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Abstract Background The emergence of vertebrates is characterized by a strong increase in miRNA families. MicroRNAs interact broadly with many transcripts, and the evolution of such a system is intriguing. However, evolutionary questions concerning the origin of miRNA genes and their subsequent evolution remain unexplained. Results In order to systematically understand the evolutionary relationship between miRNAs gene and their function, we classified human known miRNAs into eight groups based on their evolutionary ages estimated by maximum parsimony method. New miRNA genes with new functional sequences accumulated more dynamically in vertebrates than that observed in Drosophila. Different levels of evolutionary selection were observed over miRNA gene sequences with different time of origin. Most genic miRNAs differ from their host genes in time of origin, there is no particular relationship between the age of a miRNA and the age of its host genes, genic miRNAs are mostly younger than the corresponding host genes. MicroRNAs originated over different time-scales are often predicted/verified to target the same or overlapping sets of genes, opening the possibility of substantial functional redundancy among miRNAs of different ages. Higher degree of tissue specificity and lower expression level was found in young miRNAs. Conclusions Our data showed that compared with protein coding genes, miRNA genes are more dynamic in terms of emergence and decay. Evolution patterns are quite different between miRNAs of different ages. MicroRNAs activity is under tight control with well-regulated expression increased and targeting decreased over time. Our work calls attention to the study of miRNA activity with a consideration of their origin time.

Abstract Background A number of vertebrate highly conserved elements (HCEs) have been detected and their genomic interval distances have been reported to be more conserved than protein coding genes among mammalian genomes. A characteristic of the human – non-mammalian comparisons is a bimodal distribution of relative distance difference of conserved consecutive HCE pairs; and it is difficult to attribute such profile to a random assortment. We therefore undertook an analysis of the human genomic regions confined by consecutive HCE pairs common to eight genomes (human, mouse, rat, chicken, frog, zebrafish, tetradon and fugu). Results Among HCE pairs, we found that some consistently preserve highly conserved interval distance among genomes while others have relatively low distance conservation. Using a partition method, we detected two groups of inter-HCE regions (IHRs) with distinct distance conservation pattern in vertebrate genomes: IHR1s that are bordered by HCE pairs with relative small distance variation, and IHR2s with larger distance difference values. Compared to random background, annotated repeat sequences are significantly less frequent in IHR1s than IHR2s, which reflects a correlation between repeat sequences and the length expansion of IHRs. Both groups of IHRs are unexpectedly enriched in human indel (i.e. insertion and deletion) polymorphism-variations than random background. The correlation between the percentage of conserved sequence and human IHR length was stronger for IHR1 than IHR2. Both groups of IHRs are significantly enriched for CpG islands. Conclusion The data suggest that subsets of HCE pairs may undergo different evolutionary paths in light of their genomic distance conservation, and that sets of genomic regions pertain to HCEs, as well as the region in which HCEs reside, should be treated as integrated domains.

Abstract Background A major goal of post-genomics research is the integrated analysis of genes, regulatory elements and the chromatin architecture on a genome-wide scale. Mapping DNase I hypersensitive sites within the nuclear chromatin is a powerful and well-established method of identifying regulatory element candidates. Results Here, we report the first genome-wide analysis of DNase I hypersensitive sites (DHSs) in Caenorhabditis elegans. The data was obtained by hybridizing DNase I-treated and end-captured material from young adult worms to a high-resolution tiling microarray. The data show that C. elegans DHSs were significantly enriched within intergenic regions located 2 kb upstream and downstream of coding genes, and also that a considerable fraction of all DHSs mapped to intergenic positions distant to annotated coding genes. Annotated transcribed loci were generally depleted in DHSs relative to intergenic regions, but DHSs were nonetheless enriched in coding exons and UTRs, whereas introns were significantly depleted in DHSs. Many DHSs appeared to be associated with annotated non-coding RNAs and recently detected transcripts of unknown function. It has been reported that nematode highly conserved non-coding elements were associated with cis-regulatory elements, and we also found that DHSs, particularly distal intergenic DHSs, were significantly enriched in regions that were conserved between the C. elegans and C. briggsae genomes. Conclusion We describe the first genome-wide analysis of C. elegans DHSs, and show that the distribution of DHSs is strongly associated with functional elements in the genome.

