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Thousands of novel translated open reading frames in humans inferred by ribosome footprint profiling

View Article: PubMed Central - PubMed

ABSTRACT

Accurate annotation of protein coding regions is essential for understanding how genetic information is translated into function. We describe riboHMM, a new method that uses ribosome footprint data to accurately infer translated sequences. Applying riboHMM to human lymphoblastoid cell lines, we identified 7273 novel coding sequences, including 2442 translated upstream open reading frames. We observed an enrichment of footprints at inferred initiation sites after drug-induced arrest of translation initiation, validating many of the novel coding sequences. The novel proteins exhibit significant selective constraint in the inferred reading frames, suggesting that many are functional. Moreover, ~40% of bicistronic transcripts showed negative correlation in the translation levels of their two coding sequences, suggesting a potential regulatory role for these novel regions. Despite known limitations of mass spectrometry to detect protein expressed at low level, we estimated a 14% validation rate. Our work significantly expands the set of known coding regions in humans.

Doi:: http://dx.doi.org/10.7554/eLife.13328.001

No MeSH data available.


Related in: MedlinePlus

Decision rules to identify novel mCDS.Illustrating the decisions that identify novel mCDS. The mCDS that do not overlap any known CDS (pooling GENCODE, UCSC and CCDS annotations) are labeled as novel mCDS -- these include mCDS from both novel and annotated transcripts.DOI:http://dx.doi.org/10.7554/eLife.13328.011
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fig2s1: Decision rules to identify novel mCDS.Illustrating the decisions that identify novel mCDS. The mCDS that do not overlap any known CDS (pooling GENCODE, UCSC and CCDS annotations) are labeled as novel mCDS -- these include mCDS from both novel and annotated transcripts.DOI:http://dx.doi.org/10.7554/eLife.13328.011

Mentions: Thus, in summary, it is likely that most of the 814 mCDS that were identified within GENCODE annotated protein-coding transcripts and have a distinct reading frame compared to GENCODE annotations are indeed novel alternate translated sequences. To ensure that an mCDS is truly novel, we verified that it does not overlap any known CDS annotated by GENCODE, UCSC (Rosenbloom et al., 2015), or CCDS (Farrell et al., 2014) in the same frame. (See Figure 2 for the different classes of LCL transcripts that contain a novel mCDS; Figure 2—figure supplement 1 illustrates the decision rules used to identify a novel mCDS). Among these 814 novel mCDS, 386 mCDS overlap an annotated CDS but have a different reading frame (labeled ‘dual-coding’) and 156 do not overlap the annotated CDS. An example of a novel dual-coding region – an mRNA sequence that codes for proteins in two different frames – inferred in the POLR2M gene is illustrated in Figure 3A. Using tandem mass-spectrometry data (Battle et al., 2015), we found four unique spectra matching peptides in the mCDS and no spectra matching peptides in the annotated CDS (protein level posterior error probability = 3 × 10−35. However, our simulations suggest that most, or all, of the 39% of genes where riboHMM infers the annotated reading frame but disagrees with the annotated start site are false discoveries, and these are not considered further here.10.7554/eLife.13328.010Figure 2.Overview of novel coding sequences.


Thousands of novel translated open reading frames in humans inferred by ribosome footprint profiling
Decision rules to identify novel mCDS.Illustrating the decisions that identify novel mCDS. The mCDS that do not overlap any known CDS (pooling GENCODE, UCSC and CCDS annotations) are labeled as novel mCDS -- these include mCDS from both novel and annotated transcripts.DOI:http://dx.doi.org/10.7554/eLife.13328.011
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4940163&req=5

fig2s1: Decision rules to identify novel mCDS.Illustrating the decisions that identify novel mCDS. The mCDS that do not overlap any known CDS (pooling GENCODE, UCSC and CCDS annotations) are labeled as novel mCDS -- these include mCDS from both novel and annotated transcripts.DOI:http://dx.doi.org/10.7554/eLife.13328.011
Mentions: Thus, in summary, it is likely that most of the 814 mCDS that were identified within GENCODE annotated protein-coding transcripts and have a distinct reading frame compared to GENCODE annotations are indeed novel alternate translated sequences. To ensure that an mCDS is truly novel, we verified that it does not overlap any known CDS annotated by GENCODE, UCSC (Rosenbloom et al., 2015), or CCDS (Farrell et al., 2014) in the same frame. (See Figure 2 for the different classes of LCL transcripts that contain a novel mCDS; Figure 2—figure supplement 1 illustrates the decision rules used to identify a novel mCDS). Among these 814 novel mCDS, 386 mCDS overlap an annotated CDS but have a different reading frame (labeled ‘dual-coding’) and 156 do not overlap the annotated CDS. An example of a novel dual-coding region – an mRNA sequence that codes for proteins in two different frames – inferred in the POLR2M gene is illustrated in Figure 3A. Using tandem mass-spectrometry data (Battle et al., 2015), we found four unique spectra matching peptides in the mCDS and no spectra matching peptides in the annotated CDS (protein level posterior error probability = 3 × 10−35. However, our simulations suggest that most, or all, of the 39% of genes where riboHMM infers the annotated reading frame but disagrees with the annotated start site are false discoveries, and these are not considered further here.10.7554/eLife.13328.010Figure 2.Overview of novel coding sequences.

View Article: PubMed Central - PubMed

ABSTRACT

Accurate annotation of protein coding regions is essential for understanding how genetic information is translated into function. We describe riboHMM, a new method that uses ribosome footprint data to accurately infer translated sequences. Applying riboHMM to human lymphoblastoid cell lines, we identified 7273 novel coding sequences, including 2442 translated upstream open reading frames. We observed an enrichment of footprints at inferred initiation sites after drug-induced arrest of translation initiation, validating many of the novel coding sequences. The novel proteins exhibit significant selective constraint in the inferred reading frames, suggesting that many are functional. Moreover, ~40% of bicistronic transcripts showed negative correlation in the translation levels of their two coding sequences, suggesting a potential regulatory role for these novel regions. Despite known limitations of mass spectrometry to detect protein expressed at low level, we estimated a 14% validation rate. Our work significantly expands the set of known coding regions in humans.

Doi:: http://dx.doi.org/10.7554/eLife.13328.001

No MeSH data available.


Related in: MedlinePlus