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Exploring mRNA 3'-UTR G-quadruplexes: evidence of roles in both alternative polyadenylation and mRNA shortening.

Beaudoin JD, Perreault JP - Nucleic Acids Res. (2013)

Bottom Line: To shed light on the role(s) of these structures, those found in the LRP5 and FXR1 genes were characterized both in vitro and in cellulo.The 3'-UTR G-quadruplexes were found to increase the efficiencies of alternative polyadenylation sites, leading to the expression of shorter transcripts and to possess the ability to interfere with the miRNA regulatory network of a specific mRNA.Clearly, G-quadruplexes located in the 3'-UTRs of mRNAs are cis-regulatory elements that have a significant impact on gene expression.

View Article: PubMed Central - PubMed

Affiliation: RNA Group/Groupe ARN, Département de biochimie, Faculté de médecine et des sciences de santé, Pavillon de recherche appliquée au cancer, Université de Sherbrooke, QC J1E 4K8, Canada.

ABSTRACT
Guanine-rich RNA sequences can fold into non-canonical, four stranded helical structures called G-quadruplexes that have been shown to be widely distributed within the mammalian transcriptome, as well as being key regulatory elements in various biological mechanisms. That said, their role within the 3'-untranslated region (UTR) of mRNA remains to be elucidated and appreciated. A bioinformatic analysis of the 3'-UTRs of mRNAs revealed enrichment in G-quadruplexes. To shed light on the role(s) of these structures, those found in the LRP5 and FXR1 genes were characterized both in vitro and in cellulo. The 3'-UTR G-quadruplexes were found to increase the efficiencies of alternative polyadenylation sites, leading to the expression of shorter transcripts and to possess the ability to interfere with the miRNA regulatory network of a specific mRNA. Clearly, G-quadruplexes located in the 3'-UTRs of mRNAs are cis-regulatory elements that have a significant impact on gene expression.

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Related in: MedlinePlus

LRP5 3′-UTR PG4 folds into a G4 structure in vitro. (a) Sequence and numbering of the wt LRP5 PG4 used in the in vitro experiments. The lowercase guanosines (g) correspond to those mutated to adenosines in the G/A-mutant version. Nucleotides that were hydrolyzed significantly more in the presence of KCl during the in-line probing are both in bold and underlined. (b, c) CD spectra for the LRP5 PG4 sequence using 4 µM of either the wt (b) or the G/A-mutant (c) versions performed either in the absence of salt (closed circle) or in the presence of 100 mM of either LiCl (inverted closed triangle), NaCl (open circle) or KCl (open triangle). (d) Autoradiogram of a 10% denaturing polyacrylamide gel of the in-line probing of the 5′-end-labeled LRP5 wt and G/A-mutant PG4 versions performed either in the absence of salt (NS), or in the presence of 100 mM of either LiCl, NaCl or KCl. Lanes L and T1 correspond to alkaline hydrolysis and RNase T1 mapping of the wt version, respectively. The positions of the guanosines are indicated on the left of the gel, whereas the domains of the G4 structure are indicated on the right.
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gkt265-F1: LRP5 3′-UTR PG4 folds into a G4 structure in vitro. (a) Sequence and numbering of the wt LRP5 PG4 used in the in vitro experiments. The lowercase guanosines (g) correspond to those mutated to adenosines in the G/A-mutant version. Nucleotides that were hydrolyzed significantly more in the presence of KCl during the in-line probing are both in bold and underlined. (b, c) CD spectra for the LRP5 PG4 sequence using 4 µM of either the wt (b) or the G/A-mutant (c) versions performed either in the absence of salt (closed circle) or in the presence of 100 mM of either LiCl (inverted closed triangle), NaCl (open circle) or KCl (open triangle). (d) Autoradiogram of a 10% denaturing polyacrylamide gel of the in-line probing of the 5′-end-labeled LRP5 wt and G/A-mutant PG4 versions performed either in the absence of salt (NS), or in the presence of 100 mM of either LiCl, NaCl or KCl. Lanes L and T1 correspond to alkaline hydrolysis and RNase T1 mapping of the wt version, respectively. The positions of the guanosines are indicated on the left of the gel, whereas the domains of the G4 structure are indicated on the right.

Mentions: Initially the PG4 located in the 3′-UTR of the low-density lipoprotein receptor-related protein 5 (LRP5) mRNA was studied as a model candidate. This PG4 sequence possesses small loops, a high number of guanosines and a low number of cystosines in flanking sequences; consequently, it possesses a strong predisposition to fold into a G4 structure (Figure 1a). Moreover, the full-length LRP5 3′-UTR is relatively short (203 nt), which significantly simplifies both the manipulations and the analysis of the data.Figure 1.