Abstract Background Short non-coding RNAs (ncRNAs) perform their cellular functions in ribonucleoprotein (RNP) complexes, which are also essential for maintaining the stability of the ncRNAs. Depletion of individual protein components of non-coding ribonucleoprotein (ncRNP) particles by RNA interference (RNAi) may therefore affect expression levels of the corresponding ncRNA, and depletion of candidate associated proteins may constitute an alternative strategy when investigating ncRNA-protein interactions and ncRNA functions. Therefore, we carried out a pilot study in which the effects of RNAi against protein components of small nucleolar RNPs (snoRNPs) in Caenorhabditis elegans were observed on an ncRNA microarray. Results RNAi against individual C. elegans protein components of snoRNPs produced strongly reduced mRNA levels and distinct phenotypes for all targeted proteins. For each type of snoRNP, individual depletion of at least three of the four protein components produced significant (P ≦ 1.2 × 10-5) reductions in the expression levels of the corresponding small nucleolar RNAs (snoRNAs), whereas the expression levels of other ncRNAs were largely unaffected. The effects of depletion of individual proteins were in accordance with snoRNP structure analyses obtained in other species for all but two of the eight targeted proteins. Variations in snoRNA size, sequence and secondary structure characteristics were not systematically reflected in the affinity for individual protein component of snoRNPs. The data supported the classification of nearly all annotated snoRNAs and suggested the presence of several novel snoRNAs among unclassified short ncRNA transcripts. A number of transcripts containing canonical Sm binding element sequences (Sm Y RNAs) also showed reduced expression after depletion of protein components of C/D box snoRNPs, whereas the expression of some stem-bulge RNAs (sbRNAs) was increased after depletion of the same proteins. Conclusion The study confirms observations made for other organisms, where reduced ncRNA levels after depletion of protein components of ncRNPs were noted, and shows that such reductions in expression levels occur across entire sets of ncRNA. Thereby, the study also demonstrates the feasibility of combining RNAi against candidate proteins with ncRNA microarray analysis to investigate ncRNA-protein interactions and hence ncRNA cellular functions.

Abstract Background Recent analysis of the mouse transcriptional data has revealed the existence of ~34,000 messenger-like non-coding RNAs (ml-ncRNAs). Whereas the functional properties of these ml-ncRNAs are beginning to be unravelled, no functional information is available for the large majority of these transcripts. Results A few ml-ncRNA have been shown to have genomic loci that overlap with microRNA loci, leading us to suspect that a fraction of ml-ncRNA may encode microRNAs. We therefore developed an algorithm (PriMir) for specifically detecting potential microRNA-encoding transcripts in the entire set of 34,030 mouse full-length ml-ncRNAs. In combination with mouse-rat sequence conservation, this algorithm detected 97 (80 of them were novel) strong miRNA-encoding candidates, and for 52 of these we obtained experimental evidence for the existence of their corresponding mature microRNA by microarray and stem-loop RT-PCR. Sequence analysis of the microRNA-encoding RNAs revealed an internal motif, whose presence correlates strongly (R2 = 0.9, P-value = 2.2 × 10-16) with the occurrence of stem-loops with characteristics of known pre-miRNAs, indicating the presence of a larger number microRNA-encoding RNAs (from 300 up to 800) in the ml-ncRNAs population. Conclusion Our work highlights a unique group of ml-ncRNAs and offers clues to their functions.

