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Diverse RNA-binding proteins interact with functionally related sets of RNAs, suggesting an extensive regulatory system.

Hogan DJ, Riordan DP, Gerber AP, Herschlag D, Brown PO - PLoS Biol. (2008)

Bottom Line: These potential RNA-recognition elements were diverse in sequence, structure, and location: some were found predominantly in 3'-untranslated regions, others in 5'-untranslated regions, some in coding sequences, and many in two or more of these features.Although this study only examined a small fraction of the universe of yeast RBPs, 70% of the mRNA transcriptome had significant associations with at least one of these RBPs, and on average, each distinct yeast mRNA interacted with three of the RBPs, suggesting the potential for a rich, multidimensional network of regulation.These results strongly suggest that combinatorial binding of RBPs to specific recognition elements in mRNAs is a pervasive mechanism for multi-dimensional regulation of their post-transcriptional fate.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Stanford University School of Medicine, Stanford, California, United States of America.

ABSTRACT
RNA-binding proteins (RBPs) have roles in the regulation of many post-transcriptional steps in gene expression, but relatively few RBPs have been systematically studied. We searched for the RNA targets of 40 proteins in the yeast Saccharomyces cerevisiae: a selective sample of the approximately 600 annotated and predicted RBPs, as well as several proteins not annotated as RBPs. At least 33 of these 40 proteins, including three of the four proteins that were not previously known or predicted to be RBPs, were reproducibly associated with specific sets of a few to several hundred RNAs. Remarkably, many of the RBPs we studied bound mRNAs whose protein products share identifiable functional or cytotopic features. We identified specific sequences or predicted structures significantly enriched in target mRNAs of 16 RBPs. These potential RNA-recognition elements were diverse in sequence, structure, and location: some were found predominantly in 3'-untranslated regions, others in 5'-untranslated regions, some in coding sequences, and many in two or more of these features. Although this study only examined a small fraction of the universe of yeast RBPs, 70% of the mRNA transcriptome had significant associations with at least one of these RBPs, and on average, each distinct yeast mRNA interacted with three of the RBPs, suggesting the potential for a rich, multidimensional network of regulation. These results strongly suggest that combinatorial binding of RBPs to specific recognition elements in mRNAs is a pervasive mechanism for multi-dimensional regulation of their post-transcriptional fate.

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Diverse Combinatorial Patterns of RNA-Binding Protein Interactions with a Choice Sample of mRNAs(A–K) Putative binding sites of RBPs in target mRNAs. The relative lengths of the 5′-UTR, coding sequence, and 3′-UTR are drawn to scale. For mRNAs for which there are reliable measurements for untranslated sequence lengths (SUN4, DSE2, SCW4, CLN2, PUF2, PMA1, SUR7, and HHT1) [63], we added 50 bases onto the estimated 5′-UTR and 3′-UTR lengths, because the estimated UTR lengths are likely conservative. For mRNAs that do not have reliable untranslated region measurements (CTS1, EGT2, and MRP1), we used 250 bases upstream and downstream of the coding sequence as the 5′-UTR and 3′-UTR, respectively. The positions of the start and stop codons are indicated by stop signals. Putative binding sites for RBPs with strong evidence for association (1% FDR) are marked (Puf3-REFINE, Puf4-FIRE, Puf5-REFINE, Pub1-FIRE, Puf1/2-REFINE, Ssd1-REFINE, Nsr1-REFINE, Yll032c-REFINE, Pin4-REFINE, and Nrd1/Nab3-REFINE) (Figure 5 and Table S4). RBPs that we found to be associated with the mRNA, but for which the recognition elements are not yet known, are listed to the right of the mRNA. The number of Pab1 molecules shown bound to the poly(A) tail represents the degree of enrichment of the corresponding mRNA in the Pab1 IPs (log2 immunopurification enrichment −6 = 0, −5 = 1, etc.) and not the number of Pab1 molecules bound per mRNA. The cap-binding proteins, Cbc1/2 and eiF4e, are shown by default at the cap site.
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pbio-0060255-g006: Diverse Combinatorial Patterns of RNA-Binding Protein Interactions with a Choice Sample of mRNAs(A–K) Putative binding sites of RBPs in target mRNAs. The relative lengths of the 5′-UTR, coding sequence, and 3′-UTR are drawn to scale. For mRNAs for which there are reliable measurements for untranslated sequence lengths (SUN4, DSE2, SCW4, CLN2, PUF2, PMA1, SUR7, and HHT1) [63], we added 50 bases onto the estimated 5′-UTR and 3′-UTR lengths, because the estimated UTR lengths are likely conservative. For mRNAs that do not have reliable untranslated region measurements (CTS1, EGT2, and MRP1), we used 250 bases upstream and downstream of the coding sequence as the 5′-UTR and 3′-UTR, respectively. The positions of the start and stop codons are indicated by stop signals. Putative binding sites for RBPs with strong evidence for association (1% FDR) are marked (Puf3-REFINE, Puf4-FIRE, Puf5-REFINE, Pub1-FIRE, Puf1/2-REFINE, Ssd1-REFINE, Nsr1-REFINE, Yll032c-REFINE, Pin4-REFINE, and Nrd1/Nab3-REFINE) (Figure 5 and Table S4). RBPs that we found to be associated with the mRNA, but for which the recognition elements are not yet known, are listed to the right of the mRNA. The number of Pab1 molecules shown bound to the poly(A) tail represents the degree of enrichment of the corresponding mRNA in the Pab1 IPs (log2 immunopurification enrichment −6 = 0, −5 = 1, etc.) and not the number of Pab1 molecules bound per mRNA. The cap-binding proteins, Cbc1/2 and eiF4e, are shown by default at the cap site.

