Suppression of WHITE COLLAR-independent frequency Transcription by Histone H3 Lysine 36 Methyltransferase SET-2 Is Necessary for Clock Function in Neurospora

Guangyan Sun,Zhipeng Zhou,Xiao Liu,Kexin Gai,Qingqing Liu,Joonseok Cha,Farah Naz Kaleri,Ying Wang,Qun He

doi:10.1074/jbc.m115.711333

Abstract

The circadian system in Neurospora is based on the transcriptional/translational feedback loops and rhythmic frequency (frq) transcription requires the WHITE COLLAR (WC) complex. Our previous paper has shown that frq could be transcribed in a WC-independent pathway in a strain lacking the histone H3K36 methyltransferase, SET-2 (su(var)3-9-enhancer-of-zeste-trithorax-2) (1), but the mechanism was unclear. Here we disclose that loss of histone H3K36 methylation, due to either deletion of SET-2 or H3K36R mutation, results in arrhythmic frq transcription and loss of overt rhythmicity. Histone acetylation at frq locus increases in set-2(KO) mutant. Consistent with these results, loss of H3K36 methylation readers, histone deacetylase RPD-3 (reduced potassium dependence 3) or EAF-3 (essential SAS-related acetyltransferase-associated factor 3), also leads to hyperacetylation of histone at frq locus and WC-independent frq expression, suggesting that proper chromatin modification at frq locus is required for circadian clock operation. Furthermore, a mutant strain with three amino acid substitutions (histone H3 lysine 9, 14, and 18 to glutamine) was generated to mimic the strain with hyperacetylation state of histone H3. H3K9QK14QK18Q mutant exhibits the same defective clock phenotype as rpd-3(KO) mutant. Our results support a scenario in which H3K36 methylation is required to establish a permissive chromatin state for circadian frq transcription by maintaining proper acetylation status at frq locus.

Highlights

Despite evolutionary distance, circadian clock oscillators are conserved among the organisms to perform their cellular and behavioral activities
As there are nonspecific bands recognized by SET-2 antibody, set-2KO strain was used as the negative control in the chromatin immunoprecipitation (ChIP) assay
These results indicate that SET-2 is recruited to frq locus during transcription (Fig. 1, B and C)

Summary

Introduction

Circadian clock oscillators are conserved among the organisms to perform their cellular and behavioral activities. Our previous study indicates that RCO-1 functions to regulate H3K36 methylation levels at the frq locus to suppress WC-independent frq transcription. Consistent with the decreased H3K36me3 and increased H3K9ac at frq locus in rco-1KO mutant, deletion of set-2 gene results in WC-independent frq transcription and impaired circadian rhythm [1], suggesting that crosstalk of different histone modifications correlates with the rhythmic transcription of frq gene.

Results

Conclusion