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Significance The circadian clock is predominantly regulated by transcriptional negative feedback, where the protein(s) arising from the central clock gene(s) inhibit their own expression. InNeurospora and mammals, the clock genes have natural antisense transcripts (NATs): long noncoding RNAs that overlap and are expressed in opposite orientation to the protein coding genes. Previously, we demonstrated thatNeurospora frequency NAT is needed for normal DNA methylation. This report demonstrates that proper regulation of thefrequency antisense transcript is needed to establish repressive heterochromatin. However, prior to establishing heterochromatin, NAT expression creates a transcriptionally permissive state that helps promote sense transcript expression. Broader implications indicate that NATs, in general, may first promote sense gene expression prior to establishing heterochromatin.

The frequency natural antisense transcript first promotes, then represses, frequency gene expression via facultative heterochromatin