The Med31 Conserved Component of the Divergent Mediator Complex in Tetrahymena thermophila Participates in Developmental Regulation

Garg, Jyoti; Saettone, Alejandro; Nabeel‐Shah, Syed; Cadorin, Matthew D. R.; Ponce, Marcelo; Marquez, Susanna; Pu, Shuye; Greenblatt, Jack; Pearlman, Ronald E.; Fillingham, Jeffrey

doi:10.1016/j.cub.2019.06.052

Cited by 17 publications

(73 citation statements)

References 61 publications

(71 reference statements)

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“…b Although there are 40 physical cores in the TDS/Niagara nodes, hyperthreading is enabled so it can be used up to 80 logical cores. c Ibd1-2 data set for T.thermophila [16]. d MED31-1 data set for T.thermophila [48]. e MED31-2 data set for T.thermophila [48].…”

Section: Resultsmentioning

confidence: 99%

“…d MED31-1 data set for T.thermophila [48]. e MED31-2 data set for T.thermophila [48]. f Data set for O.trifallax .As it can be seen, the working space (in this case memory utilization ) can reach up to a factor of 9-10 × the size of the initial data to be processed.…”

Section: Resultsmentioning

confidence: 99%

“…Other tools, such as, MACS2 and metagene using deepTools analysis [45] complement RACS. Recently several ChIP-Seq studies [46–48] have emerged for T.thermophila . However, there is a lack of standardized computational methods for this model organism, hence it becomes difficult to reliably reach at the same conclusions when replicating the findings.…”

Section: Discussionmentioning

confidence: 99%

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RACS: rapid analysis of ChIP-Seq data for contig based genomes

et al. 2019

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BackgroundChromatin immunoprecipitation coupled to next generation sequencing (ChIP-Seq) is a widely-used molecular method to investigate the function of chromatin-related proteins by identifying their associated DNA sequences on a genomic scale. ChIP-Seq generates large quantities of data that is difficult to process and analyze, particularly for organisms with a contig-based sequenced genomes that typically have minimal annotation on their associated set of genes other than their associated coordinates primarily predicted by gene finding programs. Poorly annotated genome sequence makes comprehensive analysis of ChIP-Seq data difficult and as such standardized analysis pipelines are lacking.ResultsWe present a one-stop computational pipeline, “Rapid Analysis of ChIP-Seq data” (RACS), that utilizes traditional High-Performance Computing (HPC) techniques in association with open source tools for processing and analyzing raw ChIP-Seq data. RACS is an open source computational pipeline available from any of the following repositories https://bitbucket.org/mjponce/RACS or https://gitrepos.scinet.utoronto.ca/public/?a=summary&p=RACS. RACS is particularly useful for ChIP-Seq in organisms with contig-based genomes that have poor gene annotation to aid protein function discovery.To test the performance and efficiency of RACS, we analyzed ChIP-Seq data previously published in a model organism Tetrahymena thermophila which has a contig-based genome. We assessed the generality of RACS by analyzing a previously published data set generated using the model organism Oxytricha trifallax, whose genome sequence is also contig-based with poor annotation.ConclusionsThe RACS computational pipeline presented in this report is an efficient and reliable tool to analyze genome-wide raw ChIP-Seq data generated in model organisms with poorly annotated contig-based genome sequence. Because RACS segregates the found read accumulations between genic and intergenic regions, it is particularly efficient for rapid downstream analyses of proteins involved in gene expression.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

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RACS: rapid analysis of ChIP-Seq data for contig based genomes

et al. 2019

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“…The Mediator transcriptional co-activator complex is widely required for transcription in yeast and human cells. Through AP-MS of conserved Mediator subunit Med31-FZZ [146], we recently identified a divergent Tetrahymena Mediator complex. Indirect immunofluorescence of Med31-FZZ demonstrates it to localize to the MAC during growth and development consistent with function as a transcriptional regulator and also to the meiotic MIC implicating it in expression of ncRNAs [146].…”

