scholarly journals GC content but not nucleosome positioning directly contributes to intron-splicing efficiency in Paramecium

2021 ◽  
Author(s):  
Stefano Gnan ◽  
Melody Matelot ◽  
Marion Weiman ◽  
Olivier Arnaiz ◽  
Frederic Guerin ◽  
...  
Keyword(s):  
Nucleosome Occupancy ◽  
Average Length ◽  
Gc Content ◽  
Nucleosome Positioning ◽  
Evolutionary Constraint ◽  
Paramecium Tetraurelia ◽  
Intron Splicing ◽  
Nonsense Mediated Decay ◽  
Eukaryotic Genes ◽  
Splicing Efficiency

Eukaryotic genes are interrupted by introns that must be accurately spliced from mRNA precursors. With an average length of 25 nt, the >90,000 introns of Paramecium tetraurelia stand among the shortest introns reported in eukaryotes. The mechanisms specifying the correct recognition of these tiny introns remain poorly understood. Splicing can occur co-transcriptionally and it has been proposed that chromatin structure might influence splice site recognition. To investigate the roles of nucleosome positioning in intron recognition, we determined the nucleosome occupancy along the P. tetraurelia genome. We showed that P. tetraurelia displays a regular nucleosome array with a nucleosome repeat length of ~151 bp, amongst the smallest periodicities reported. Our analysis revealed that introns are frequently associated with inter-nucleosomal DNA, pointing to an evolutionary constraint to locate introns at the AT-rich nucleosome edge sequences. Using accurate splicing efficiency data from cells depleted for the nonsense-mediated decay effectors, we showed that introns located at the edge of nucleosomes display higher splicing efficiency than those at the centre. However, multiple regression analysis indicated that the GC content, rather than nucleosome positioning, directly contributes to intron splicing efficiency. Our data reveal a complex link between GC content, nucleosome positioning and intron evolution in Paramecium.

scholarly journals A deformation energy model reveals sequence-dependent property of nucleosome positioning

Chromosoma ◽  
2021 ◽  
Vol 130 (1) ◽  
pp. 27-40
Author(s):  
Guoqing Liu ◽  
Hongyu Zhao ◽  
Hu Meng ◽  
Yongqiang Xing ◽  
Lu Cai
Keyword(s):  
Budding Yeast ◽  
Nucleosome Occupancy ◽  
Nucleosome Positioning ◽  
Deformation Energy ◽  
Energy Model ◽  
Structural Parameter ◽  
High Deformation ◽  
Dna Deformation ◽  
Genome Wide ◽  
Genomic Regions

AbstractWe present a deformation energy model for predicting nucleosome positioning, in which a position-dependent structural parameter set derived from crystal structures of nucleosomes was used to calculate the DNA deformation energy. The model is successful in predicting nucleosome occupancy genome-wide in budding yeast, nucleosome free energy, and rotational positioning of nucleosomes. Our model also indicates that the genomic regions underlying the MNase-sensitive nucleosomes in budding yeast have high deformation energy and, consequently, low nucleosome-forming ability, while the MNase-sensitive non-histone particles are characterized by much lower DNA deformation energy and high nucleosome preference. In addition, we also revealed that remodelers, SNF2 and RSC8, are likely to act in chromatin remodeling by binding to broad nucleosome-depleted regions that are intrinsically favorable for nucleosome positioning. Our data support the important role of position-dependent physical properties of DNA in nucleosome positioning.

scholarly journals Transcriptome analysis of two contrasting genotypes of pearl millet to gain insight into heat stress responses

2020 ◽  
Author(s):  
Albert Maibam ◽  
Sunil Nigombam ◽  
Harinder Vishwakarma ◽  
Showkat Ahmad Lone ◽  
Kishor Gaikwad ◽  
...  
Keyword(s):  
Heat Stress ◽  
Pearl Millet ◽  
Stress Responses ◽  
Simple Sequence Repeats ◽  
Pennisetum Glaucum ◽  
Average Length ◽  
Gc Content ◽  
Functional Markers ◽  
Sequencing Data ◽  
Simple Sequence