Abstract Background As phenotypic features derived from heritable characters, the topologies of metabolic pathways contain both phylogenetic and phenetic components. In the post-genomic era, it is possible to measure the "phylophenetic" contents of different pathways topologies from a global perspective. Results We reconstructed phylophenetic trees for all available metabolic pathways based on topological similarities, and compared them to the corresponding 16S rRNA-based trees. Similarity values for each pair of trees ranged from 0.044 to 0.297. Using the quartet method, single pathways trees were merged into a comprehensive tree containing information from a large part of the entire metabolic networks. This tree showed considerably higher similarity (0.386) to the corresponding 16S rRNA-based tree than any tree based on a single pathway, but was, on the other hand, sufficiently distinct to preserve unique phylogenetic information not reflected by the 16S rRNA tree. Conclusion We observed that the topology of different metabolic pathways provided different phylogenetic and phenetic information, depicting the compromise between phylogenetic information and varying evolutionary pressures forming metabolic pathway topologies in different organisms. The phylogenetic information content of the comprehensive tree is substantially higher than that of any tree based on a single pathway, which also gave clues to constraints working on the topology of the global metabolic networks, information that is only partly reflected by the topologies of individual metabolic pathways.

Motivation: PIWI-interacting RNAs (piRNAs) are a class of small non-coding RNAs that are highly abundant in the germline. One important role of piRNAs is to defend genome integrity by guiding PIWI proteins to silence transposable elements (TEs), which have a high potential to cause deleterious effects on their host. The mechanism of piRNA-mediated post-transcriptional silencing was also observed to affect mRNAs, suggesting that piRNAs might play a broad role in gene expression regulation. However, there has been no systematic report with regard to how many protein-coding genes might be targeted and regulated by piRNAs. Results: We trained a support vector machine classifier based on a combination of Miwi CLIP-Seq-derived features and position-derived features to predict the potential targets of piRNAs on mRNAs in the mouse. Reanalysis of a published microarray dataset suggested that the expression level of the 2587 protein-coding genes predicted as piRNA targets showed significant upregulation as a whole after abolishing the slicer activity of Miwi, supporting the conclusion that they are subject to piRNA-mediated regulation. Availability and implementation: A web version of the method called pirnaPre as well as our results for browse is available at http://www.regulatoryrna.org/software/piRNA/piRNA_target_mRNA/index.php. [ABSTRACT FROM AUTHOR]

piRNAs are a class of small RNAs that is most abundantly expressed in the animal germ line. Presently, substantial research is going on to reveal the functions of piRNAs in the epigenetic and post-transcriptional regulation of transposons and genes. A piRNA database for collection, annotation and structuring of these data will be a valuable contribution to the field, and we have therefore developed the piRBase platform which integrates various piRNA-related high-throughput data. piRBase has the largest collection of piRNAs among existing databases, and contains at present 77 million piRNA sequences from nine organisms. Repeat-derived and gene-derived piRNAs, which possibly participate in the regulation of the corresponding elements, have been given particular attention. Furthermore, epigenetic data and reported piRNA targets were also collected. To our knowledge, this is the first piRNA database that systematically integrates epigenetic and post-transcriptional regulation data to support piRNA functional analysis. We believe that piRBase will contribute to a better understanding of the piRNA functions. [ABSTRACT FROM AUTHOR]

Introduction. Small noncoding RNAs have important regulatory functions in different cell pathways. It is believed that most of them mainly play role in gene post-transcriptional regulation in the cytoplasm. Recent evidence suggests miRNA and siRNA activity in the nucleus. Here, we show distinct genome-wide sub-cellular localization distribution profiles of small noncoding RNAs in human breast cancer cells. Methods. We separated breast cancer cell nuclei from cytoplasm, and identified small RNA sequences using a high-throughput sequencing platform. To determine the relationship betweenmiRNA sub-cellular distribution and cancer progression, we used microarray analysis to examine the miRNA expression levels in nucleus and cytoplasm of three human cell lines, one normal breast cell line and two breast cancer cell lines. Logistic regression and SVMwere used for further analysis. Results. The sub-cellular distribution of small noncoding RNAs shows that numerousmiRNAs and their isoforms (isomiR) not only locate to the cytoplasm but also appeare in the nucleus. Subsequent microarray analyses indicated that the miRNA nuclear-cytoplasmicratio is a significant characteristic of different cancer cell lines. Conclusions. Our results indicate that the sub-cellular distribution is important for miRNA function, and that the characterization of the small RNAs sub-cellular localizome may contribute to cancer research and diagnosis. [ABSTRACT FROM AUTHOR]