Section: Transcriptionmentioning

confidence: 99%

Functional Proteomics of Nuclear Proteins in Tetrahymena thermophila: A Review

Saettone

Nabeel‐Shah

Garg

et al. 2019

Genes

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Identification and characterization of protein complexes and interactomes has been essential to the understanding of fundamental nuclear processes including transcription, replication, recombination, and maintenance of genome stability. Despite significant progress in elucidation of nuclear proteomes and interactomes of organisms such as yeast and mammalian systems, progress in other models has lagged. Protists, including the alveolate ciliate protozoa with Tetrahymena thermophila as one of the most studied members of this group, have a unique nuclear biology, and nuclear dimorphism, with structurally and functionally distinct nuclei in a common cytoplasm. These features have been important in providing important insights about numerous fundamental nuclear processes. Here, we review the proteomic approaches that were historically used as well as those currently employed to take advantage of the unique biology of the ciliates, focusing on Tetrahymena, to address important questions and better understand nuclear processes including chromatin biology of eukaryotes.

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“…Both the MAC ( Eisen et al, 2006 ) and MIC ( Hamilton et al, 2016 ) genomes have been sequenced and annotated ( Stover et al, 2006 ). A well-developed functional proteomic pipeline exists for the study of epigenetic regulators in particular for a ffinity p urification coupled to m ass s pectrometry (AP-MS) to effectively solubilize native (i.e., un-crosslinked) protein complexes both bound and unbound to chromatin ( Xiong et al, 2011 ; Garg et al, 2013 , 2019 ; Saettone et al, 2018 , 2019a ; Ashraf et al, 2019 ; Nabeel-Shah et al, 2020 ). A critical feature of this approach is the expression of the epitope tagged proteins at levels closely approximating that of the endogenous protein.…”

Section: Introduction/backgroundmentioning

confidence: 99%

Exploring the Histone Acetylation Cycle in the Protozoan Model Tetrahymena thermophila

Wahab

Saettone

Nabeel‐Shah

et al. 2020

Front. Cell Dev. Biol.

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The eukaryotic histone acetylation cycle is composed of three classes of proteins, histone acetyltransferases (HATs) that add acetyl groups to lysine amino acids, bromodomain (BRD) containing proteins that are one of the most characterized of several protein domains that recognize acetyl-lysine (Kac) and effect downstream function, and histone deacetylases (HDACs) that catalyze the reverse reaction. Dysfunction of selected proteins of these three classes is associated with human disease such as cancer. Additionally, the HATs, BRDs, and HDACs of fungi and parasitic protozoa present potential drug targets. Despite their importance, the function and mechanisms of HATs, BRDs, and HDACs and how they relate to chromatin remodeling (CR) remain incompletely understood. Tetrahymena thermophila (Tt) provides a highly tractable single-celled free-living protozoan model for studying histone acetylation, featuring a massively acetylated somatic genome, a property that was exploited in the identification of the first nuclear/type A HAT Gcn5 in the 1990s. Since then, Tetrahymena remains an under-explored model for the molecular analysis of HATs, BRDs, and HDACs. Studies of HATs, BRDs, and HDACs in Tetrahymena have the potential to reveal the function of HATs and BRDs relevant to both fundamental eukaryotic biology and to the study of disease mechanisms in parasitic protozoa.

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The Med31 Conserved Component of the Divergent Mediator Complex in Tetrahymena thermophila Participates in Developmental Regulation

Cited by 17 publications

References 61 publications

RACS: rapid analysis of ChIP-Seq data for contig based genomes

RACS: rapid analysis of ChIP-Seq data for contig based genomes

Functional Proteomics of Nuclear Proteins in Tetrahymena thermophila: A Review

Exploring the Histone Acetylation Cycle in the Protozoan Model Tetrahymena thermophila

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