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Differential and coherent processing patterns from small RNAs.

Pundhir S, Gorodkin J - Sci Rep (2015)

Bottom Line: While the annotated loci predominantly consist of ~24 nt short RNAs, the unannotated loci comparatively consist of ~17 nt short RNAs.Furthermore, these ~17 nt short RNAs are significantly enriched for overlap to transcription start sites and DNase I hypersensitive sites (p-value < 0.01) that are characteristic features of transcription initiation RNAs.We discuss how the computational pipeline developed in this study has the potential to be applied to other forms of RNA-seq data for further transcriptome-wide studies of differential and coherent processing.

View Article: PubMed Central - PubMed

Affiliation: Center for non-coding RNA in Technology and Health, IKVH, University of Copenhagen, Grønnegårdsvej 3, 1870, Frederiksberg C, Denmark.

ABSTRACT
Post-transcriptional processing events related to short RNAs are often reflected in their read profile patterns emerging from high-throughput sequencing data. MicroRNA arm switching across different tissues is a well-known example of what we define as differential processing. Here, short RNAs from the nine cell lines of the ENCODE project, irrespective of their annotation status, were analyzed for genomic loci representing differential or coherent processing. We observed differential processing predominantly in RNAs annotated as miRNA, snoRNA or tRNA. Four out of five known cases of differentially processed miRNAs that were in the input dataset were recovered and several novel cases were discovered. In contrast to differential processing, coherent processing is observed widespread in both annotated and unannotated regions. While the annotated loci predominantly consist of ~24 nt short RNAs, the unannotated loci comparatively consist of ~17 nt short RNAs. Furthermore, these ~17 nt short RNAs are significantly enriched for overlap to transcription start sites and DNase I hypersensitive sites (p-value < 0.01) that are characteristic features of transcription initiation RNAs. We discuss how the computational pipeline developed in this study has the potential to be applied to other forms of RNA-seq data for further transcriptome-wide studies of differential and coherent processing.

No MeSH data available.


The fold change (log2) in the expression of twelve genes targeted by either of the two differentially processed miRNAs, has-mir-30a and has-mir-30e.In Embryonic stem cell (Esc), the 3p-arm of both the miRNAs is dominantly expressed. Consequently, we see a decrease in the expression of genes targeted by 3p-arm (green bars) in Esc. In contrast, the genes targeted by the 5p-arm, which is dominantly expressed in epithelial cells, show increase in their expression (yellow bar) in the Esc. A significant difference in the expression is computed using DESeq237 and the significance level (p-value) is indicated adjacent to each bar.
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f2: The fold change (log2) in the expression of twelve genes targeted by either of the two differentially processed miRNAs, has-mir-30a and has-mir-30e.In Embryonic stem cell (Esc), the 3p-arm of both the miRNAs is dominantly expressed. Consequently, we see a decrease in the expression of genes targeted by 3p-arm (green bars) in Esc. In contrast, the genes targeted by the 5p-arm, which is dominantly expressed in epithelial cells, show increase in their expression (yellow bar) in the Esc. A significant difference in the expression is computed using DESeq237 and the significance level (p-value) is indicated adjacent to each bar.

Mentions: We further analyzed the expression of twelve genes, which are targeted by either of the two differentially processed miRNAs (hsa-mir-30a and hsa-mir-30e; Fig. 1A,B). The miRTarBase database that contains the experimentally validated miRNA-target interactions was used to determine the twelve target genes36. These twelve genes were selected because they have been validated by three independent experimental methods (luciferase reporter assay, qRT-PCR and western blot)36. The fold change in the expression of the target genes was compared, using DESeq237 between embryonic stem cell (H1-hESC) and epithelial cell (A549) lines in which the alternative arms of both the miRNAs are dominantly expressed (Fig. 1A,B). To determine the expression of the genes in H1-hESC and A549 cell lines, we used the corresponding polyA long RNA-seq read data sets, downloaded from the ENCODE project24 in SAM format38. For all the genes, except DTL, we observed an anti-correlation between the expression of miRNA arm (5p or 3p) and the target genes (Fig. 2), which is a characteristic feature of miRNA-target interaction. Interestingly, one of the key genes, SNAI1 involved in epithelial-mesenchymal transition and target of the miRNA, hsa-mir30e-5p is also highly expressed in H1-hESC in comparison to A549. Although the higher expression of SNAI1 in H1-hESC does not reach the significance level of <0.05, an apparent increase in its expression relative to that in the A549 cell line is in agreement with the role of SNAI1 to induce epithelial to mesenchymal transition by repressing the expression of E-cadherin protein. E-cadherin is highly expressed in epithelial cells and is responsible for maintaining cell adhesion, a primary feature of epithelial cells. Thus, the lower expression of hsa-mir-30e-5p and the up-regulation of target gene, SNAI1 in H1-hESC as compared to in the A549 cell line supports the role of hsa-mir-30e in epithelial-mesenchymal transition as suggested in an earlier study29.


