Simple sequence repeats in organellar genomes of rice: frequency and distribution in genic and intergenic regions

MOTIVATION: Simple sequence repeats (SSRs) are abundant across genomes. However, the significance of SSRs in organellar genomes of rice has not been completely understood. The availability of organellar genome sequences allows us to understand the organization of SSRs in their genic and intergenic regions. RESULTS: We have analyzed SSRs in mitochondrial and chloroplast genomes of rice. We identified 2528 SSRs in the mitochondrial genome and average 870 SSRs in the chloroplast genomes. About 8.7% of the mitochondrial and 27.5% of the chloroplast SSRs were observed in the genic region. Dinucleotides were the most abundant repeats in genic and intergenic regions of the mitochondrial genome while mononucleotides were predominant in the chloroplast genomes. The rps and nad gene clusters of mitochondria had the maximum repeats, while the rpo and ndh gene clusters of chloroplast had the maximum repeats. We identified SSRs in both organellar genomes and validated in different cultivars and species.     

Compact intergenic regions of the pufferfish genome facilitate isolation of gene promoters: characterization of Fugu 3'-phosphoadenosine 5'-phosphosulfate synthase 2 (fPapss2) gene promoter function in transgenic Xenopus

The highly compact nature of the pufferfish (Fugu rubripes) genome renders it a useful tool not only for annotating coding regions within vertebrate genomes, but also for the identification of sequences important to gene regulation. Indeed, owing to this compaction it will be feasible in many instances to initiate analyses using entire intergenic regions when mapping gene promoters; a strategy that is very rarely feasible with the expanded genomes of other species. Stemming from our interest in studying promoters expressed in chondrocytes, we selected for study the intergenic region upstream of Fugu 3'-phosphoadenosine 5'-phosphosulfate synthase 2, fPapss2, a gene required for the normal development of cartilage extracellular matrix. Functional characterization of the entire fPapss2 5' intergenic region was carried out by monitoring expression of the enhanced green fluorescent protein (EGFP) gene reporter in the developing cartilage of transgenic Xenopus laevis. By evaluating a series of 5' intergenic region deletions we defined a minimal fPapss2 sequence of approximately 300 bp that was essential for EGFP expression in tadpole cartilage. This functional analysis of an entire Fugu intergenic region, combined with the efficiency of Xenopus transgenesis, serves as a model for the rapid characterization of evolutionarily-conserved regulatory regions of other pufferfish genes.     

Intergenics: A tool for extraction of intergenicregions

For the past one decade, there has been considerable explosion of interest in searching novel regulatory elements in the intergenic region between the protein coding regions. The microbial genomes are the most exploited in terms of intergenic (noncoding) regions due to its less complexity. We think, the increasing pace of genome sequencing calls for a tool which will be useful for the extraction of intergenic regions. IntergenicS (Intergenic Sequence) is a tool which can extract the intergenic regions of microbial genomes at NCBI. All the unannotated regions between annotated protein coding genes and noncoding RNA genes can be extracted. It also deals with the calculation of GC base composition of the intergenic regions. This will be a useful tool for the analysis of noncoding regions of both bacterial and archael genomes.     

Microsatellite analysis in organelle genomes of Chlorophyta

Simple Sequence Repeats (SSRs) or microsatellites constitute a significant portion of genomes however; their significance in organellar genomes has not been completely understood. The availability of organelle genome sequences allows us to understand the organization of SSRs in their genic and intergenic regions. In the present work, SSRs were identified and categorized in 14 mitochondrial and 22 chloroplast genomes of algal species belonging to Chlorophyta. Based on the study, it was observed that number of SSRs in non-coding region were more as compared to coding region and frequency of mononucleotides repeats were highest followed by dinucleotides in both mitochondrial and chloroplast genomes. It was also observed that maximum number of SSRs was found in genes encoding for beta subunit of RNA polymerase in chloroplast genomes and NADH dehydrogenase in mitochondrial genomes. This is the first and original report on whole genomes sequence analysis of organellar genomes of green algae.     

Evolution of Conserved Non-Coding Sequences Within the Vertebrate Hox Clusters Through the Two-Round Whole Genome Duplications Revealed by Phylogenetic Footprinting Analysis

As a result of two-round whole genome duplications, four or more paralogous Hox clusters exist in vertebrate genomes. The paralogous genes in the Hox clusters show similar expression patterns, implying shared regulatory mechanisms for expression of these genes. Previous studies partly revealed the expression mechanisms of Hox genes. However, cis-regulatory elements that control these paralogous gene expression are still poorly understood. Toward solving this problem, the authors searched conserved non-coding sequences (CNSs), which are candidates of cis-regulatory elements. When comparing orthologous Hox clusters of 19 vertebrate species, 208 intergenic conserved regions were found. The authors then searched for CNSs that were conserved not only between orthologous clusters but also among the four paralogous Hox clusters. The authors found three regions that are conserved among all the four clusters and eight regions that are conserved between intergenic regions of two paralogous Hox clusters. In total, 28 CNSs were identified in the paralogous Hox clusters, and nine of them were newly found in this study. One of these novel regions bears a RARE motif. These CNSs are candidates for gene expression regulatory regions among paralogous Hox clusters. The authors also compared vertebrate CNSs with amphioxus CNSs within the Hox cluster, and found that two CNSs in the HoxA and HoxB clusters retain homology with amphioxus CNSs through the two-round whole genome duplications.     

