scholarly journals High-Resolution Mapping of Transcription Initiation in the Asexual Stages of Toxoplasma gondii

2021 ◽  
Vol 10 ◽  
Author(s):  
Benedikt M. Markus ◽  
Benjamin S. Waldman ◽  
Hernan A. Lorenzi ◽  
Sebastian Lourido
Keyword(s):  
Toxoplasma Gondii ◽  
Transcription Initiation ◽  
Nucleosome Occupancy ◽  
Mrna Isoforms ◽  
Protein Coding ◽  
Transcription Start Sites ◽  
Promoter Architecture ◽  
Depth Analysis ◽  
Life Threatening ◽  
Specific Alternative

Toxoplasma gondii is a common parasite of humans and animals, causing life-threatening disease in the immunocompromized, fetal abnormalities when contracted during gestation, and recurrent ocular lesions in some patients. Central to the prevalence and pathogenicity of this protozoan is its ability to adapt to a broad range of environments, and to differentiate between acute and chronic stages. These processes are underpinned by a major rewiring of gene expression, yet the mechanisms that regulate transcription in this parasite are only partially characterized. Deciphering these mechanisms requires a precise and comprehensive map of transcription start sites (TSSs); however, Toxoplasma TSSs have remained incompletely defined. To address this challenge, we used 5′-end RNA sequencing to genomically assess transcription initiation in both acute and chronic stages of Toxoplasma. Here, we report an in-depth analysis of transcription initiation at promoters, and provide empirically-defined TSSs for 7603 (91%) protein-coding genes, of which only 1840 concur with existing gene models. Comparing data from acute and chronic stages, we identified instances of stage-specific alternative TSSs that putatively generate mRNA isoforms with distinct 5′ termini. Analysis of the nucleotide content and nucleosome occupancy around TSSs allowed us to examine the determinants of TSS choice, and outline features of Toxoplasma promoter architecture. We also found pervasive divergent transcription at Toxoplasma promoters, clustered within the nucleosomes of highly-symmetrical phased arrays, underscoring chromatin contributions to transcription initiation. Corroborating previous observations, we asserted that Toxoplasma 5′ leaders are among the longest of any eukaryote studied thus far, displaying a median length of approximately 800 nucleotides. Further highlighting the utility of a precise TSS map, we pinpointed motifs associated with transcription initiation, including the binding sites of the master regulator of chronic-stage differentiation, BFD1, and a novel motif with a similar positional arrangement present at 44% of Toxoplasma promoters. This work provides a critical resource for functional genomics in Toxoplasma, and lays down a foundation to study the interactions between genomic sequences and the regulatory factors that control transcription in this parasite.

scholarly journals RSC-mediated nucleosome positioning and GRFs form barriers in promoters to limit divergent noncoding transcription

2021 ◽  
Author(s):  
Andrew Wu ◽  
Claudia Vivori ◽  
Harshil Patel ◽  
Theodora Sideri ◽  
Folkert van Werven
Keyword(s):  
Transcription Initiation ◽  
Nucleosome Occupancy ◽  
Nucleosome Positioning ◽  
Gene Promoters ◽  
Protein Coding ◽  
Transcription Start Sites ◽  
Binding Factor ◽  
Cryptic Transcription ◽  
Divergent Transcription ◽  
Noncoding Transcription

The directionality of gene promoters - the ratio of protein-coding over divergent noncoding transcription - is highly variable and regulated. How promoter directionality is controlled remains poorly understood. We show that the chromatin remodelling complex RSC and general regulatory factors (GRFs) dictate promoter directionality by attenuating divergent transcription. Depletion of RSC increased divergent noncoding transcription and decreased protein-coding transcription at promoters with strong directionality. Consistent with RSCs role in regulating chromatin, RSC depletion impacts nucleosome occupancy upstream of the nucleosome depleted region where divergent transcription initiates, suggesting that nucleosome positioning at the 5 prime border of gene promoters physically blocks the recruitment of the transcription machinery and inhibits initiation of divergent transcription. Highly directional promoters were also enriched for the binding of GRFs such as Reb1 and Abf1. Furthermore, ectopic targeting of divergent transcription initiation sites with GRFs or the dCas9 protein can suppress divergent transcription. Our data suggest that RSC-mediated nucleosome positioning and GRFs play a pervasive role in repressing divergent transcription. We propose that any DNA binding factor, when stably associated with cryptic transcription start sites, can form a barrier for repressing divergent transcription. Our study provides an explanation as to why certain promoters are more directional than others.

scholarly journals A selector of transcription initiation in the protozoan parasite Toxoplasma gondii.

