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A phospho-mimicking Atf1 mutant bypasses the transcription activating function of the MAP kinase Sty1

ABSTRACT: The MAP kinase Sty1 of fission yeast accumulates in the nucleus upon stress, and stimulates expression of hundreds of genes, including atf1, via the constitutively nuclear transcription factor Atf1. The role on transcription of the signal-dependent phosphorylation of Atf1 by Sty1 is unclear. We have expressed the transcription factor and mutant derivatives from a constitutive promoter, to unchain Atf1 activity from its protein levels. Expression of a non-phosphorylatable Atf1 renders cells sensitive to oxidative stress, due to the impaired transcription of a subset of genes, those dependent on another transcription factor, Pap1. Cells expressing a phospho-mimicking mutant display enhanced resistance to stress, and while expression of the Pap1-dependent genes is still stress-dependent, a second subset of genes is constitutively expressed. Our study demonstrates that phosphorylation of Atf1 by Sty1 does not stimulate binding of the transcription factor to DNA but rather establishes a platform of interaction with the transcriptional machinery.

ORGANISM(S): Schizosaccharomyces pombe

PROVIDER: GSE97057 | GEO | 2017/07/10

SECONDARY ACCESSION(S): PRJNA380603

REPOSITORIES: GEO

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Project description:The high osmolarity glycerol response (HOG) pathway plays a pivotal role in the stress response, virulence regulation, and differentiation of fungi, including Cryptococcus neoformans that causes fatal meningoencephalitis. Core signaling components of in the HOG pathway, including the Tco-Ypd1-Ssk1 phosphorelay system and the Ssk2-Pbs2-Hog1 MAPK module, have been elucidated but its downstream transcription factors remain unclear. Here we demonstrated that Atf1 with a basic leucine zipper domain is the transcription factor downstream of Hog1 in C. neoformans. We found that ATF1 expression was differentially regulated by oxidative damaging agents, mainly in a Hog1-dependent, but Mpk1-independent, manner. Interestingly, Atf1 not only promoted oxidative stress response and adaptation, but also played an opposing role to Hog1 in the process. Atf1 primarily localized to the nucleus under both unstressed and oxidative stress conditions in a Hog1-independent manner. Our data demonstrated that Atf1 promoted pheromone production and sexual differentiation under negative control by Hog1. Finally, a DNA microarray-based transcriptome analysis of the atf1M-bM-^HM-^F mutant under unstressed and oxidative stress conditions revealed that Atf1 regulated oxidative stress response genes, including a sulfiredoxin gene (SRX1). Intriguing, the array data further demonstrated that Atf1 modulated basal expression of genes involved in DNA repair and genotoxic stress response. Supporting this, we found that the atf1M-bM-^HM-^F mutant was highly sensitive to genotoxic agents. In conclusion, this study provided a further insight into the Hog1-dependent oxidative and genotoxic stress response and differentiation mechanism of C. neoformans. There are more than 95% of genome homology between JEC21 and H99. Therefore 6 slides of JEC21 (Cryptococcus neoformans var. neoformans serotype D) 70-mer oligo are used in this analysis, 3 biological replicate experiments are performed, total RNAs are extracted 2 conditions (with or without treatment of Hydroten peroxide) from H99 (H99 Wild type strain (Cryptococcus neoformans var. grubii serotype A), and atf1M-NM-^T). We use the mixed all of total RNAs from this experiment as a control RNA. We use Cy3 as Sample dye and Cy5 as a control dye.

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A phospho-mimicking Atf1 mutant bypasses the transcription activating function of the MAP kinase Sty1

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