for silencing at both HMR and telomeric loci and is associated with heterochromatin (22). Whereas the chromatin remodeling activities of Isw1p and Snf2p have been long established, Fun30p has only recently been shown to possess histone H2A/H2B dimer exchange and nucleosome sliding activities (23).…”

Keeping chromatin quiet

“…9 We propose therefore that SMARCAD1 mediates maintenance of heterochromatin structures. [31][32][33] The fact that interfering with SMARCAD1 function perturbs heterochromatin organization so profoundly is in line with the recognized prerequisite for histone deacetylation in establishing silent chromatin structures and maintaining chromosome stability from yeast to man. 22,[34][35][36] Interestingly, the global changes in histone modification patterns observed in SMARCAD1 knockdown cells coincide with S-phase progression.…”

“…74 SMARCAD1 and its homologs in other species contain potential mono-ubiquitin binding motifs, termed CUE domains, raising the possibility that SMARCAD1 could target ubiquitinylated proteins like PCNA or H2A at replication sites. 32,75,76 Outlook It is likely that SMARCAD1 directs histone deacetylation at replication sites through its physical association with HDAC1 and HDAC2. Coupling HDACs with ATP-dependent chromatin remodeling activities within a multi-enzyme complex is a strategy also employed by Mi-2/ NURD, NCoR1, NoRC and the yeast SHREC complexes involved in transcriptional repression.…”