Heterochromatin has puzzled scientists for decades. It was identified almost 90 years ago by its darker appearance under the microscope in relation to euchromatin. Only later did scientists begin to understand that this tightly packaged DNA silences gene expression and has roles in chromosome maintenance. Today, heterochromatin continues to maintain its silence, only reluctantly giving up secrets to its quiescence.
Danesh Moazed, HMS professor of cell biology, and colleagues have just uncovered two mechanisms that contribute to heterochromatin silencing in fission yeast. Paradoxically, both depend on the very thing heterochromatin is thought to prevent, transcription.
The first mechanism relies on RNA interference. Though RNAi has been linked to heterochromatin silencing before—key RNAi enzymes, including Dicer and Argonaute, are essential for heterochromatin formation—it is unclear whether RNAi targets heterochromatic transcripts themselves or merely regulates other genes that contribute to heterochromatin formation. If the former, then cells would have to make small interfering RNAs (siRNAs) that complement heterochromatin genes; however, various research labs, including Moazed’s, have failed to detect such siRNAs in yeast—until now.
To pinpoint heterochromatin siRNAs, Moazed and colleagues, including HMS associate professor of cell biology Steven Gygi, turned to a more sensitive detection method and to yeast cells that are deficient in exoribonuclease, which degrades siRNA. Writing in the May 18 Cell, postdoctoral research fellow Marc Buhler and colleagues say that when they inserted the reporter gene ura4+ into yeast centromeric heterochromatin, they were able to detect ura4+ siRNAs with the more sensitive assay, albeit in low amounts. Nevertheless, “because siRNAs are a signature of processing by RNAi, this establishes RNAi as one part of the mechanism that contributes to heterochromatin silencing,” said Moazed.
But it turns out that this is only part of the picture, because when Buhler inserted the ura4+ transgene into a different heterochromatin location called the silent mating–type locus, ura4+ siRNAs were not detected. And that’s where the second mechanism comes in. The researchers wondered if the exosome, a nuclear ribonuclease complex that degrades aberrant RNA transcripts, might also play a role in gene silencing. Specifically, they tested whether a protein called Cid14 might be involved because it is a homolog of a budding yeast polyadenylase that marks transcripts for exosome-mediated degradation. Sure enough, by knocking out Cid14 in fission yeast, the researchers abolished silencing of the ura4+ reporter.
If these mechanisms are conserved in humans, it could have important implications for RNAi-based therapies. “It suggests that you may be able to design RNAi therapies that can have permanent effects at the chromatin level, leading to very stable changes in gene expression patterns,” said Moazed.
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