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Genomic analysis of ADAR1 binding and its involvement in multiple RNA processing pathways.

Bahn JH, Ahn J, Lin X, Zhang Q, Lee JH, Civelek M, Xiao X - Nat Commun (2015)

Bottom Line: Similarly, ADAR1 interacts with DROSHA and DGCR8 in the nucleus and possibly out-competes DGCR8 in primary miRNA binding, which enhances mature miRNA expression.These functions are dependent on ADAR1's editing activity, at least for a subset of targets.Our study unfolds a broad landscape of the functional roles of ADAR1.

View Article: PubMed Central - PubMed

Affiliation: Department of Integrative Biology and Physiology and the Molecular Biology Institute, University of California Los Angeles, Los Angeles, California 90095, USA.

ABSTRACT
Adenosine deaminases acting on RNA (ADARs) are the primary factors underlying adenosine to inosine (A-to-I) editing in metazoans. Here we report the first global study of ADAR1-RNA interaction in human cells using CLIP-seq. A large number of CLIP sites are observed in Alu repeats, consistent with ADAR1's function in RNA editing. Surprisingly, thousands of other CLIP sites are located in non-Alu regions, revealing functional and biophysical targets of ADAR1 in the regulation of alternative 3' UTR usage and miRNA biogenesis. We observe that binding of ADAR1 to 3' UTRs precludes binding by other factors, causing 3' UTR lengthening. Similarly, ADAR1 interacts with DROSHA and DGCR8 in the nucleus and possibly out-competes DGCR8 in primary miRNA binding, which enhances mature miRNA expression. These functions are dependent on ADAR1's editing activity, at least for a subset of targets. Our study unfolds a broad landscape of the functional roles of ADAR1.

No MeSH data available.


ADAR1 binding signature reflects its function in RNA editing(a) Shortest distance between ADAR1-bound Alu sites and RNA editing sites (DARNED database, same below) in the same gene. Linear: linear genomic distance; structural: distance calculated between predicted dsRNA structures harboring the CLIP cluster and editing sites; control: distance between CLIP clusters and random A's chosen from the same region as authentic editing sites. Both linear and structural distances are significantly smaller than control (p < 2.2e-16, Kolmogorov–Smirnov (KS) test). (b) Histogram of distance (up to 100nt) between deletions in CLIP reads and closest RNA editing sites in the same gene. Red dashed line represents the average distance in the range shown. (c) Histogram of closest distance between ADAR1-bound Alu clusters in the same gene. A and B denote the bottom and top 5% of distances respectively. (d) Genomic distribution of editing sites within 100nt of Alu clusters in groups A and B as defined in (c). The distribution of these editing sites in different regions of annotated genes is shown. Note that no editing sites were found in coding exons. (e) Conservation level of positions surrounding editing sites in groups A and B across primates. DNA conservation was calculated as % sequence identify. Light shaded area represents confidence intervals.
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Figure 2: ADAR1 binding signature reflects its function in RNA editing(a) Shortest distance between ADAR1-bound Alu sites and RNA editing sites (DARNED database, same below) in the same gene. Linear: linear genomic distance; structural: distance calculated between predicted dsRNA structures harboring the CLIP cluster and editing sites; control: distance between CLIP clusters and random A's chosen from the same region as authentic editing sites. Both linear and structural distances are significantly smaller than control (p < 2.2e-16, Kolmogorov–Smirnov (KS) test). (b) Histogram of distance (up to 100nt) between deletions in CLIP reads and closest RNA editing sites in the same gene. Red dashed line represents the average distance in the range shown. (c) Histogram of closest distance between ADAR1-bound Alu clusters in the same gene. A and B denote the bottom and top 5% of distances respectively. (d) Genomic distribution of editing sites within 100nt of Alu clusters in groups A and B as defined in (c). The distribution of these editing sites in different regions of annotated genes is shown. Note that no editing sites were found in coding exons. (e) Conservation level of positions surrounding editing sites in groups A and B across primates. DNA conservation was calculated as % sequence identify. Light shaded area represents confidence intervals.

Mentions: We next examined the relationship between ADAR1 binding and RNA editing in detail with a focus on CLIP sites within Alu repeats. We analyzed the distance between ADAR1 CLIP clusters and their respective closest known A-to-I editing sites. As shown in Fig. 2a, the linear distance from binding to editing sites was significantly smaller than to controls calculated for random A's in the same region. Moreover, the binding sites were even closer to editing sites if the distances were calculated between the editing sites and predicted dsRNA structures harboring the CLIP cluster. In particular, >20% of Alu-containing structures overlapped with A-to-I editing sites and about 50% of the CLIP clusters were located relative to editing sites in a distance of at least two orders of magnitude closer than expected by chance. It should be noted that the absolute distance between CLIP clusters and editing sites is relatively high (median ~1kb for the structured ones) possibly due to the facts that many more editing site are yet to be identified and/or the CLIP experiments did not capture all ADAR1 binding sites.


