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Kinetic competition during the transcription cycle results in stochastic RNA processing.

Coulon A, Ferguson ML, de Turris V, Palangat M, Chow CC, Larson DR - Elife (2014)

Bottom Line: We find that kinetic competition results in multiple competing pathways for pre-mRNA splicing.Splicing of the terminal intron occurs stochastically both before and after transcript release, indicating there is not a strict quality control checkpoint.The majority of pre-mRNAs are spliced after release, while diffusing away from the site of transcription.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Biological Modeling, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, United States.

ABSTRACT
Synthesis of mRNA in eukaryotes involves the coordinated action of many enzymatic processes, including initiation, elongation, splicing, and cleavage. Kinetic competition between these processes has been proposed to determine RNA fate, yet such coupling has never been observed in vivo on single transcripts. In this study, we use dual-color single-molecule RNA imaging in living human cells to construct a complete kinetic profile of transcription and splicing of the β-globin gene. We find that kinetic competition results in multiple competing pathways for pre-mRNA splicing. Splicing of the terminal intron occurs stochastically both before and after transcript release, indicating there is not a strict quality control checkpoint. The majority of pre-mRNAs are spliced after release, while diffusing away from the site of transcription. A single missense point mutation (S34F) in the essential splicing factor U2AF1 which occurs in human cancers perturbs this kinetic balance and defers splicing to occur entirely post-release.

No MeSH data available.


Related in: MedlinePlus

Correlation functions with several gene copies at the TS reflect single-transcript kinetics.The same principles as shown on Figure 3 can be applied to a case where several independent genes are at the transcription site (TS). The contributions of pairs of transcripts from distinct genes (inter-gene) behave like inter-transcript correlations in Figure 3B and hence distribute uniformly on the delay axis and simply average out.DOI:http://dx.doi.org/10.7554/eLife.03939.017
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fig3s1: Correlation functions with several gene copies at the TS reflect single-transcript kinetics.The same principles as shown on Figure 3 can be applied to a case where several independent genes are at the transcription site (TS). The contributions of pairs of transcripts from distinct genes (inter-gene) behave like inter-transcript correlations in Figure 3B and hence distribute uniformly on the delay axis and simply average out.DOI:http://dx.doi.org/10.7554/eLife.03939.017

Mentions: Finally, we emphasize that an essential advantage of this approach is that correlation functions reveal single-transcript kinetics even from signals where multiple transcripts are present at any given time (Supplementary file 1—§1). As illustrated in Figure 3, a single transcription and splicing event results in correlation functions with a peak near zero delay, reflecting the intra-transcript kinetics (Figure 3A). If three transcripts are present, additional peaks appear at non-zero delay, due to inter-transcript kinetics but all the correlations resulting from intra-transcript kinetics accumulate around 0 (Figure 3B). After averaging over many transcription and splicing events, inter-transcript correlations disappear, leaving only a central peak which reflects the kinetics of single transcripts (Figure 3C). See also Video 4 and Figure 3—figure supplement 1.10.7554/eLife.03939.016Figure 3.Correlation functions reflect single-transcript kinetics.


Kinetic competition during the transcription cycle results in stochastic RNA processing.

Coulon A, Ferguson ML, de Turris V, Palangat M, Chow CC, Larson DR - Elife (2014)

Correlation functions with several gene copies at the TS reflect single-transcript kinetics.The same principles as shown on Figure 3 can be applied to a case where several independent genes are at the transcription site (TS). The contributions of pairs of transcripts from distinct genes (inter-gene) behave like inter-transcript correlations in Figure 3B and hence distribute uniformly on the delay axis and simply average out.DOI:http://dx.doi.org/10.7554/eLife.03939.017
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4210818&req=5

fig3s1: Correlation functions with several gene copies at the TS reflect single-transcript kinetics.The same principles as shown on Figure 3 can be applied to a case where several independent genes are at the transcription site (TS). The contributions of pairs of transcripts from distinct genes (inter-gene) behave like inter-transcript correlations in Figure 3B and hence distribute uniformly on the delay axis and simply average out.DOI:http://dx.doi.org/10.7554/eLife.03939.017
Mentions: Finally, we emphasize that an essential advantage of this approach is that correlation functions reveal single-transcript kinetics even from signals where multiple transcripts are present at any given time (Supplementary file 1—§1). As illustrated in Figure 3, a single transcription and splicing event results in correlation functions with a peak near zero delay, reflecting the intra-transcript kinetics (Figure 3A). If three transcripts are present, additional peaks appear at non-zero delay, due to inter-transcript kinetics but all the correlations resulting from intra-transcript kinetics accumulate around 0 (Figure 3B). After averaging over many transcription and splicing events, inter-transcript correlations disappear, leaving only a central peak which reflects the kinetics of single transcripts (Figure 3C). See also Video 4 and Figure 3—figure supplement 1.10.7554/eLife.03939.016Figure 3.Correlation functions reflect single-transcript kinetics.

Bottom Line: We find that kinetic competition results in multiple competing pathways for pre-mRNA splicing.Splicing of the terminal intron occurs stochastically both before and after transcript release, indicating there is not a strict quality control checkpoint.The majority of pre-mRNAs are spliced after release, while diffusing away from the site of transcription.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Biological Modeling, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, United States.

ABSTRACT
Synthesis of mRNA in eukaryotes involves the coordinated action of many enzymatic processes, including initiation, elongation, splicing, and cleavage. Kinetic competition between these processes has been proposed to determine RNA fate, yet such coupling has never been observed in vivo on single transcripts. In this study, we use dual-color single-molecule RNA imaging in living human cells to construct a complete kinetic profile of transcription and splicing of the β-globin gene. We find that kinetic competition results in multiple competing pathways for pre-mRNA splicing. Splicing of the terminal intron occurs stochastically both before and after transcript release, indicating there is not a strict quality control checkpoint. The majority of pre-mRNAs are spliced after release, while diffusing away from the site of transcription. A single missense point mutation (S34F) in the essential splicing factor U2AF1 which occurs in human cancers perturbs this kinetic balance and defers splicing to occur entirely post-release.

No MeSH data available.


Related in: MedlinePlus