Abstract Background Pennisetum glaucum (L.) R. Br. is mainly grown in arid and semi-arid regions. Being naturally tolerant to various adverse condtitions, it is a good biological resource for deciphering the molecular basis of abiotic stresses such as heat stress in plants but limited studies have been carried out till date to this effect. Here, we performed RNA-sequencing from the leaf of two contrasting genotypes of pearl millet (841-B and PPMI-69) subjected to heat stress (42 °C for 6 h). Results Over 274 million high quality reads with an average length of 150 nt were generated. Assembly was carried out using trinity, obtaining 47,310 unigenes having an average length of 1254 nucleotides, N50 length of 1853 nucleotides and GC content of 53.11%. Blastx resulted in annotation of 35,628 unigenes and functional classification showed 15,950 unigenes designated to 51 Gene Ontology terms, 13,786 unigenes allocated to 23 Clusters of Orthologous Groups and 4,255 unigenes distributed into 132 functional KEGG pathways. 12,976 simple sequence repeats were identified from 10,294 unigenes for the development of functional markers. A total of 3,05,759 SNPs were observed in the transcriptome data. Out of 2,301 differentially expressed genes, 10 potential candidates genes were selected based on log2 fold change and adjusted p-value parameters for their differential gene expression by qRT-PCR. Conclusions The dynamic expression changes in two genotypes of P. glaucum reflect transcriptome regulation of signaling pathways in heat stress response. In order to develop genetic markers, 12,976 simple sequence repeats (SSRs) were identified. The sequencing data generated in this study shall serve as an important resource for further research in the area of crop biotechnology.

scholarly journals Actively translated uORFs reduce translation and mRNA stability independent of NMD in Arabidopsis

2021 ◽  
Author(s):  
Hsin-Yen Larry Wu ◽  
Polly Yingshan Hsu
Keyword(s):  
Gene Expression ◽  
Mrna Stability ◽  
Gene Expression Regulation ◽  
Regulatory Elements ◽  
Expression Regulation ◽  
Ribosome Profiling ◽  
Translation Efficiency ◽  
Nonsense Mediated Decay ◽  
Protein Levels ◽  
Eukaryotic Genes

ABSTRACTUpstream ORFs (uORFs) are widespread cis-regulatory elements in the 5’ untranslated regions of eukaryotic genes. Translation of uORFs could negatively regulate protein synthesis by repressing main ORF (mORF) translation and by reducing mRNA stability presumably through nonsense-mediated decay (NMD). While the above expectations were supported in animals, they have not been extensively tested in plants. Using ribosome profiling, we systematically identified 2093 Actively Translated uORFs (ATuORFs) in Arabidopsis seedlings and examined their roles in gene expression regulation by integrating multiple genome-wide datasets. Compared with genes without uORFs, we found ATuORFs result in 38%, 14%, and 43% reductions in translation efficiency, mRNA stability, and protein levels, respectively. The effects of predicted but not actively translated uORFs are much weaker than those of ATuORFs. Interestingly, ATuORF-containing genes are also expressed at higher levels and encode longer proteins with conserved domains, features that are common in evolutionarily older genes. Moreover, we provide evidence that uORF translation in plants, unlike in vertebrates, generally does not trigger NMD. We found ATuORF-containing transcripts are degraded through 5’ to 3’ decay, while NMD targets are degraded through both 5’ to 3’ and 3’ to 5’ decay, suggesting uORF-associated mRNA decay and NMD have distinct genetic requirements. Furthermore, we showed ATuORFs and NMD repress translation through separate mechanisms. Our results reveal that the potent inhibition of uORFs on mORF translation and mRNA stability in plants are independent of NMD, highlighting a fundamental difference in gene expression regulation by uORFs in the plant and animal kingdoms.

scholarly journals Exploration of the Germline Genome of the Ciliate Chilodonella uncinata through Single-Cell Omics (Transcriptomics and Genomics)

mBio ◽  
2018 ◽  
Vol 9 (1) ◽  
Author(s):  
Xyrus X. Maurer-Alcalá ◽  
Rob Knight ◽  
Laura A. Katz
Keyword(s):  
Single Cell ◽  
Large Scale ◽  
Gc Content ◽  
Gene Families ◽  
Genome Rearrangements ◽  
Paramecium Tetraurelia ◽  
Protein Coding ◽  
Genome Data ◽  
Large Gene ◽  
Disproportionate Number

ABSTRACTSeparate germline and somatic genomes are found in numerous lineages across the eukaryotic tree of life, often separated into distinct tissues (e.g., in plants, animals, and fungi) or distinct nuclei sharing a common cytoplasm (e.g., in ciliates and some foraminifera). In ciliates, germline-limited (i.e., micronuclear-specific) DNA is eliminated during the development of a new somatic (i.e., macronuclear) genome in a process that is tightly linked to large-scale genome rearrangements, such as deletions and reordering of protein-coding sequences. Most studies of germline genome architecture in ciliates have focused on the model ciliatesOxytricha trifallax,Paramecium tetraurelia, andTetrahymena thermophila, for which the complete germline genome sequences are known. Outside of these model taxa, only a few dozen germline loci have been characterized from a limited number of cultivable species, which is likely due to difficulties in obtaining sufficient quantities of “purified” germline DNA in these taxa. Combining single-cell transcriptomics and genomics, we have overcome these limitations and provide the first insights into the structure of the germline genome of the ciliateChilodonella uncinata, a member of the understudied classPhyllopharyngea. Our analyses reveal the following: (i) large gene families contain a disproportionate number of genes from scrambled germline loci; (ii) germline-soma boundaries in the germline genome are demarcated by substantial shifts in GC content; (iii) single-cell omics techniques provide large-scale quality germline genome data with limited effort, at least for ciliates with extensively fragmented somatic genomes. Our approach provides an efficient means to understand better the evolution of genome rearrangements between germline and soma in ciliates.IMPORTANCEOur understanding of the distinctions between germline and somatic genomes in ciliates has largely relied on studies of a few model genera (e.g.,Oxytricha,Paramecium,Tetrahymena). We have used single-cell omics to explore germline-soma distinctions in the ciliateChilodonella uncinata, which likely diverged from the better-studied ciliates ~700 million years ago. The analyses presented here indicate that developmentally regulated genome rearrangements between germline and soma are demarcated by rapid transitions in local GC composition and lead to diversification of protein families. The approaches used here provide the basis for future work aimed at discerning the evolutionary impacts of germline-soma distinctions among diverse ciliates.