Background: A number of vertebrate highly conserved elements (HCEs) have been detected and their genomic interval distances have been reported to be more conserved than protein coding genes among mammalian genomes. A characteristic of the human - non-mammalian comparisons is a bimodal distribution of relative distance difference of conserved consecutive HCE pairs; and it is difficult to attribute such profile to a random assortment. We therefore undertook an analysis of the human genomic regions confined by consecutive HCE pairs common to eight genomes (human, mouse, rat, chicken, frog, zebrafish, tetradon and fugu). Results: Among HCE pairs, we found that some consistently preserve highly conserved interval distance among genomes while others have relatively low distance conservation. Using a partition method, we detected two groups of inter-HCE regions (IHRs) with distinct distance conservation pattern in vertebrate genomes: IHR1s that are bordered by HCE pairs with relative small distance variation, and IHR2s with larger distance difference values. Compared to random background, annotated repeat sequences are significantly less frequent in IHR1s than IHR2s, which reflects a correlation between repeat sequences and the length expansion of IHRs. Both groups of IHRs are unexpectedly enriched in human indel (i.e. insertion and deletion) polymorphism-variations than random background. The correlation between the percentage of conserved sequence and human IHR length was stronger for IHR1 than IHR2. Both groups of IHRs are significantly enriched for CpG islands. Conclusion: The data suggest that subsets of HCE pairs may undergo different evolutionary paths in light of their genomic distance conservation, and that sets of genomic regions pertain to HCEs, as well as the region in which HCEs reside, should be treated as integrated domains. [ABSTRACT FROM AUTHOR]

Background: Small noncoding RNAs (ncRNAs), including short interfering RNAs (siRNAs) and microRNAs (miRNAs), can silence genes at the transcriptional, post-transcriptional or translational level [1, 2]. Results: Here, we show that microRNA-10a (miR-10a) targets a homologous DNA region in the promoter region of the hoxd4 gene and represses its expression at the transcriptional level. Mutational analysis of the miR-10a sequence revealed that the 3' end of the miRNA sequence is the most critical element for the silencing effect. MicroRNA-10a-induced transcriptional gene inhibition requires the presence of Dicer and Argonautes 1 and 3, and it is related to promoter associated noncoding RNAs. Bisulfite sequencing analysis showed that the reduced hoxd4 expression was accompanied by de novo DNA methylation at the hoxd4 promoter. We further demonstrated that trimethylation of histone 3 lysine 27 (H3K27me3) is involved in the miR-10a-induced hoxd4 transcriptional gene silence. Conclusion: In conclusion, our results demonstrate that miR-10a can regulate human gene expression in a transcriptional manner, and indicate that endogenous small noncoding RNA-induced control of transcription may be a potential system for expressional regulation in human breast cancer cells. [ABSTRACT FROM AUTHOR]

Background: A major goal of post-genomics research is the integrated analysis of genes, regulatory elements and the chromatin architecture on a genome-wide scale. Mapping DNase I hypersensitive sites within the nuclear chromatin is a powerful and well-established method of identifying regulatory element candidates. Results: Here, we report the first genome-wide analysis of DNase I hypersensitive sites (DHSs) in Caenorhabditis elegans. The data was obtained by hybridizing DNase I-treated and end-captured material from young adult worms to a high-resolution tiling microarray. The data show that C. elegans DHSs were significantly enriched within intergenic regions located 2 kb upstream and downstream of coding genes, and also that a considerable fraction of all DHSs mapped to intergenic positions distant to annotated coding genes. Annotated transcribed loci were generally depleted in DHSs relative to intergenic regions, but DHSs were nonetheless enriched in coding exons and UTRs, whereas introns were significantly depleted in DHSs. Many DHSs appeared to be associated with annotated non-coding RNAs and recently detected transcripts of unknown function. It has been reported that nematode highly conserved non-coding elements were associated with cisregulatory elements, and we also found that DHSs, particularly distal intergenic DHSs, were significantly enriched in regions that were conserved between the C. elegans and C. briggsae genomes. Conclusion: We describe the first genome-wide analysis of C. elegans DHSs, and show that the distribution of DHSs is strongly associated with functional elements in the genome. [ABSTRACT FROM AUTHOR]