1995 ◽  
Vol 15 (1) ◽  
pp. 87-93 ◽  
Author(s):  
D Soldati ◽  
J C Boothroyd
Keyword(s):  
Toxoplasma Gondii ◽  
Gene Transcription ◽  
Transcription Initiation ◽  
Protozoan Parasite ◽  
Intracellular Parasite ◽  
Transcription Start ◽  
Transcription Start Sites ◽  
Obligate Intracellular ◽  
Obligate Intracellular Parasite ◽  
Sag1 Gene

The recent development of an efficient transfection system for the apicomplexan Toxoplasma gondii allows a comprehensive dissection of the elements involved in gene transcription in this obligate intracellular parasite. We demonstrate here that for the SAG1 gene, a stretch of six repeated sequences in the region 35 to 190 bp upstream of the first of two transcription start sites is essential for efficient and accurate transcription initiation. This repeat element shows characteristics of a selector in determining the position of the transcription start sites.

scholarly journals Transcription initiation mapping in 31 bovine tissues reveals complex promoter activity, pervasive transcription, and tissue-specific promoter usage

2020 ◽  
Author(s):  
D.E. Goszczynski ◽  
M.M. Halstead ◽  
A.D. Islas-Trejo ◽  
H. Zhou ◽  
P.J. Ross
Keyword(s):  
Transcription Initiation ◽  
Promoter Activity ◽  
Bovine Genome ◽  
Transcription Start ◽  
Protein Coding ◽  
Tissue Specific ◽  
Transcription Start Sites ◽  
Expression Control ◽  
Tissue Specific Promoter ◽  
Genome Annotations

ABSTRACTCharacterizing transcription start sites is essential for understanding the regulatory mechanisms that control gene expression. Recently, a new bovine genome assembly (ARS-UCD1.2) with high continuity, accuracy, and completeness was released; however, the functional annotation of the bovine genome lacks precise transcription start sites and includes a low number of transcripts in comparison to human and mouse. Using the RAMPAGE approach, this study identified transcription start sites at high resolution in a large collection of bovine tissues. We found several known and novel transcription start sites attributed to promoters of protein coding and lncRNA genes that were validated through experimental and in silico evidence. With these findings, the annotation of transcription start sites in cattle reached a level comparable to the mouse and human genome annotations. In addition, we identified and characterized transcription start sites for antisense transcripts derived from bidirectional promoters, potential lncRNAs, mRNAs, and pre-miRNAs. We also analyzed the quantitative aspects of RAMPAGE data for producing a promoter activity atlas, reaching highly reproducible results comparable to traditional RNA-Seq. Lastly, gene co-expression networks revealed an impressive use of tissue-specific promoters, especially between brain and testicle, which expressed several genes in common from alternate transcription start sites. Regions surrounding co-expressed modules were enriched in binding factor motifs representative of their tissues. This annotation will be highly useful for future studies on expression control in cattle and other species. Furthermore, these data provide significant insight into transcriptional activity for a comprehensive set of tissues.

scholarly journals The origin and evolution of a distinct mechanism of transcription initiation in yeasts

2020 ◽  
Author(s):  
Zhaolian Lu ◽  
Zhenguo Lin
Keyword(s):  
Rna Polymerase Ii ◽  
Transcription Initiation ◽  
Core Promoter ◽  
Model Species ◽  
Initiation Mechanism ◽  
Protein Coding ◽  
Origin And Evolution ◽  
Transcription Start Sites ◽  
Mrna Maturation ◽  
Nucleotide Resolution

ABSTRACTThe molecular process of transcription by RNA Polymerase II is highly conserved among eukaryotes (“classic model”). Intriguingly, a distinct way of locating transcription start sites (TSSs) was found in a budding yeast Saccharomyces cerevisiae (“scanning model”). The origin of the “scanning model” and its underlying genetic mechanisms remain unsolved. Herein, we applied genomic approaches to address these questions. We first identified TSSs at a single-nucleotide resolution for 12 yeast species using the nAnT-iCAGE technique, which significantly improved the annotations of these genomes by providing accurate 5’boundaries of protein-coding genes. We then infer the initiation mechanism of a species based on its TSS maps and genome sequences. We found that the “scanning model” had originated after the split of Yarrowia lipolytica and the rest of budding yeasts. An adenine-rich region immediately upstream of TSS had appeared during the evolution of the “scanning model” species, which might facilitate TSS selection in these species. Both initiation mechanisms share a strong preference for pyrimidine-purine dinucleotides surrounding the TSS. Our results suggested that the purine is required for accurately recruiting the first nucleotide, increasing the chance of being capped during mRNA maturation, which is critical for efficient translation initiation. Based on our findings, we proposed a model of TSS selection for the “scanning model” species. Besides, our study also demonstrated that the intrinsic sequence feature primarily determines the distribution of initiation activities within a core promoter (core promoter shape).

