Limits...
Correlation between the secondary structure of pre-mRNA introns and the efficiency of splicing in Saccharomyces cerevisiae.

Rogic S, Montpetit B, Hoos HH, Mackworth AK, Ouellette BF, Hieter P - BMC Genomics (2008)

Bottom Line: The nature of these base-pairing interactions and their effect on splicing efficiency were most extensively studied in ribosomal protein gene RPS17B (previously known as RP51B).We show that it is necessary to consider suboptimal structure predictions and to compute the structural branchpoint distances in order to explain previously published splicing efficiency results.Our findings provide further insights into the role of pre-mRNA secondary structure in gene splicing in yeast and also offer basis for improvement of computational methods for splice site identification and gene-finding.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Computer Science, University of British Columbia, Vancouver, Canada. rogic@bioinformatics.ubc.ca

ABSTRACT

Background: Secondary structure interactions within introns have been shown to be essential for efficient splicing of several yeast genes. The nature of these base-pairing interactions and their effect on splicing efficiency were most extensively studied in ribosomal protein gene RPS17B (previously known as RP51B). It was determined that complementary pairing between two sequence segments located downstream of the 5' splice site and upstream of the branchpoint sequence promotes efficient splicing of the RPS17B pre-mRNA, presumably by shortening the branchpoint distance. However, no attempts were made to compute a shortened, 'structural' branchpoint distance and thus the functional relationship between this distance and the splicing efficiency remains unknown.

Results: In this paper we use computational RNA secondary structure prediction to analyze the secondary structure of the RPS17B intron. We show that it is necessary to consider suboptimal structure predictions and to compute the structural branchpoint distances in order to explain previously published splicing efficiency results. Our study reveals that there is a tight correlation between this distance and splicing efficiency levels of intron mutants described in the literature. We experimentally test this correlation on additional RPS17B mutants and intron mutants within two other yeast genes.

Conclusion: The proposed model of secondary structure requirements for efficient splicing is the first attempt to specify the functional relationship between pre-mRNA secondary structure and splicing. Our findings provide further insights into the role of pre-mRNA secondary structure in gene splicing in yeast and also offer basis for improvement of computational methods for splice site identification and gene-finding.

Show MeSH
Protein expression results for the APE2 gene containing the newly designed mutant introns. Protein expression level is normalized with respect to wildtype expression level. Shaded boxes represent the mean value for several different samples and error bars represent +/- 1 standard deviation for these samples.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC2536676&req=5

Figure 8: Protein expression results for the APE2 gene containing the newly designed mutant introns. Protein expression level is normalized with respect to wildtype expression level. Shaded boxes represent the mean value for several different samples and error bars represent +/- 1 standard deviation for these samples.

Mentions: The experimental results are consistent with our prediction for five out of seven mutants: mutants ape2-S1, ape2-S2, ape2-S3 and ape2-S5 all have a level of protein abundance similar to the wildtype (Figure 8) and mutant ape2-L1 shows significantly reduced expression as expected. Mutant ape2-L2, which was expected to have reduced protein abundance as a consequence of reduced splicing efficiency, is expressed at the same level as the wildtype. Also, mutant ape2-S4 has reduced splicing despite the fact that it has a similar distribution of structural branchpoint distances as the wildtype intron. Since this mutant has the mutation at the same location as ape2-L1 (see Materials and Methods), it is possible that the intron segment that we mutated was important for splicing (e.g., contained a splicing enhancer). Overall, the results for APE2 mutants support our hypothesis of the role of structural branchpoint distance in gene splicing.


Correlation between the secondary structure of pre-mRNA introns and the efficiency of splicing in Saccharomyces cerevisiae.

Rogic S, Montpetit B, Hoos HH, Mackworth AK, Ouellette BF, Hieter P - BMC Genomics (2008)

Protein expression results for the APE2 gene containing the newly designed mutant introns. Protein expression level is normalized with respect to wildtype expression level. Shaded boxes represent the mean value for several different samples and error bars represent +/- 1 standard deviation for these samples.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2536676&req=5

Figure 8: Protein expression results for the APE2 gene containing the newly designed mutant introns. Protein expression level is normalized with respect to wildtype expression level. Shaded boxes represent the mean value for several different samples and error bars represent +/- 1 standard deviation for these samples.
Mentions: The experimental results are consistent with our prediction for five out of seven mutants: mutants ape2-S1, ape2-S2, ape2-S3 and ape2-S5 all have a level of protein abundance similar to the wildtype (Figure 8) and mutant ape2-L1 shows significantly reduced expression as expected. Mutant ape2-L2, which was expected to have reduced protein abundance as a consequence of reduced splicing efficiency, is expressed at the same level as the wildtype. Also, mutant ape2-S4 has reduced splicing despite the fact that it has a similar distribution of structural branchpoint distances as the wildtype intron. Since this mutant has the mutation at the same location as ape2-L1 (see Materials and Methods), it is possible that the intron segment that we mutated was important for splicing (e.g., contained a splicing enhancer). Overall, the results for APE2 mutants support our hypothesis of the role of structural branchpoint distance in gene splicing.

Bottom Line: The nature of these base-pairing interactions and their effect on splicing efficiency were most extensively studied in ribosomal protein gene RPS17B (previously known as RP51B).We show that it is necessary to consider suboptimal structure predictions and to compute the structural branchpoint distances in order to explain previously published splicing efficiency results.Our findings provide further insights into the role of pre-mRNA secondary structure in gene splicing in yeast and also offer basis for improvement of computational methods for splice site identification and gene-finding.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Computer Science, University of British Columbia, Vancouver, Canada. rogic@bioinformatics.ubc.ca

ABSTRACT

Background: Secondary structure interactions within introns have been shown to be essential for efficient splicing of several yeast genes. The nature of these base-pairing interactions and their effect on splicing efficiency were most extensively studied in ribosomal protein gene RPS17B (previously known as RP51B). It was determined that complementary pairing between two sequence segments located downstream of the 5' splice site and upstream of the branchpoint sequence promotes efficient splicing of the RPS17B pre-mRNA, presumably by shortening the branchpoint distance. However, no attempts were made to compute a shortened, 'structural' branchpoint distance and thus the functional relationship between this distance and the splicing efficiency remains unknown.

Results: In this paper we use computational RNA secondary structure prediction to analyze the secondary structure of the RPS17B intron. We show that it is necessary to consider suboptimal structure predictions and to compute the structural branchpoint distances in order to explain previously published splicing efficiency results. Our study reveals that there is a tight correlation between this distance and splicing efficiency levels of intron mutants described in the literature. We experimentally test this correlation on additional RPS17B mutants and intron mutants within two other yeast genes.

Conclusion: The proposed model of secondary structure requirements for efficient splicing is the first attempt to specify the functional relationship between pre-mRNA secondary structure and splicing. Our findings provide further insights into the role of pre-mRNA secondary structure in gene splicing in yeast and also offer basis for improvement of computational methods for splice site identification and gene-finding.

Show MeSH