Mentions: A selective sample of 11 mRNAs provides an unfinished, but revealing, picture of the organization of the information that specifies interactions with, and perhaps regulation by, specific RBPs examined in this study (Figure 6). For each mRNA, the location of high-confidence RNA recognition elements for RBPs that interact with the mRNA are indicated, while RBPs that interact with the mRNA, but whose binding site is uncertain, are shown to the right of the mRNA. The relative lengths of the 5′-UTR, coding sequence, and 3′-UTR are drawn to scale, and the translation start and stop codons are depicted with the corresponding “traffic signal.” Each of these mRNAs has specific interactions with overlapping, but distinct, subsets of RBPs in the study. The putative binding patterns of specific RBPs, with respect to the number and locations of sites, vary considerably among the mRNAs, which may have important functional consequences. The first five mRNAs (SUN4, DSE2, CTS1, SCW4, and EGT2) encode cell-wall enzymes (Figure 6A–6E). Each of these mRNAs associated with five to nine RBPs in this study, including all five with Pub1, Khd1, and Ypl184c, four with Ssd1 (SUN4, DSE2, CTS1, and SCW4), three with Scp160 (CTS1, SCW4, and EGT2), and two with Nab6 (CTS1 and SCW4) and Nrd1 (DSE2 and EGT2). In addition to these overlapping interactions, most of these mRNAs associated with a unique set of additional RBPs; for instance, SUN4 contains two Puf5-binding sites in its 3′-UTR and EGT2 contains eight She2-binding sites in its coding sequence. CLN2 encodes a G1 cyclin and associated with many of the same RBPs as SUN4, DSE2, CTS1, SCW4, and EGT2 (Figure 6F). PUF2 associated with several RBPs, including its cognate protein, which is common among RBPs in this study (Text S8); there are 12 Puf2-binding sites in its coding sequence (Figure 6G). PMA1 associated with a similar set of RBPs as PUF2, including Pub1 and Puf2, but the locations and numbers of binding sites for these RBPs are very different in the two mRNAs (Figure 6H). The putative binding sites for Puf4 and Puf5 in the 3′-UTR of HHT1 partially overlap, suggesting these RBPs may compete for binding to this mRNA (Figure 6J). These diagrams represent only a partial picture of the RBP interactions with these mRNAs; the mRNA targets have only been defined for a small fraction of all yeast RBPs, and the sequence elements that specify many of the interactions we have identified are not yet known.