Genomic analysis of ADAR1 binding and its involvement in multiple RNA processing pathways.

Bahn JH, Ahn J, Lin X, Zhang Q, Lee JH, Civelek M, Xiao X - Nat Commun (2015)

ADAR1 binding signature reflects its function in RNA editing(a) Shortest distance between ADAR1-bound Alu sites and RNA editing sites (DARNED database, same below) in the same gene. Linear: linear genomic distance; structural: distance calculated between predicted dsRNA structures harboring the CLIP cluster and editing sites; control: distance between CLIP clusters and random A's chosen from the same region as authentic editing sites. Both linear and structural distances are significantly smaller than control (p < 2.2e-16, Kolmogorov–Smirnov (KS) test). (b) Histogram of distance (up to 100nt) between deletions in CLIP reads and closest RNA editing sites in the same gene. Red dashed line represents the average distance in the range shown. (c) Histogram of closest distance between ADAR1-bound Alu clusters in the same gene. A and B denote the bottom and top 5% of distances respectively. (d) Genomic distribution of editing sites within 100nt of Alu clusters in groups A and B as defined in (c). The distribution of these editing sites in different regions of annotated genes is shown. Note that no editing sites were found in coding exons. (e) Conservation level of positions surrounding editing sites in groups A and B across primates. DNA conservation was calculated as % sequence identify. Light shaded area represents confidence intervals.
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Related In: Results  -  Collection

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Figure 2: ADAR1 binding signature reflects its function in RNA editing(a) Shortest distance between ADAR1-bound Alu sites and RNA editing sites (DARNED database, same below) in the same gene. Linear: linear genomic distance; structural: distance calculated between predicted dsRNA structures harboring the CLIP cluster and editing sites; control: distance between CLIP clusters and random A's chosen from the same region as authentic editing sites. Both linear and structural distances are significantly smaller than control (p < 2.2e-16, Kolmogorov–Smirnov (KS) test). (b) Histogram of distance (up to 100nt) between deletions in CLIP reads and closest RNA editing sites in the same gene. Red dashed line represents the average distance in the range shown. (c) Histogram of closest distance between ADAR1-bound Alu clusters in the same gene. A and B denote the bottom and top 5% of distances respectively. (d) Genomic distribution of editing sites within 100nt of Alu clusters in groups A and B as defined in (c). The distribution of these editing sites in different regions of annotated genes is shown. Note that no editing sites were found in coding exons. (e) Conservation level of positions surrounding editing sites in groups A and B across primates. DNA conservation was calculated as % sequence identify. Light shaded area represents confidence intervals.
Mentions: We next examined the relationship between ADAR1 binding and RNA editing in detail with a focus on CLIP sites within Alu repeats. We analyzed the distance between ADAR1 CLIP clusters and their respective closest known A-to-I editing sites. As shown in Fig. 2a, the linear distance from binding to editing sites was significantly smaller than to controls calculated for random A's in the same region. Moreover, the binding sites were even closer to editing sites if the distances were calculated between the editing sites and predicted dsRNA structures harboring the CLIP cluster. In particular, >20% of Alu-containing structures overlapped with A-to-I editing sites and about 50% of the CLIP clusters were located relative to editing sites in a distance of at least two orders of magnitude closer than expected by chance. It should be noted that the absolute distance between CLIP clusters and editing sites is relatively high (median ~1kb for the structured ones) possibly due to the facts that many more editing site are yet to be identified and/or the CLIP experiments did not capture all ADAR1 binding sites.

Bottom Line: Similarly, ADAR1 interacts with DROSHA and DGCR8 in the nucleus and possibly out-competes DGCR8 in primary miRNA binding, which enhances mature miRNA expression.These functions are dependent on ADAR1's editing activity, at least for a subset of targets.Our study unfolds a broad landscape of the functional roles of ADAR1.

View Article: PubMed Central - PubMed

Affiliation: Department of Integrative Biology and Physiology and the Molecular Biology Institute, University of California Los Angeles, Los Angeles, California 90095, USA.

ABSTRACT
Adenosine deaminases acting on RNA (ADARs) are the primary factors underlying adenosine to inosine (A-to-I) editing in metazoans. Here we report the first global study of ADAR1-RNA interaction in human cells using CLIP-seq. A large number of CLIP sites are observed in Alu repeats, consistent with ADAR1's function in RNA editing. Surprisingly, thousands of other CLIP sites are located in non-Alu regions, revealing functional and biophysical targets of ADAR1 in the regulation of alternative 3' UTR usage and miRNA biogenesis. We observe that binding of ADAR1 to 3' UTRs precludes binding by other factors, causing 3' UTR lengthening. Similarly, ADAR1 interacts with DROSHA and DGCR8 in the nucleus and possibly out-competes DGCR8 in primary miRNA binding, which enhances mature miRNA expression. These functions are dependent on ADAR1's editing activity, at least for a subset of targets. Our study unfolds a broad landscape of the functional roles of ADAR1.

No MeSH data available.