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Tunable protein synthesis by transcript isoforms in human cells.

Floor SN, Doudna JA - Elife (2016)

Bottom Line: However, the relationship between human transcript diversity and protein production is complex as each isoform can be translated differently.We fractionated a polysome profile and reconstructed transcript isoforms from each fraction, which we term Transcript Isoforms in Polysomes sequencing (TrIP-seq).Select 5' untranslated regions exert robust translational control between cell lines, while 3' untranslated regions can confer cell type-specific expression.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States.

ABSTRACT
Eukaryotic genes generate multiple RNA transcript isoforms though alternative transcription, splicing, and polyadenylation. However, the relationship between human transcript diversity and protein production is complex as each isoform can be translated differently. We fractionated a polysome profile and reconstructed transcript isoforms from each fraction, which we term Transcript Isoforms in Polysomes sequencing (TrIP-seq). Analysis of these data revealed regulatory features that control ribosome occupancy and translational output of each transcript isoform. We extracted a panel of 5' and 3' untranslated regions that control protein production from an unrelated gene in cells over a 100-fold range. Select 5' untranslated regions exert robust translational control between cell lines, while 3' untranslated regions can confer cell type-specific expression. These results expose the large dynamic range of transcript-isoform-specific translational control, identify isoform-specific sequences that control protein output in human cells, and demonstrate that transcript isoform diversity must be considered when relating RNA and protein levels.

No MeSH data available.


Clustering of human preimplantation embryo data(A) Hierarchical clustering of 45,895 transcript isoforms across human preimplantation embryogenesis with the stages indicated below. Yellow: above isoform average, cyan: below isoform average.DOI:http://dx.doi.org/10.7554/eLife.10921.015
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fig4s1: Clustering of human preimplantation embryo data(A) Hierarchical clustering of 45,895 transcript isoforms across human preimplantation embryogenesis with the stages indicated below. Yellow: above isoform average, cyan: below isoform average.DOI:http://dx.doi.org/10.7554/eLife.10921.015

Mentions: (A) For each embryonic stage, transcripts are mapped onto all eight TrIP-seq clusters, as in Figure 2. The percentage of transcripts mapping to each cluster per stage is then calculated, and compared to the percentage for TrIP-seq data. hESC – human embryonic stem cell; p0, p10 – passage zero or passage ten. Error is s.d. between single cells at each embryonic stage or between TrIP-seq biological replicates. (B) Expression levels for two transcripts of CSDE1 across embryonic development and polysome fractions demonstrating a switch in translational status. Error is S.D. TPM – transcripts per million. (C) Diagram of two transcript isoforms of CSDE1. Regions different between the two isoforms are in yellow and select shared intronic regions have been shortened for clarity. See also Figure 4—figure supplement 1.


Tunable protein synthesis by transcript isoforms in human cells.

Floor SN, Doudna JA - Elife (2016)

Clustering of human preimplantation embryo data(A) Hierarchical clustering of 45,895 transcript isoforms across human preimplantation embryogenesis with the stages indicated below. Yellow: above isoform average, cyan: below isoform average.DOI:http://dx.doi.org/10.7554/eLife.10921.015
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4764583&req=5

fig4s1: Clustering of human preimplantation embryo data(A) Hierarchical clustering of 45,895 transcript isoforms across human preimplantation embryogenesis with the stages indicated below. Yellow: above isoform average, cyan: below isoform average.DOI:http://dx.doi.org/10.7554/eLife.10921.015
Mentions: (A) For each embryonic stage, transcripts are mapped onto all eight TrIP-seq clusters, as in Figure 2. The percentage of transcripts mapping to each cluster per stage is then calculated, and compared to the percentage for TrIP-seq data. hESC – human embryonic stem cell; p0, p10 – passage zero or passage ten. Error is s.d. between single cells at each embryonic stage or between TrIP-seq biological replicates. (B) Expression levels for two transcripts of CSDE1 across embryonic development and polysome fractions demonstrating a switch in translational status. Error is S.D. TPM – transcripts per million. (C) Diagram of two transcript isoforms of CSDE1. Regions different between the two isoforms are in yellow and select shared intronic regions have been shortened for clarity. See also Figure 4—figure supplement 1.

Bottom Line: However, the relationship between human transcript diversity and protein production is complex as each isoform can be translated differently.We fractionated a polysome profile and reconstructed transcript isoforms from each fraction, which we term Transcript Isoforms in Polysomes sequencing (TrIP-seq).Select 5' untranslated regions exert robust translational control between cell lines, while 3' untranslated regions can confer cell type-specific expression.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States.

ABSTRACT
Eukaryotic genes generate multiple RNA transcript isoforms though alternative transcription, splicing, and polyadenylation. However, the relationship between human transcript diversity and protein production is complex as each isoform can be translated differently. We fractionated a polysome profile and reconstructed transcript isoforms from each fraction, which we term Transcript Isoforms in Polysomes sequencing (TrIP-seq). Analysis of these data revealed regulatory features that control ribosome occupancy and translational output of each transcript isoform. We extracted a panel of 5' and 3' untranslated regions that control protein production from an unrelated gene in cells over a 100-fold range. Select 5' untranslated regions exert robust translational control between cell lines, while 3' untranslated regions can confer cell type-specific expression. These results expose the large dynamic range of transcript-isoform-specific translational control, identify isoform-specific sequences that control protein output in human cells, and demonstrate that transcript isoform diversity must be considered when relating RNA and protein levels.

No MeSH data available.