Diverse RNA-binding proteins interact with functionally related sets of RNAs, suggesting an extensive regulatory system.

Hogan DJ, Riordan DP, Gerber AP, Herschlag D, Brown PO - PLoS Biol. (2008)

Diverse Combinatorial Patterns of RNA-Binding Protein Interactions with a Choice Sample of mRNAs(A–K) Putative binding sites of RBPs in target mRNAs. The relative lengths of the 5′-UTR, coding sequence, and 3′-UTR are drawn to scale. For mRNAs for which there are reliable measurements for untranslated sequence lengths (SUN4, DSE2, SCW4, CLN2, PUF2, PMA1, SUR7, and HHT1) [63], we added 50 bases onto the estimated 5′-UTR and 3′-UTR lengths, because the estimated UTR lengths are likely conservative. For mRNAs that do not have reliable untranslated region measurements (CTS1, EGT2, and MRP1), we used 250 bases upstream and downstream of the coding sequence as the 5′-UTR and 3′-UTR, respectively. The positions of the start and stop codons are indicated by stop signals. Putative binding sites for RBPs with strong evidence for association (1% FDR) are marked (Puf3-REFINE, Puf4-FIRE, Puf5-REFINE, Pub1-FIRE, Puf1/2-REFINE, Ssd1-REFINE, Nsr1-REFINE, Yll032c-REFINE, Pin4-REFINE, and Nrd1/Nab3-REFINE) (Figure 5 and Table S4). RBPs that we found to be associated with the mRNA, but for which the recognition elements are not yet known, are listed to the right of the mRNA. The number of Pab1 molecules shown bound to the poly(A) tail represents the degree of enrichment of the corresponding mRNA in the Pab1 IPs (log2 immunopurification enrichment −6 = 0, −5 = 1, etc.) and not the number of Pab1 molecules bound per mRNA. The cap-binding proteins, Cbc1/2 and eiF4e, are shown by default at the cap site.
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Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC2573929&req=5

pbio-0060255-g006: Diverse Combinatorial Patterns of RNA-Binding Protein Interactions with a Choice Sample of mRNAs(A–K) Putative binding sites of RBPs in target mRNAs. The relative lengths of the 5′-UTR, coding sequence, and 3′-UTR are drawn to scale. For mRNAs for which there are reliable measurements for untranslated sequence lengths (SUN4, DSE2, SCW4, CLN2, PUF2, PMA1, SUR7, and HHT1) [63], we added 50 bases onto the estimated 5′-UTR and 3′-UTR lengths, because the estimated UTR lengths are likely conservative. For mRNAs that do not have reliable untranslated region measurements (CTS1, EGT2, and MRP1), we used 250 bases upstream and downstream of the coding sequence as the 5′-UTR and 3′-UTR, respectively. The positions of the start and stop codons are indicated by stop signals. Putative binding sites for RBPs with strong evidence for association (1% FDR) are marked (Puf3-REFINE, Puf4-FIRE, Puf5-REFINE, Pub1-FIRE, Puf1/2-REFINE, Ssd1-REFINE, Nsr1-REFINE, Yll032c-REFINE, Pin4-REFINE, and Nrd1/Nab3-REFINE) (Figure 5 and Table S4). RBPs that we found to be associated with the mRNA, but for which the recognition elements are not yet known, are listed to the right of the mRNA. The number of Pab1 molecules shown bound to the poly(A) tail represents the degree of enrichment of the corresponding mRNA in the Pab1 IPs (log2 immunopurification enrichment −6 = 0, −5 = 1, etc.) and not the number of Pab1 molecules bound per mRNA. The cap-binding proteins, Cbc1/2 and eiF4e, are shown by default at the cap site.
Mentions: A selective sample of 11 mRNAs provides an unfinished, but revealing, picture of the organization of the information that specifies interactions with, and perhaps regulation by, specific RBPs examined in this study (Figure 6). For each mRNA, the location of high-confidence RNA recognition elements for RBPs that interact with the mRNA are indicated, while RBPs that interact with the mRNA, but whose binding site is uncertain, are shown to the right of the mRNA. The relative lengths of the 5′-UTR, coding sequence, and 3′-UTR are drawn to scale, and the translation start and stop codons are depicted with the corresponding “traffic signal.” Each of these mRNAs has specific interactions with overlapping, but distinct, subsets of RBPs in the study. The putative binding patterns of specific RBPs, with respect to the number and locations of sites, vary considerably among the mRNAs, which may have important functional consequences. The first five mRNAs (SUN4, DSE2, CTS1, SCW4, and EGT2) encode cell-wall enzymes (Figure 6A–6E). Each of these mRNAs associated with five to nine RBPs in this study, including all five with Pub1, Khd1, and Ypl184c, four with Ssd1 (SUN4, DSE2, CTS1, and SCW4), three with Scp160 (CTS1, SCW4, and EGT2), and two with Nab6 (CTS1 and SCW4) and Nrd1 (DSE2 and EGT2). In addition to these overlapping interactions, most of these mRNAs associated with a unique set of additional RBPs; for instance, SUN4 contains two Puf5-binding sites in its 3′-UTR and EGT2 contains eight She2-binding sites in its coding sequence. CLN2 encodes a G1 cyclin and associated with many of the same RBPs as SUN4, DSE2, CTS1, SCW4, and EGT2 (Figure 6F). PUF2 associated with several RBPs, including its cognate protein, which is common among RBPs in this study (Text S8); there are 12 Puf2-binding sites in its coding sequence (Figure 6G). PMA1 associated with a similar set of RBPs as PUF2, including Pub1 and Puf2, but the locations and numbers of binding sites for these RBPs are very different in the two mRNAs (Figure 6H). The putative binding sites for Puf4 and Puf5 in the 3′-UTR of HHT1 partially overlap, suggesting these RBPs may compete for binding to this mRNA (Figure 6J). These diagrams represent only a partial picture of the RBP interactions with these mRNAs; the mRNA targets have only been defined for a small fraction of all yeast RBPs, and the sequence elements that specify many of the interactions we have identified are not yet known.