Exploring mRNA 3'-UTR G-quadruplexes: evidence of roles in both alternative polyadenylation and mRNA shortening.

Beaudoin JD, Perreault JP - Nucleic Acids Res. (2013)

LRP5 3′-UTR PG4 folds into a G4 structure in vitro. (a) Sequence and numbering of the wt LRP5 PG4 used in the in vitro experiments. The lowercase guanosines (g) correspond to those mutated to adenosines in the G/A-mutant version. Nucleotides that were hydrolyzed significantly more in the presence of KCl during the in-line probing are both in bold and underlined. (b, c) CD spectra for the LRP5 PG4 sequence using 4 µM of either the wt (b) or the G/A-mutant (c) versions performed either in the absence of salt (closed circle) or in the presence of 100 mM of either LiCl (inverted closed triangle), NaCl (open circle) or KCl (open triangle). (d) Autoradiogram of a 10% denaturing polyacrylamide gel of the in-line probing of the 5′-end-labeled LRP5 wt and G/A-mutant PG4 versions performed either in the absence of salt (NS), or in the presence of 100 mM of either LiCl, NaCl or KCl. Lanes L and T1 correspond to alkaline hydrolysis and RNase T1 mapping of the wt version, respectively. The positions of the guanosines are indicated on the left of the gel, whereas the domains of the G4 structure are indicated on the right.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3675481&req=5

gkt265-F1: LRP5 3′-UTR PG4 folds into a G4 structure in vitro. (a) Sequence and numbering of the wt LRP5 PG4 used in the in vitro experiments. The lowercase guanosines (g) correspond to those mutated to adenosines in the G/A-mutant version. Nucleotides that were hydrolyzed significantly more in the presence of KCl during the in-line probing are both in bold and underlined. (b, c) CD spectra for the LRP5 PG4 sequence using 4 µM of either the wt (b) or the G/A-mutant (c) versions performed either in the absence of salt (closed circle) or in the presence of 100 mM of either LiCl (inverted closed triangle), NaCl (open circle) or KCl (open triangle). (d) Autoradiogram of a 10% denaturing polyacrylamide gel of the in-line probing of the 5′-end-labeled LRP5 wt and G/A-mutant PG4 versions performed either in the absence of salt (NS), or in the presence of 100 mM of either LiCl, NaCl or KCl. Lanes L and T1 correspond to alkaline hydrolysis and RNase T1 mapping of the wt version, respectively. The positions of the guanosines are indicated on the left of the gel, whereas the domains of the G4 structure are indicated on the right.
Mentions: Initially the PG4 located in the 3′-UTR of the low-density lipoprotein receptor-related protein 5 (LRP5) mRNA was studied as a model candidate. This PG4 sequence possesses small loops, a high number of guanosines and a low number of cystosines in flanking sequences; consequently, it possesses a strong predisposition to fold into a G4 structure (Figure 1a). Moreover, the full-length LRP5 3′-UTR is relatively short (203 nt), which significantly simplifies both the manipulations and the analysis of the data.Figure 1.

Bottom Line: To shed light on the role(s) of these structures, those found in the LRP5 and FXR1 genes were characterized both in vitro and in cellulo.The 3'-UTR G-quadruplexes were found to increase the efficiencies of alternative polyadenylation sites, leading to the expression of shorter transcripts and to possess the ability to interfere with the miRNA regulatory network of a specific mRNA.Clearly, G-quadruplexes located in the 3'-UTRs of mRNAs are cis-regulatory elements that have a significant impact on gene expression.

View Article: PubMed Central - PubMed

Affiliation: RNA Group/Groupe ARN, Département de biochimie, Faculté de médecine et des sciences de santé, Pavillon de recherche appliquée au cancer, Université de Sherbrooke, QC J1E 4K8, Canada.

ABSTRACT
Guanine-rich RNA sequences can fold into non-canonical, four stranded helical structures called G-quadruplexes that have been shown to be widely distributed within the mammalian transcriptome, as well as being key regulatory elements in various biological mechanisms. That said, their role within the 3'-untranslated region (UTR) of mRNA remains to be elucidated and appreciated. A bioinformatic analysis of the 3'-UTRs of mRNAs revealed enrichment in G-quadruplexes. To shed light on the role(s) of these structures, those found in the LRP5 and FXR1 genes were characterized both in vitro and in cellulo. The 3'-UTR G-quadruplexes were found to increase the efficiencies of alternative polyadenylation sites, leading to the expression of shorter transcripts and to possess the ability to interfere with the miRNA regulatory network of a specific mRNA. Clearly, G-quadruplexes located in the 3'-UTRs of mRNAs are cis-regulatory elements that have a significant impact on gene expression.

Show MeSH
Related in: MedlinePlus