ABSTRACT
In eukaryotes, repetitive DNA can become silenced de novo, either transcriptionally or post-transcriptionally, by processes independent of strong sequence-specific cues. The mechanistic nature of such processes remains poorly understood. We found that in the fungus Neurospora crassa, de novo initiation of both transcriptional and post-transcriptional silencing was linked to perturbed chromatin, which was produced experimentally by the aberrant activity of transcription factors at the tetO operator array. Transcriptional silencing was mediated by canonical constitutive heterochromatin. On the other hand, post-transcriptional silencing resembled repeat-induced quelling but occurred normally when homologous recombination was inactivated. All silencing of the tetO array was dependent on SAD-6 (a fungal ortholog of the SWI/SNF chromatin remodeler ATRX), which was required to maintain nucleosome occupancy at the perturbed locus. In addition, we found that two other types of sequences (the lacO array and native AT-rich DNA) could also undergo recombination-independent quelling associated with perturbed chromatin. These results suggested a model in which the de novo initiation of transcriptional and post-transcriptional silencing is coupled to the remodeling of perturbed chromatin.
SIGNIFICANCE STATEMENT This study addresses an enigmatic question of how transcriptional and post-transcriptional gene silencing can be initiated de novo in the absence of strong sequence-specific cues. Using the fungus Neurospora crassa as a model organism, we found that both types of silencing can be triggered in mitotic cells by the remodeling of a transiently perturbed (nucleosome-depleted) chromatin state. In this system, the initiation of silencing requires SAD-6, a conserved SWI/SNF chromatin remodeler orthologous to ATRX that has been already implicated in repetitive DNA silencing in fungi, plants, and animals. Thus, the model proposed in this study may underpin a range of gene-silencing phenomena observed in other eukaryotes.
Competing Interest Statement
The authors have declared no competing interest.
Footnotes
Incorporated fluorescence microscopy; several figures and text were also edited for clarity.