Bottom Line: These potential RNA-recognition elements were diverse in sequence, structure, and location: some were found predominantly in 3'-untranslated regions, others in 5'-untranslated regions, some in coding sequences, and many in two or more of these features.Although this study only examined a small fraction of the universe of yeast RBPs, 70% of the mRNA transcriptome had significant associations with at least one of these RBPs, and on average, each distinct yeast mRNA interacted with three of the RBPs, suggesting the potential for a rich, multidimensional network of regulation.These results strongly suggest that combinatorial binding of RBPs to specific recognition elements in mRNAs is a pervasive mechanism for multi-dimensional regulation of their post-transcriptional fate.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Stanford University School of Medicine, Stanford, California, United States of America.

ABSTRACT
RNA-binding proteins (RBPs) have roles in the regulation of many post-transcriptional steps in gene expression, but relatively few RBPs have been systematically studied. We searched for the RNA targets of 40 proteins in the yeast Saccharomyces cerevisiae: a selective sample of the approximately 600 annotated and predicted RBPs, as well as several proteins not annotated as RBPs. At least 33 of these 40 proteins, including three of the four proteins that were not previously known or predicted to be RBPs, were reproducibly associated with specific sets of a few to several hundred RNAs. Remarkably, many of the RBPs we studied bound mRNAs whose protein products share identifiable functional or cytotopic features. We identified specific sequences or predicted structures significantly enriched in target mRNAs of 16 RBPs. These potential RNA-recognition elements were diverse in sequence, structure, and location: some were found predominantly in 3'-untranslated regions, others in 5'-untranslated regions, some in coding sequences, and many in two or more of these features. Although this study only examined a small fraction of the universe of yeast RBPs, 70% of the mRNA transcriptome had significant associations with at least one of these RBPs, and on average, each distinct yeast mRNA interacted with three of the RBPs, suggesting the potential for a rich, multidimensional network of regulation. These results strongly suggest that combinatorial binding of RBPs to specific recognition elements in mRNAs is a pervasive mechanism for multi-dimensional regulation of their post-transcriptional fate.

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Related in: MedlinePlus