scholarly journals Differential RNA Editing and Intron Splicing in Soybean Mitochondria during Nodulation

2020 ◽  
Vol 21 (24) ◽  
pp. 9378
Author(s):  
Yuzhe Sun ◽  
Min Xie ◽  
Zhou Xu ◽  
Koon Chuen Chan ◽  
Jia Yi Zhong ◽  
...  
Keyword(s):  
Rna Editing ◽  
Mitochondrial Genes ◽  
Protein Abundance ◽  
Intron Splicing ◽  
Rna Seq ◽  
Intron 1 ◽  
Tremendous Amount ◽  
Mitochondrial Transcripts ◽  
Splicing Efficiency ◽  
Transcript Profiles

Nitrogen fixation in soybean consumes a tremendous amount of energy, leading to substantial differences in energy metabolism and mitochondrial activities between nodules and uninoculated roots. While C-to-U RNA editing and intron splicing of mitochondrial transcripts are common in plant species, their roles in relation to nodule functions are still elusive. In this study, we performed RNA-seq to compare transcript profiles and RNA editing of mitochondrial genes in soybean nodules and roots. A total of 631 RNA editing sites were identified on mitochondrial transcripts, with 12% or 74 sites differentially edited among the transcripts isolated from nodules, stripped roots, and uninoculated roots. Eight out of these 74 differentially edited sites are located on the matR transcript, of which the degrees of RNA editing were the highest in the nodule sample. The degree of mitochondrial intron splicing was also examined. The splicing efficiencies of several introns in nodules and stripped roots were higher than in uninoculated roots. These include nad1 introns 2/3/4, nad4 intron 3, nad5 introns 2/3, cox2 intron 1, and ccmFc intron 1. A greater splicing efficiency of nad4 intron 1, a higher NAD4 protein abundance, and a reduction in supercomplex I + III2 were also observed in nodules, although the causal relationship between these observations requires further investigation.

scholarly journals The genome and microbiome of a dikaryotic fungus (Inocybe terrigena, Inocybaceae) revealed by metagenomics

2017 ◽  
Author(s):  
Mohammad Bahram ◽  
Dan Vanderpool ◽  
Mari Pent ◽  
Markus Hiltunen ◽  
Martin Ryberg
Keyword(s):  
Fruiting Body ◽  
Gc Content ◽  
White Rot ◽  
Fungal Genome ◽  
Metagenomic Sequencing ◽  
Ectomycorrhizal Symbiosis ◽  
High Coverage ◽  
Symbiotic Associations ◽  
Body Tissues ◽  
Eukaryotic Genes

Although recent advances in molecular methods have facilitated understanding the evolution of fungal symbiosis, little is known about genomic and microbiome features of most uncultured symbiotic fungal clades. Here, we analysed the genome and microbiome of Inocybaceae (Basidiomycota), a largely uncultured ectomycorrhizal clade that form symbiotic associations with a wide variety of plant species. Using metagenomic sequencing and assembly of dikaryotic fruiting-body tissues from Inocybe terrigena (Fr.) Kuyper, followed by classifying contigs into fungi and bacteria based on BLAST alignments as well as their differential coverage and GC content, we obtained a highly complete fungal genome, containing 93% of core eukaryotic genes. I. terrigena genome was more related to previously published ectmycorrhizal and brown rot than white rot genomes; however, it showed a significant reduction in lignin degradation capacity compared to closely related ectomycorrhizal clades, supporting independent evolution of ectomycorrhizal symbiosis in Inocybe. The microbiome of I. terrigena harboured bacteria with relatively high-coverage assemblies as well as with known symbiotic functions in hypogeous fungal tissues, suggesting the symbiotic functions of these bacteria in fungal tissues independent of habitat conditions. Our study demonstrates the usefulness of direct metagenomics analysis of fruiting-body tissues for characterizing fungal genomes and microbiome.

