Normal chromosome conformation depends on subtelomeric facultative heterochromatin in Neurospora crassa

Abstract

High-throughput chromosome conformation capture (Hi-C) analyses revealed that the 3D structure of the Neurospora crassa genome is dominated by intra- and interchromosomal links between regions of heterochromatin, especially constitutive heterochromatin. Elimination of trimethylation of lysine 9 on histone H3 (H3K9me3) or its binding partner Heterochromatin Protein 1 (HP1)—both prominent features of constitutive heterochromatin—have little effect on the Hi-C pattern. It remained possible that di- or trimethylation of lysine 27 on histone H3 (H3K27me2/3), which becomes localized in regions of constitutive heterochromatin when H3K9me3 or HP1 are lost, plays a critical role in the 3D structure of the genome. We found that H3K27me2/3, catalyzed by the Polycomb Repressive Complex 2 (PRC2) member SET-7 (SET domain protein-7), does indeed play a prominent role in the Hi-C pattern of WT, but that its presence in regions normally occupied by H3K9me3 is not responsible for maintenance of the genome architecture when H3K9me3 is lost. The Hi-C pattern of a mutant defective in the PRC2 member N. crassa p55 (NPF), which is predominantly required for subtelomeric H3K27me2/3, was equivalent to that of the set-7 deletion strain, suggesting that subtelomeric facultative heterochromatin is paramount for normal chromosome conformation. Both PRC2 mutants showed decreased heterochromatin–heterochromatin contacts and increased euchromatin–heterochromatin contacts. Cytological observations suggested elimination of H3K27me2/3 leads to partial displacement of telomere clusters from the nuclear periphery. Transcriptional profiling of Δdim-5, Δset-7, Δset-7; Δdim-5, and Δnpf strains detailed anticipated changes in gene expression but did not support the idea that global changes in genome architecture, per se, led to altered transcription.

Polycomb “polypacks” the chromatin

“…Furthermore, H3K27me2/3 cannot maintain constitutive heterochromatin silencing in the absence of H3K9me3 in N. crassa (12). Thus, as noted by Klocko et al (4), H3K27 methylation cannot functionally replace H3K9me3.…”

“…Thus, it is conceivable that decreased pericentromere interactions would also account for the abnormal centromere-euchromatin connections observed in the N. crassa strain that lacked H3K27 methylation. In this case, Klocko et al (4) indicate that subtelomeric H3K27me3 prevent such ectopic centromere-euchromatin interactions, which possibly occurred in an indirect manner. Alternatively, but not exclusively, PRC2 may have an H3K27me3-independent function at the centromere to restrain the spatial compartmentalization of centromeres from chromosome arms.…”

“…Thus, an additional question is whether facultative heterochromatin could substitute the function of constitutive heterochromatin in chromosome organization. The results from Klocko et al (4) show that the Hi-C pattern in a strain that lacks both H3K9 and H3K27 methylation highly resembles that of the Neurospora Δset-7 strain. Moreover, losing both H3K9 and H3K27 methylation does not change the dominance of constitutive heterochromatin interactions in chromosome organization, which suggests that there are alternative mechanisms that sustain global genome organization and support basal nuclear activity.…”

“…Although the features of chromosome organization have been characterized in several organisms, the principles and mechanisms governing the formation of chromosome organization have remained a mystery. In PNAS, Klocko et al (4) report that the Polycomb Repressive Complex 2 (PRC2) and H3K27me3 have a role in shaping genome organization in Neurospora crassa.…”

“…Thus, the detachment of heterochromatin from the silent nuclear peripheral compartments will distort the global genome conformation, especially in organisms constructing their genome structure primarily through heterochromatin interactions. Klocko et al (4) reveal that telomeres detach from the nuclear periphery and move to the interior of the nucleus in the Neurospora Δset-7 strain. As they note, such changes may produce aberrant chromosomal contacts and affect transcription.…”