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Transcriptional interference by antisense RNA is required for circadian clock function.

Xue Z, Ye Q, Anson SR, Yang J, Xiao G, Kowbel D, Glass NL, Crosthwaite SK, Liu Y - Nature (2014)

Bottom Line: Natural antisense RNAs are found in a wide range of eukaryotic organisms.Moreover, our results suggest that antisense transcription inhibits sense expression by mediating chromatin modifications and premature termination of transcription.Taken together, our results establish antisense transcription as an essential feature in a circadian system and shed light on the importance and mechanism of antisense action.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, The University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, Texas 75390, USA.

ABSTRACT
Eukaryotic circadian oscillators consist of negative feedback loops that generate endogenous rhythmicities. Natural antisense RNAs are found in a wide range of eukaryotic organisms. Nevertheless, the physiological importance and mode of action of most antisense RNAs are not clear. frequency (frq) encodes a component of the Neurospora core circadian negative feedback loop, which was thought to generate sustained rhythmicity. Transcription of qrf, the long non-coding frq antisense RNA, is induced by light, and its level oscillates in antiphase to frq sense RNA. Here we show that qrf transcription is regulated by both light-dependent and light-independent mechanisms. Light-dependent qrf transcription represses frq expression and regulates clock resetting. Light-independent qrf expression, on the other hand, is required for circadian rhythmicity. frq transcription also inhibits qrf expression and drives the antiphasic rhythm of qrf transcripts. The mutual inhibition of frq and qrf transcription thus forms a double negative feedback loop that is interlocked with the core feedback loop. Genetic and mathematical modelling analyses indicate that such an arrangement is required for robust and sustained circadian rhythmicity. Moreover, our results suggest that antisense transcription inhibits sense expression by mediating chromatin modifications and premature termination of transcription. Taken together, our results establish antisense transcription as an essential feature in a circadian system and shed light on the importance and mechanism of antisense action.

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Neither RNAi nor DNA methylation pathways play a significant role in the clock. (a-b) Strand-specific RT-qPCR results showing the induction of frq after a light pulse are similar in the indicated trains at DD24. (c) Strand-specific RT-qPCR results showing similar expression levels of frq and qrf in the indicated strains in LL. (d) A table showing the period lengths of the conidiation rhythms in the wild-type and different RNAi mutants. (e-f) Strand-specific RT-qPCR results showing the induction of frq after a light pulse is similar in the indicated trains at DD24. Error bars are standard deviations (n=3).
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Figure 5: Neither RNAi nor DNA methylation pathways play a significant role in the clock. (a-b) Strand-specific RT-qPCR results showing the induction of frq after a light pulse are similar in the indicated trains at DD24. (c) Strand-specific RT-qPCR results showing similar expression levels of frq and qrf in the indicated strains in LL. (d) A table showing the period lengths of the conidiation rhythms in the wild-type and different RNAi mutants. (e-f) Strand-specific RT-qPCR results showing the induction of frq after a light pulse is similar in the indicated trains at DD24. Error bars are standard deviations (n=3).

Mentions: How do qrf and frq inhibit each other? Light-induced frq transcription, frq and qrf levels and circadian rhythms were normal in RNAi mutants (Extended Data Figure 5a-d)19. Convergent transcription is also known to induce DNA methylation in the frq region20,21 but deletion of genes required for DNA methylation also did not affect frq expression (Extended Data Figure 5e-f).


Transcriptional interference by antisense RNA is required for circadian clock function.

Xue Z, Ye Q, Anson SR, Yang J, Xiao G, Kowbel D, Glass NL, Crosthwaite SK, Liu Y - Nature (2014)

Neither RNAi nor DNA methylation pathways play a significant role in the clock. (a-b) Strand-specific RT-qPCR results showing the induction of frq after a light pulse are similar in the indicated trains at DD24. (c) Strand-specific RT-qPCR results showing similar expression levels of frq and qrf in the indicated strains in LL. (d) A table showing the period lengths of the conidiation rhythms in the wild-type and different RNAi mutants. (e-f) Strand-specific RT-qPCR results showing the induction of frq after a light pulse is similar in the indicated trains at DD24. Error bars are standard deviations (n=3).
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4214883&req=5

Figure 5: Neither RNAi nor DNA methylation pathways play a significant role in the clock. (a-b) Strand-specific RT-qPCR results showing the induction of frq after a light pulse are similar in the indicated trains at DD24. (c) Strand-specific RT-qPCR results showing similar expression levels of frq and qrf in the indicated strains in LL. (d) A table showing the period lengths of the conidiation rhythms in the wild-type and different RNAi mutants. (e-f) Strand-specific RT-qPCR results showing the induction of frq after a light pulse is similar in the indicated trains at DD24. Error bars are standard deviations (n=3).
Mentions: How do qrf and frq inhibit each other? Light-induced frq transcription, frq and qrf levels and circadian rhythms were normal in RNAi mutants (Extended Data Figure 5a-d)19. Convergent transcription is also known to induce DNA methylation in the frq region20,21 but deletion of genes required for DNA methylation also did not affect frq expression (Extended Data Figure 5e-f).

Bottom Line: Natural antisense RNAs are found in a wide range of eukaryotic organisms.Moreover, our results suggest that antisense transcription inhibits sense expression by mediating chromatin modifications and premature termination of transcription.Taken together, our results establish antisense transcription as an essential feature in a circadian system and shed light on the importance and mechanism of antisense action.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, The University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, Texas 75390, USA.

ABSTRACT
Eukaryotic circadian oscillators consist of negative feedback loops that generate endogenous rhythmicities. Natural antisense RNAs are found in a wide range of eukaryotic organisms. Nevertheless, the physiological importance and mode of action of most antisense RNAs are not clear. frequency (frq) encodes a component of the Neurospora core circadian negative feedback loop, which was thought to generate sustained rhythmicity. Transcription of qrf, the long non-coding frq antisense RNA, is induced by light, and its level oscillates in antiphase to frq sense RNA. Here we show that qrf transcription is regulated by both light-dependent and light-independent mechanisms. Light-dependent qrf transcription represses frq expression and regulates clock resetting. Light-independent qrf expression, on the other hand, is required for circadian rhythmicity. frq transcription also inhibits qrf expression and drives the antiphasic rhythm of qrf transcripts. The mutual inhibition of frq and qrf transcription thus forms a double negative feedback loop that is interlocked with the core feedback loop. Genetic and mathematical modelling analyses indicate that such an arrangement is required for robust and sustained circadian rhythmicity. Moreover, our results suggest that antisense transcription inhibits sense expression by mediating chromatin modifications and premature termination of transcription. Taken together, our results establish antisense transcription as an essential feature in a circadian system and shed light on the importance and mechanism of antisense action.

Show MeSH