DNA STRUCTURAL FEATURES AND ARCHITECTURE OF PROMOTER REGIONS PLAY A ROLE IN GENE RESPONSIVENESS OF S. cerevisiae

2013 ◽  
Vol 11 (06) ◽  
pp. 1343001 ◽  
Author(s):  
VENKATA RAJESH YELLA ◽  
MANJU BANSAL
Keyword(s):  
Gene Expression ◽  
Structural Properties ◽  
Transcription Initiation ◽  
Nucleosome Occupancy ◽  
Regulation Of Gene Expression ◽  
Structural Features ◽  
Tata Box ◽  
Promoter Regions ◽  
Expression Variability ◽  
Promoter Architecture

Gene expression is the most fundamental biological process, which is essential for phenotypic variation. It is regulated by various external (environment and evolution) and internal (genetic) factors. The level of gene expression depends on promoter architecture, along with other external factors. Presence of sequence motifs, such as transcription factor binding sites (TFBSs) and TATA-box, or DNA methylation in vertebrates has been implicated in the regulation of expression of some genes in eukaryotes, but a large number of genes lack these sequences. On the other hand, several experimental and computational studies have shown that promoter sequences possess some special structural properties, such as low stability, less bendability, low nucleosome occupancy, and more curvature, which are prevalent across all organisms. These structural features may play role in transcription initiation and regulation of gene expression. We have studied the relationship between the structural features of promoter DNA, promoter directionality and gene expression variability in S. cerevisiae. This relationship has been analyzed for seven different measures of gene expression variability, along with two different regulatory effect measures. We find that a few of the variability measures of gene expression are linked to DNA structural properties, nucleosome occupancy, TATA-box presence, and bidirectionality of promoter regions. Interestingly, gene responsiveness is most intimately correlated with DNA structural features and promoter architecture.

scholarly journals Promoter scanning during transcription initiation in Saccharomyces cerevisiae: Pol II in the “shooting gallery”

2019 ◽  
Author(s):  
Chenxi Qiu ◽  
Huiyan Jin ◽  
Irina Vvedenskaya ◽  
Jordi Abante Llenas ◽  
Tingting Zhao ◽  
...  
Keyword(s):  
Catalytic Activity ◽  
Transcription Initiation ◽  
Promoter Sequence ◽  
Preinitiation Complex ◽  
Pol Ii ◽  
Transcription Start Sites ◽  
Promoter Architecture ◽  
General Transcription Factors ◽  
Promoter Class ◽  
Shooting Gallery

ABSTRACTBackgroundThe majority of eukaryotic promoters utilize multiple transcription start sites (TSSs). How multiple TSSs are specified at individual promoters across eukaryotes is not understood for most species. In S. cerevisiae, a preinitiation complex comprised of Pol II and conserved general transcription factors (GTFs) assembles and opens DNA upstream of TSSs. Evidence from model promoters indicates that the preinitiation complex (PIC) scans from upstream to downstream to identify TSSs. Prior results suggest that TSS distributions at promoters where scanning occurs shift in a polar fashion upon alteration in Pol II catalytic activity or GTF function.ResultsTo determine extent of promoter scanning across promoter classes in S. cerevisiae, we perturbed Pol II catalytic activity and GTF function and analyzed their effects on TSS usage genome-wide. We find that alterations to Pol II, TFIIB, or TFIIF function widely alter the initiation landscape consistent with promoter scanning operating at all yeast promoters, regardless of promoter class. Promoter architecture, however, can determine extent of promoter sensitivity to altered Pol II activity in ways that are predicted by a scanning model.ConclusionsOur observations coupled with previous data validate key predictions of the scanning model for Pol II initiation in yeast – which we term the “shooting gallery”. In this model, Pol II catalytic activity, and the rate and processivity of Pol II scanning together with promoter sequence determine the distribution of TSSs and their usage.

scholarly journals Discovery of widespread transcription initiation at microsatellites predictable by sequence-based deep neural network

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Mathys Grapotte ◽  
Manu Saraswat ◽  
Chloé Bessière ◽  
Christophe Menichelli ◽  
Jordan A. Ramilowski ◽  
...  
Keyword(s):  
Transcription Initiation ◽  
Tandem Repeats ◽  
Specific Gene ◽  
Rna Seq ◽  
Transcription Start Sites ◽  
Long Read ◽  
Cap Analysis ◽  
Dna Tandem Repeats ◽  
Short Tandem ◽  
Str Polymorphism