scholarly journals Distinct nucleosome distribution patterns in two structurally and functionally differentiated nuclei of a unicellular eukaryote

2015 ◽  
Author(s):  
Jie Xiong ◽  
Shan Gao ◽  
Wen Dui ◽  
Wentao Yang ◽  
Xiao Chen ◽  
...  
Keyword(s):  
Rna Polymerase Ii ◽  
Dna Sequences ◽  
Germ Line ◽  
Nucleosome Occupancy ◽  
Gc Content ◽  
Distribution Patterns ◽  
Unicellular Eukaryote ◽  
Transcription Start Sites ◽  
Highly Correlated ◽  
Cis And Trans

The ciliate protozoan Tetrahymena thermophila contains two types of structurally and functionally differentiated nuclei: the transcriptionally active somatic macronucleus (MAC) and the transcriptionally silent germ-line micronucleus (MIC). Here we demonstrate that MAC features well-positioned nucleosomes downstream of transcription start sites (TSS) likely connected with promoter proximal pausing of RNA polymerase II, as well as in exonic regions flanking both the 5′ and 3′ splice sites. In contrast, nucleosomes in MIC are more delocalized. Nucleosome occupancy in MAC and MIC are nonetheless highly correlated with each other and with predictions based upon DNA sequence features. Arrays of well-positioned nucleosomes are often correlated with GC content oscillations, suggesting significant contributions from cis-determinants. We propose that cis- and trans-determinants may coordinately accommodate some well-positioned nucleosomes with important functions, driven by a process in which positioned nucleosomes shape the mutational landscape of associated DNA sequences, while the DNA sequences in turn reinforce nucleosome positioning.

scholarly journals Nucleosome positioning on large tandem DNA repeats of the '601' sequence engineered in Saccharomyces cerevisiae

2021 ◽  
Author(s):  
Astrid Lancrey ◽  
Alexandra Joubert ◽  
Evelyne Duvernois-Berthet ◽  
Etienne Routhier ◽  
Saurabh Raj ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Sequence ◽  
Dna Sequences ◽  
Nucleosome Occupancy ◽  
Nucleosome Positioning ◽  
Dna Repeats ◽  
Repeat Array ◽  
Synthetic Dna

The so-called 601 DNA sequence is often used to constrain the position of nucleosomes on a DNA molecule in vitro. Although the ability of the 147 base pair sequence to precisely position a nucleosome in vitro is well documented, in vivo application of this property has been explored only in a few studies and yielded contradictory conclusions. Our goal in the present study was to test the ability of the 601 sequence to dictate nucleosome positioning in Saccharomyces cerevisiae in the context of a long tandem repeat array inserted in a yeast chromosome. We engineered such arrays with three different repeat size, namely 167, 197 and 237 base pairs. Although our arrays are able to position nucleosomes in vitro as expected, analysis of nucleosome occupancy on these arrays in vivo revealed that nucleosomes are not preferentially positioned as expected on the 601-core sequence along the repeats and that the measured nucleosome repeat length does not correspond to the one expected by design. Altogether our results demonstrate that the rules defining nucleosome positions on this DNA sequence in vitro are not valid in vivo, at least in this chromosomal context, questioning the relevance of using the 601 sequence in vivo to achieve precise nucleosome positioning on designer synthetic DNA sequences.

scholarly journals Genetic, parental and lifestyle factors influence telomere length

2021 ◽  
Author(s):  
Sergio Andreu-Sanchez ◽  
Geraldine Aubert ◽  
Aida Ripoll-Cladellas ◽  
Sandra Henkelman ◽  
Daria V. Zhernakova ◽  
...  
Keyword(s):  
Gene Expression ◽  
Average Length ◽  
Cell Types ◽  
Paternal Age ◽  
Specific Gene ◽  
Cell Type ◽  
Sequencing Data ◽  
Nonsense Mediated Decay ◽  
Cell Type Specific ◽  
Telomere Biology

The average length of telomere repeats (TL) declines with age and is considered to be a marker of biological ageing. Here, we measured TL in six blood cell types from 1,046 individuals using the clinically validated Flow-FISH method. We identified remarkable cell-type-specific variations in TL. Host genetics, environmental, parental and intrinsic factors such as sex, parental age, and smoking are associated to variations in TL. By analysing the genome-wide methylation patterns, we identified that the association of maternal, but not paternal, age to TL is mediated by epigenetics. Coupling these measurements to single-cell RNA-sequencing data for 62 participants revealed differential gene expression in T-cells. Genes negatively associated with TL were enriched for pathways related to translation and nonsense-mediated decay. Altogether, this study addresses cell-type-specific differences in telomere biology and its relation to cell-type-specific gene expression and highlights how perinatal factors play a role in determining TL, on top of genetics and lifestyle.

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