Characterization of chloroplast DNA microsatellites from Saccharum spp and related species

Microsatellites, or simple sequence repeats (SSRs), and their flanking regions in chloroplast genomes (plastomes) of some species of the family Poaceae were analyzed in silico to look for DNA sequence variations. Comparison of the complete chloroplast DNA sequences (cpDNAs) of sugarcane (Saccharum hybrid cv. SP-80-3280 and S. officinarum cv. NCo310) and related species, Agrostis stolonifera, Brachypodium distachyon, Hordeum vulgare subsp vulgare, Lolium perenne, Oryza nivara, O. sativa subsp indica, O. sativa subsp japonica, Sorghum bicolor, Triticum aestivum, Zea mays, and Z. mays cv. B73, allowed us to examine the organization of chloroplast SSRs (cpSSRs) in genic and intergenic regions. We identified 204 cpSSRs in the sugarcane cpDNA; 22.5% were in genic regions. The ndh, rps, trn, and rpl gene clusters of the chloroplasts had the most repeats. Mononucleotide repeats were the most abundant cpSSRs in these species; however, di-, tri-, tetra-, penta-, and hexanucleotide repeats were also identified. Many base substitutions and deletions/insertions were identified in the cpSSR loci and their flanking regions. Multiple alignments of all cpSSR sequences of Poaceae species made identification of nucleotide variability possible; repeat motifs are not uniformly distributed across the Poaceae plastomes, but are mostly confined to intergenic regions. Phylogeny was determined by maximum parsimony and neighbor-joining inference methods. The cpSSRs of these species were found to be polymorphic. It appears that individual cpSSRs in the Poaceae are stable, at least over short periods of evolutionary time. We conclude that the plastome database can be exploited for phylogenetic analysis and biotechnological development.     

Selection on Accessible Chromatin Regions in Capsella grandiflora

Accurate estimates of genome-wide rates and fitness effects of new mutations are essential for an improved understanding of molecular evolutionary processes. Although eukaryotic genomes generally contain a large noncoding fraction, functional noncoding regions and fitness effects of mutations in such regions are still incompletely characterized. A promising approach to characterize functional noncoding regions relies on identifying accessible chromatin regions (ACRs) tightly associated with regulatory DNA. Here, we applied this approach to identify and estimate selection on ACRs in Capsella grandiflora, a crucifer species ideal for population genomic quantification of selection due to its favorable population demography. We describe a population-wide ACR distribution based on ATAC-seq data for leaf samples of 16 individuals from a natural population. We use population genomic methods to estimate fitness effects and proportions of positively selected fixations (α) in ACRs and find that intergenic ACRs harbor a considerable fraction of weakly deleterious new mutations, as well as a significantly higher proportion of strongly deleterious mutations than comparable inaccessible intergenic regions. ACRs are enriched for expression quantitative trait loci (eQTL) and depleted of transposable element insertions, as expected if intergenic ACRs are under selection because they harbor regulatory regions. By integrating empirical identification of intergenic ACRs with analyses of eQTL and population genomic analyses of selection, we demonstrate that intergenic regulatory regions are an important source of nearly neutral mutations. These results improve our understanding of selection on noncoding regions and the role of nearly neutral mutations for evolutionary processes in outcrossing Brassicaceae species.     

Analysis of similarity within 142 pairs of orthologous intergenic regions of Caenorhabditis elegans and Caenorhabditis briggsae

Patterns of similarity between genomes of related species reflect the distribution of selective constraint within DNA. We analyzed alignments of 142 orthologous intergenic regions of Caenorhabditis elegans and Caenorhabditis briggsae and found a mosaic pattern with regions of high similarity (phylogenetic footprints) interspersed with non-alignable sequences. Footprints cover ~20% of intergenic regions, often occur in clumps and are rare within 5′ UTRs but common within 3′ UTRs. The footprints have a higher ratio of transitions to transversions than expected at random and a higher GC content than the rest of the intergenic region. The number of footprints and the GC content of footprints within an intergenic region are higher when genes are oriented so that their 5′ ends form the boundaries of the intergenic region. Overall, the patterns and characteristics identified here, along with other comparative and experimental studies, suggest that many footprints have a regulatory function, although other types of function are also possible. These conclusions may be quite general across eukaryotes, and the characteristics of conserved regulatory elements determined from genomic comparisons can be useful in prediction of regulation sites within individual DNA sequences.     