AbstractUsing the Cap Analysis of Gene Expression (CAGE) technology, the FANTOM5 consortium provided one of the most comprehensive maps of transcription start sites (TSSs) in several species. Strikingly, ~72% of them could not be assigned to a specific gene and initiate at unconventional regions, outside promoters or enhancers. Here, we probe these unassigned TSSs and show that, in all species studied, a significant fraction of CAGE peaks initiate at microsatellites, also called short tandem repeats (STRs). To confirm this transcription, we develop Cap Trap RNA-seq, a technology which combines cap trapping and long read MinION sequencing. We train sequence-based deep learning models able to predict CAGE signal at STRs with high accuracy. These models unveil the importance of STR surrounding sequences not only to distinguish STR classes, but also to predict the level of transcription initiation. Importantly, genetic variants linked to human diseases are preferentially found at STRs with high transcription initiation level, supporting the biological and clinical relevance of transcription initiation at STRs. Together, our results extend the repertoire of non-coding transcription associated with DNA tandem repeats and complexify STR polymorphism.

Transcriptional and posttranscriptional regulation of maize mitochondrial gene expression

1991 ◽  
Vol 11 (1) ◽  
pp. 533-543
Author(s):  
R M Mulligan ◽  
P Leon ◽  
V Walbot
Keyword(s):  
Dna Sequences ◽  
Transcription Initiation ◽  
Posttranscriptional Regulation ◽  
Rank Order ◽  
Mitochondrial Gene ◽  
Initiation Site ◽  
Gene Copy ◽  
Promoter Strength ◽  
Protein Coding

Lysed maize mitochondria synthesize RNA in the presence of radioactive nucleoside triphosphates, and this assay was utilized to compare the rates of transcription of seven genes. The rates of incorporation varied over a 14-fold range, with the following rank order: 18S rRNA greater than 26S rRNA greater than atp1 greater than atp6 greater than atp9 greater than cob greater than cox3. The products of run-on transcription hybridized specifically to known transcribed regions and selectively to the antisense DNA strand; thus, the isolated run-on transcription system appears to be an accurate representation of endogenous transcription. Although there were small differences in gene copy abundance, these differences cannot account for the differences in apparent transcription rates; we conclude that promoter strength is the main determinant. Among the protein coding genes, incorporation was greatest for atp1. The most active transcription initiation site of this gene was characterized by hybridization with in vitro-capped RNA and by primer extension analyses. The DNA sequences at this and other transcription initiation sites that we have previously mapped were analyzed with respect to the apparent promoter strengths. We propose that two short sequence elements just upstream of initiation sites form at least a portion of the sequence requirements for a maize mitochondrial promoter. In addition to modulation at the level of transcription, steady-state abundance of protein-coding mRNAs varied over a 20-fold range and did not correlate with transcriptional activity. These observations suggest that posttranscriptional processes are important in the modulation of mRNA abundance.

Unraveling Toxoplasma gondii GT1 Strain Virulence and New Protein-Coding Genes with Proteogenomic Analyses

2021 ◽  
Author(s):  
Neelam Antil ◽  
Manish Kumar ◽  
Santosh Kumar Behera ◽  
Mohammad Arefian ◽  
Chinmaya Narayana Kotimoole ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Protein Coding ◽  
Protein Coding Genes ◽  
New Protein

The Long Road to Understanding RNAPII Transcription Initiation and Related Syndromes

2021 ◽  
Vol 90 (1) ◽  
pp. 193-219
Author(s):  
Emmanuel Compe ◽  
Jean-Marc Egly
Keyword(s):  
Transcription Factors ◽  
Rna Polymerase Ii ◽  
Chromatin Remodeling ◽  
Transcription Initiation ◽  
Rna Synthesis ◽  
Regulatory Elements ◽  
Initiation Mechanism ◽  
Protein Coding ◽  
Core Promoters ◽  
General Transcription Factors

In eukaryotes, transcription of protein-coding genes requires the assembly at core promoters of a large preinitiation machinery containing RNA polymerase II (RNAPII) and general transcription factors (GTFs). Transcription is potentiated by regulatory elements called enhancers, which are recognized by specific DNA-binding transcription factors that recruit cofactors and convey, following chromatin remodeling, the activating cues to the preinitiation complex. This review summarizes nearly five decades of work on transcription initiation by describing the sequential recruitment of diverse molecular players including the GTFs, the Mediator complex, and DNA repair factors that support RNAPII to enable RNA synthesis. The elucidation of the transcription initiation mechanism has greatly benefited from the study of altered transcription components associated with human diseases that could be considered transcription syndromes.

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