The Molecular Mechanisms Underlying The Regulation Of Catalase-3 Expression By Histone Variant H2A.Z In Neurospora Crassa

Eukaryotic DNA is highly condensed into chromatins,which alters the accessibility and behavior of the DNA and recognition by the transcription factors and RNA polymerase.Thus,it is generally believed that nucleosomes must be remodeled at certain regions of a given gene to increase DNA accessibility in order to trigger efficient gene expression by transcription factors during development or stress responses.The replacement of histone variants on the core histones of nucleosomes and post-translational modifications on the tails of core histones contribute to the stability of the modified nucleosomes at defined regions of the genome.H2A.Z is a histone variant of the canonical H2A family.And it is an evolutionarily conserved histone variant which shares higher sequence similarity among different eukaryotic species than to the canonical H2A within the same organism,suggesting a functionally distinct role for H2A.Z on gene expression and genomic stability compared to H2A.In eukaryotes,deposition of the histone variant H2A.Z into nucleosomes through the chromatin remodeling complex,SWR1,is a crucial step in modulating gene transcription.Recently,H2A.Z has been shown to control the expression of responsive genes,but the underlying mechanism of how H2A.Z responds to physiological stimuli is not well understood.In this study,we explored the role of H2A.Z in transcriptional regulation of cat-3 gene which can be induced by oxidative stress from environmental or intracellular stimuli.Unlike its essential role in other organisms,H2A.Z knock-out mutant in N.crassa is viable.We reveal that,in Neurospora crassa,H2A.Z is a negative regulator of catalase-3 gene,which is responsible for resistance to oxidative stress.H2A.Z represses cat-3 gene expression through direct incorporation at cat-3 locus in a SWR1 complex dependent pathway.Loss of H2A.Z or SWR1 subunits leads to increased binding of a sequence-specific transcription factor,CPC1,at cat-3 locus.Additionally,introduction of plasmids containing gene encoding H2A.Z or SWR1 complex subunits into wild-type strains decreased CAT-3 expression,indicating that H2A.Z counteracts the positive effect of CPC1 to achieve low level cat-3 expression under non-inductive condition.Furthermore,upon oxidative stress,H2A.Z is rapidly evicted from cat-3 locus for the recruitment of CPC1,resulting in robust and full cat-3 gene expression in response to external stimuli.Interestingly,the reduction of H2A.Z at ORF locus of cat-3 gene promotes me to detect whether the loss of H2A.Z results in enhancing elongation rate of RNA Pol ?.ChIP results showed that the enrichment of Ser2p of RNA Pol ? and SET-2 increased at the ORF of cat-3 in H2A.ZKO mutants,suggesting that H2A.Z might be a barrier for transcriptional elongation.Finally,we observed that NCU00354 transcription elevated in the H2A.2KO mutants and swr defective strains,and the expression of NCU00354 cannot be induced efficiently in these mutants under oxidative stress compared to the wild-type.The results suggest that H2A.Z might carry a function required for local environment.Collectively,this study strongly demonstrates that H2A.Z antagonizes the function of transcription factor to regulate responsive gene transcription under normal conditions and to poise for gene full activation under oxidative stress.