Large numbers of sequence elements have been identified to be highly conserved among vertebrate genomes. These highly conserved elements (HCEs) are often located in or around genes that are involved in transcription regulation and early development. They have been shown to be involved in cis-regulatory activities through both in vivo and additional computational studies. We have investigated the structural relationships between such elements and genes in six vertebrate genomes human, mouse, rat, chicken, zebrafish and tetraodon and detected several thousand cases of conserved HCE-gene associations, and also cases of HCEs with no common target genes. A few examples underscore the potential significance of our findings about several individual genes. We found that the conserved association between HCE/HCEs and gene/genes are not restricted to elements by their absolute distance on the genome. Notably, long-range associations were identified and the molecular functions of the associated genes do not show any particular overrepresentation of the functional categories previously reported. HCEs in close proximity are found to be linked with different set of gene/genes. The results reflect the highly complex correlation between HCEs and their putative target genes. [ABSTRACT FROM AUTHOR]

Background: The 2,2,7-trimethylguanosine (TMG) cap structure is an important functional characteristic of ncRNAs with critical cellular roles, such as some snRNAs. Here we used immunoprecipitation with both K121 and R1131 anti-TMG antibodies to systematically identify the TMG cap structures for all presently characterized ncRNAs in C. elegans. Results: The two anti-TMG antibodies precipitated a similar group of the C. elegans ncRNAs. All snRNAs known to have a TMG cap structure were found in the precipitate, indicating that our identification system was efficient. Other ncRNA families related to splicing, such as SL RNAs and Sm Y RNAs, were also found in the precipitate, as were 7 C/D box snoRNAs. Further analysis showed that the SL RNAs and the Sm Y RNAs shared a very similar Sm binding site element (AAU4-5GGA), which sequence composition differed somewhat from those of other U snRNAs. There were also 16 ncRNAs without an Sm binding site element in the precipitate, suggesting that for these ncRNAs, TMG formation may occur independently of Sm proteins. Conclusion: Our results showed that most ncRNAs predicted to be transcribed by RNA polymerase II had a TMG cap, while those predicted to be transcribed by RNA plymerase III or located in introns did not have a TMG cap structure. Compared to ncRNAs without a TMG cap, TMG-capped ncRNAs tended to have higher expression levels. Five functionally non-annotated ncRNAs also have a TMG cap structure, which might be helpful for identifying the cellular roles of these ncRNAs. [ABSTRACT FROM AUTHOR]

Transcription factors with a large number of target genes—transcription hub(s), or THub(s)—are usually crucial components of the regulatory system of a cell, and the different patterns through which they transfer the transcriptional signal to downstream cascades are of great interest. By profiling normalized abundances (AN) of basic regulatory patterns of individual THubs in the yeast Saccharomyces cerevisiae transcriptional regulation network under five different cellular states and environmental conditions, we have investigated their preferences for different basic regulatory patterns. Subgraph-normalized abundances downstream of individual THubs often differ significantly from that of the network as a whole, and conversely, certain over-represented subgraphs are not preferred by any THub. The THub preferences changed substantially when the cellular or environmental conditions changed. This switching of regulatory pattern preferences suggests that a change in conditions does not only elicit a change in response by the regulatory network, but also a change in the mechanisms by which the response is mediated. The THub subgraph preference profile thus provides a novel tool for description of the structure and organization between the large-scale exponents and local regulatory patterns. [ABSTRACT FROM AUTHOR]