Project description:In Saccharomyces cerevisiae, the kinase Rio1 regulates rDNA transcription and segregation, pre-rRNA cleavage, and 40S ribosomal subunit maturation. Other roles are unknown. Human orthologue RIOK1; which is frequently overexpressed in malignancies, drives tumor growth and metastasis. Again, also RIOK1 biology is poorly understood. In this study, we charted the global activity of Rio1 in budding yeast. By producing and systems-integrating its protein-interaction, gene-transcription, and chromatin-binding maps we generated Rio1's multi-layered activity network, which controls protein synthesis and turnover, metabolism, growth, proliferation, and genetic stability. Rio1 regulates itself at the transcriptional level, and manages its network both directly and indirectly, via a battery of regulators and transcription factors, including Gcn4. We experimentally confirmed the network and show that Rio1 commands its downstream circuit depending on the growth conditions encountered. We also find that Rio1 and RIOK1 activities are functionally equivalent. Our data suggest that pathological RIOK1 expression may deregulate its network and fuel promiscuous transcription and ribosome production, uncontrolled metabolism, growth, proliferation, and chromosomal instability; well-known contributors to cancer initiation, maintenance and metastasis.
Project description:The conserved Saccharomyces cerevisiae kinase/ATPase Rio1 downregulates rDNA transcription to promote rDNA stability and segregation. To uncover additional roles in transcriptional regulation beyond the rDNA locus we defined the global Rio1 transcriptiome. By NGS we identify 818 differentially expressed genes that are under the transcriptional control of Rio1.
Project description:We report the the first single-molecule-based NGS analysis of chromatin regions that co-immunoprecipitate with Rio1 in the yeast Saccharomyces cerevisiae.
Project description:Kinetochores assemble on centromeres via histone H3 variant CENP-A and low levels of noncoding centromere transcripts (cenRNAs). The latter are ensured by downregulation of RNA polymerase II (RNAPII) and turnover by the nuclear exosome. Using S. cerevisiae, we now add kinase Rio1 to this scheme. Yeast cenRNAs are produced in very low amounts either as short (median lengths of 231nt) or long (4,458nt) transcripts, in a 1:1 ratio. Rio1 limits their production by reducing RNAPII access and transcription activity, and promotes their turnover by the 5’-3’exoribonuclease Rat1. Rio1 similarly curtails the concentrations of noncoding pericenRNAs, which also exist as short transcripts (225nt) at a magnitude higher level than the cenRNAs. In yeast depleted of Rio1, cen- and pericenRNAs accumulatre, and kinetochores misform causing chromosomal instability. The latter phenotypes were also observed with human cells lacking orthologue RioK1, suggesting that CEN regulation by Rio1/RioK1 is evolutionary conserved.
Project description:Kinetochores assemble on centromeres via histone H3 variant CENP-A and low levels of noncoding centromere transcripts (cenRNAs). The latter are ensured by transcription factor Cbf1 (S. cerevisiae), centromere-binding protein CENP-B (humans), and the local histone code downregulating RNA polymerase II (RNAPII) activity. CenRNAs levels are further adjusted by the nuclear exosome. Using S. cerevisiae, we now add kinase Rio1 to this scheme. Yeast cenRNAs are produced mostly from the periCEN regions as short (median lengths of 231nt) or long (4,458nt) transcripts, in a 1:1 ratio. Rio1 limits their production by reducing RNAPII access and transcription activity, and promotes their turnover by the 5’-3’exoribonuclease Rat1. Rio1 similarly curtails the concentrations of noncoding pericenRNAs, which nevertheless exist at a magnitude higher level than the cenRNAs. Similar to yeast, human cells depleted of orthologue RioK1 exhibit cenRNA buildup, kinetochore malformation, and chromosomal instability, suggesting that CEN regulation by Rio1/RioK1 is evolutionary conserved.
Project description:To sustain growth, budding yeast actively transcribes its ribosomal gene array (rDNA) in the nucoleolus to produce ribosomes and proteins. However, intense transcription during rDNA replication may provoke collisions between RNA polymerase I (Pol I) and the replisome, may cause replication fork instability, double-strand breaks, local recombinations and rDNA instability. The latter is manifested by rDNA array expansion or reduction and the formation of extrachromosomal rDNA circles, anomalies that accelerate aging in yeast. Transcription also interferes with the resolution, condensation and segregation of the sister chromatid rDNA arrays. As a consequence, rDNA segregation lags behind the rest of the yeast genome and occurs in late anaphase when rDNA transcription is temporarily shut off. How yeast promotes the stability and transmission of its rDNA array while satisfying a constant need for ribosomes remains unclear. Here we show that the downregulation of Pol I by the conserved cell cycle kinase Rio1 spatiotemporally coordinates rDNA transcription, replication and segregation. More specifically, Rio1 activity promotes copy-number stability of the replicating rDNA array by curtailing Pol I activity and by localising the histone deacetylase Sir2, which establishes a heterochromatic state that silences rDNA transcription. At anaphase entry, Rio1 and the Cdc14 phosphatase target Pol I subunit Rpa43 to dissociate Pol I from the 35S rDNA promoter. The rDNA locus then condensates and segregates, thereby concluding the genome transmission process. Rio1 is involved in ribosome maturation in the cytoplasm of budding yeast and human cells. Additional engagements in the cytoplasm or roles in the nucleus are unknown. Our study describes its first nuclear engagement as a Pol I silencing kinase. This activity may prove highly relevant as dysregulated RNA polymerase I activity has been associated with cancer initiation and proliferation.
Project description:The objective of this study was to determine the global chromosomal interaction map for exponentially growing Saccharomyces cerevisiae cells using Genome Conformation Capture. Interactions between chromosomes were identified within a population of yeast cells growing exponentially in a semi-defined medium containing glucose. The series contains the sequences of the ligated restriction fragments that identified the interactions.
Project description:In our previous work, we had found that Saccharomyces cerevisiae needs of the Hog1 and Slt2 proteins to growth in a low pH environment caused by sulfuric acid, one of the stress factors during the process of ethanol production. Then was performed the gene-wide expression analysis in the hog1∆ and slt2∆ mutants in order to reveal the function of the Hog1p and Slt2p MAP Kinases in the regulation of S. cerevisiae global gene expression upon stress by sulfuric acid.
Project description:We employed CapitalBio Corporation to investigate the global transcriptional profiling of Saccharomyces cerevisiae treated with allicin.
Project description:We employed CapitalBio Corporation to investigate the global transcriptional profiling of Saccharomyces cerevisiae treated with dictamnine. Keywords: response to dictamnine