CpG_MI: a novel approach for identifying functional CpG islands in mammalian genomes

CpG islands (CGIs) are CpG-rich regions compared to CpG-depleted bulk DNA of mammalian genomes and are generally regarded as the epigenetic regulatory regions in association with unmethylation, promoter activity and histone modifications. Accurate identification of CpG islands with epigenetic regulatory function in bulk genomes is of wide interest. Here, the common features of functional CGIs are identified using an average mutual information method to differentiate functional CGIs from the remaining CGIs. A new approach (CpG mutual information, CpG_MI) was further explored to identify functional CGIs based on the cumulative mutual information of physical distances between two neighboring CpGs. Compared to current approaches, CpG_MI achieved the highest prediction accuracy. This approach also identified new functional CGIs overlapping with gene promoter regions which were missed by other algorithms. Nearly all CGIs identified by CpG_MI overlapped with histone modification marks. CpG_MI could also be used to identify potential functional CGIs in other mammalian genomes, as the CpG dinucleotide contents and cumulative mutual information distributions are almost the same among six mammalian genomes in our analysis. It is a reliable quantitative tool for the identification of functional CGIs from bulk genomes and helps in understanding the relationships between genomic functional elements and epigenomic modifications.     

Evolutionary relationships between Saccharomyces cerevisiae and other fungal species as determined from genome comparisons

The increasing number of fungal genomes whose sequence has been completed permits their comparison both at the nucleotide and protein levels. The information thus obtained improves our knowledge on evolutionary relationships between fungi. Comparison of the Saccharomyces cerevisiae genome with other Hemiascomycetes genomes confirms that a whole-genome duplication occurred before the diversification between Candida glabrata and the Saccharomyces sensu stricto species and after separation from the branch leading to the other Hemiascomycetes. Duplication was followed by individual gene losses and rearrangements affecting extensive DNA regions. Although S. cerevisiae and C. glabrata are two closely related yeast species at an evolutionary scale, their different habitats and life styles correlate with specific gene differences and with more extensive gene loses having occurred in the parasitic C. glabrata. At a closer evolutive scale, diversification among the sensu stricto species began with nucleotide changes at the intergenic regions affecting sequences that are not relevant for gene regulation, together with more extensive genome rearrangements involving transposons and telomeric regions. One important characteristic of fungal genomes that is shared with other eukaryotes is the fusion of gene sequences coding for separate protein modules into a single open reading frame. This allows diversification of protein functions while saving gene information.     

Decoding cis-regulatory systems in ascidians

Ascidians, or sea squirts, are lower chordates, and share basic gene repertoires and many characteristics, both developmental and physiological, with vertebrates. Therefore, decoding cis-regulatory systems in ascidians will contribute toward elucidating the genetic regulatory systems underlying the developmental and physiological processes of vertebrates. cis-Regulatory DNAs can also be used for tissue-specific genetic manipulation, a powerful tool for studying ascidian development and physiology. Because the ascidian genome is compact compared with vertebrate genomes, both intergenic regions and introns are relatively small in ascidians. Short upstream intergenic regions contain a complete set of cis-regulatory elements for spatially regulated expression of a majority of ascidian genes. These features of the ascidian genome are a great advantage in identifying cis-regulatory sequences and in analyzing their functions. Function of cis-regulatory DNAs has been analyzed for a number of tissue-specific and developmentally regulated genes of ascidians by introducing promoter-reporter fusion constructs into ascidian embryos. The availability of the whole genome sequences of the two Ciona species, Ciona intestinalis and Ciona savignyi, facilitates comparative genomics approaches to identify cis-regulatory DNAs. Recent studies demonstrate that computational methods can help identify cis-regulatory elements in the ascidian genome. This review presents a comprehensive list of ascidian genes whose cis-regulatory regions have been subjected to functional analysis, and highlights the recent advances in bioinformatics and comparative genomics approaches to cis-regulatory systems in ascidians.     

IntergenicDB: a database for intergenic sequences

A whole genome contains not only coding regions, but also non-coding regions. These are located between the end of a given coding region and the beginning of the following coding region. For this reason, the information about gene regulation process underlies in intergenic regions. There is no easy way to obtain intergenic regions from current available databases. IntergenicDB was developed to integrate data of intergenic regions and their gene related information from NCBI databases. The main goal of INTERGENICDB is to offer friendly database for intergenic sequences of bacterial genomes.

Availability

http://intergenicdb.bioinfoucs.com/     

MRD: a microsatellite repeats database for prokaryotic and eukaryotic genomes

MRD is a database system to access the microsatellite repeats information of genomes such as archea, eubacteria, and other eukaryotic genomes whose sequence information is available in public domains. MRD stores information about simple tandemly repeated k-mer sequences where k= 1 to 6, i.e. monomer to hexamer. The web interface allows the users to search for the repeat of their interest and to know about the association of the repeat with genes and genomic regions in the specific organism. The data contains the abundance and distribution of microsatellites in the coding and non-coding regions of the genome. The exact location of repeats with respect to genomic regions of interest (such as UTR, exon, intron or intergenic regions) whichever is applicable to organism is highlighted. MRD is available on the World Wide Web at and/or . The database is designed as an open-ended system to accommodate the microsatellite repeats information of other genomes whose complete sequences will be available in future through public domain.