Differential and coherent processing patterns from small RNAs.

Pundhir S, Gorodkin J - Sci Rep (2015)

The fold change (log2) in the expression of twelve genes targeted by either of the two differentially processed miRNAs, has-mir-30a and has-mir-30e.In Embryonic stem cell (Esc), the 3p-arm of both the miRNAs is dominantly expressed. Consequently, we see a decrease in the expression of genes targeted by 3p-arm (green bars) in Esc. In contrast, the genes targeted by the 5p-arm, which is dominantly expressed in epithelial cells, show increase in their expression (yellow bar) in the Esc. A significant difference in the expression is computed using DESeq237 and the significance level (p-value) is indicated adjacent to each bar.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4499813&req=5

f2: The fold change (log2) in the expression of twelve genes targeted by either of the two differentially processed miRNAs, has-mir-30a and has-mir-30e.In Embryonic stem cell (Esc), the 3p-arm of both the miRNAs is dominantly expressed. Consequently, we see a decrease in the expression of genes targeted by 3p-arm (green bars) in Esc. In contrast, the genes targeted by the 5p-arm, which is dominantly expressed in epithelial cells, show increase in their expression (yellow bar) in the Esc. A significant difference in the expression is computed using DESeq237 and the significance level (p-value) is indicated adjacent to each bar.
Mentions: We further analyzed the expression of twelve genes, which are targeted by either of the two differentially processed miRNAs (hsa-mir-30a and hsa-mir-30e; Fig. 1A,B). The miRTarBase database that contains the experimentally validated miRNA-target interactions was used to determine the twelve target genes36. These twelve genes were selected because they have been validated by three independent experimental methods (luciferase reporter assay, qRT-PCR and western blot)36. The fold change in the expression of the target genes was compared, using DESeq237 between embryonic stem cell (H1-hESC) and epithelial cell (A549) lines in which the alternative arms of both the miRNAs are dominantly expressed (Fig. 1A,B). To determine the expression of the genes in H1-hESC and A549 cell lines, we used the corresponding polyA long RNA-seq read data sets, downloaded from the ENCODE project24 in SAM format38. For all the genes, except DTL, we observed an anti-correlation between the expression of miRNA arm (5p or 3p) and the target genes (Fig. 2), which is a characteristic feature of miRNA-target interaction. Interestingly, one of the key genes, SNAI1 involved in epithelial-mesenchymal transition and target of the miRNA, hsa-mir30e-5p is also highly expressed in H1-hESC in comparison to A549. Although the higher expression of SNAI1 in H1-hESC does not reach the significance level of <0.05, an apparent increase in its expression relative to that in the A549 cell line is in agreement with the role of SNAI1 to induce epithelial to mesenchymal transition by repressing the expression of E-cadherin protein. E-cadherin is highly expressed in epithelial cells and is responsible for maintaining cell adhesion, a primary feature of epithelial cells. Thus, the lower expression of hsa-mir-30e-5p and the up-regulation of target gene, SNAI1 in H1-hESC as compared to in the A549 cell line supports the role of hsa-mir-30e in epithelial-mesenchymal transition as suggested in an earlier study29.

Bottom Line: While the annotated loci predominantly consist of ~24 nt short RNAs, the unannotated loci comparatively consist of ~17 nt short RNAs.Furthermore, these ~17 nt short RNAs are significantly enriched for overlap to transcription start sites and DNase I hypersensitive sites (p-value < 0.01) that are characteristic features of transcription initiation RNAs.We discuss how the computational pipeline developed in this study has the potential to be applied to other forms of RNA-seq data for further transcriptome-wide studies of differential and coherent processing.

View Article: PubMed Central - PubMed

Affiliation: Center for non-coding RNA in Technology and Health, IKVH, University of Copenhagen, Grønnegårdsvej 3, 1870, Frederiksberg C, Denmark.

ABSTRACT
Post-transcriptional processing events related to short RNAs are often reflected in their read profile patterns emerging from high-throughput sequencing data. MicroRNA arm switching across different tissues is a well-known example of what we define as differential processing. Here, short RNAs from the nine cell lines of the ENCODE project, irrespective of their annotation status, were analyzed for genomic loci representing differential or coherent processing. We observed differential processing predominantly in RNAs annotated as miRNA, snoRNA or tRNA. Four out of five known cases of differentially processed miRNAs that were in the input dataset were recovered and several novel cases were discovered. In contrast to differential processing, coherent processing is observed widespread in both annotated and unannotated regions. While the annotated loci predominantly consist of ~24 nt short RNAs, the unannotated loci comparatively consist of ~17 nt short RNAs. Furthermore, these ~17 nt short RNAs are significantly enriched for overlap to transcription start sites and DNase I hypersensitive sites (p-value < 0.01) that are characteristic features of transcription initiation RNAs. We discuss how the computational pipeline developed in this study has the potential to be applied to other forms of RNA-seq data for further transcriptome-wide studies of differential and coherent processing.

No MeSH data available.