Investigation Of Transcriptional Regulation Mechanism Of Catalase Genes In Neurospora Crassa

The precise regulation of gene expression is essential for living organisms’ development and it also underlies the ability to respond to stimuli. Neurospora crassa has three catalases:Cat-1, Cat-2 and Cat-3. Cat-3 is the most important catalase which can detoxify H2O2 efficiently and regulate the asexual development of Neurospora. For Neurospora crassa, the tightly regulated expression of cat-3 represents a good system to study the mechanisms which control the specificity of gene expression. Our studies revealed that Cat-3 protein levels were increased in mutants defective in proper heterochromatin formation, including dim-5KO, hpoKO,H3K9Q, H3K9L, H3K9R and cul4KO, dcaf26KO, dim-7KO strains. Consistently, these mutants showed strong H2O2 resistance. Sequence analysis revealed a 5kb AT-rich region adjacent to the cat-3 promoter, and consequently H3K9me3 and HP1 were enriched in this region, indicating that heterochromatin forms at upstream of cat-3 promoter and is sufficient to suppress the expression of cat-3. Moreover, H3ac levels on cat-3 locus were increased in H3K9 methyltransferase mutant dim-5KO similar to those in H2O2-treated wild-type. We also detected the catalase zymograms in the strains that had defective heterochromatin formation, such as dim-5KO, these mutants exhibited elevated Cat-1 and Cat-2 activities and the more smearing bands of Cat-3.To investigate a direct correlation between cat-3 expression and its proximal heterochromatin, we generated the disruption strains of 5kb intergenic region, cat-3Δ5-3, cat-3Δ3-1, cat-3Δ5. The deletion strains exhibited slight resistance to H2O2 and higher accumulation of Cat-3 than those in dim-5KO and wild-type. And Cat-3 levels in cat-3Δ5-3 were more elevated than that in cat-3Δ3-1, cat-3Δ5. Moreover, even higher levels of Cat-3 were promoted by the combination with global heterochromatin-defective mutants. The enrichment of RNA Pol Ⅱ at cat-3 promoter regions were increased in the deletion strains, especially in cat-3Δ5, suggesting that the 5kb upstream region of cat-3 affected the transcription process of cat-3. However, unlike the lower electrophoretic mobility in dim-5KO, the Cat-3 protein in deletion strains exhibited similar pattern with that in wild-type, suggesting that modification of Cat-3 protein wasn’t altered in the deletion mutants.To sum up, in this research we investigated the transcriptional regulation mechanism of Neurospora catalase genes using genetics and molecular biological means. This study could provid some supports to understand the transcription regulation mechanism of related genes.