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Revealing Missing Human Protein Isoforms Based on Ab Initio Prediction, RNA-seq and Proteomics.

Hu Z, Scott HS, Qin G, Zheng G, Chu X, Xie L, Adelson DL, Oftedal BE, Venugopal P, Babic M, Hahn CN, Zhang B, Wang X, Li N, Wei C - Sci Rep (2015)

Bottom Line: Based on these novel transcripts, at least 36 novel proteins were detected from shotgun proteomics data of 41 breast samples.We also showed L1 retrotransposons have a more significant impact on the origin of new transcripts/genes than previously thought.In the end, the total number of human transcripts with protein-coding potential was estimated to be at least 204,950.

View Article: PubMed Central - PubMed

Affiliation: 1] School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, 800 Dongchuan Road, Shanghai 200240, China [2] Shanghai Center for Bioinformation Technology, 1278 Keyuan Road, Pudong District, Shanghai 201203, China.

ABSTRACT
Biological and biomedical research relies on comprehensive understanding of protein-coding transcripts. However, the total number of human proteins is still unknown due to the prevalence of alternative splicing. In this paper, we detected 31,566 novel transcripts with coding potential by filtering our ab initio predictions with 50 RNA-seq datasets from diverse tissues/cell lines. PCR followed by MiSeq sequencing showed that at least 84.1% of these predicted novel splice sites could be validated. In contrast to known transcripts, the expression of these novel transcripts were highly tissue-specific. Based on these novel transcripts, at least 36 novel proteins were detected from shotgun proteomics data of 41 breast samples. We also showed L1 retrotransposons have a more significant impact on the origin of new transcripts/genes than previously thought. Furthermore, we found that alternative splicing is extraordinarily widespread for genes involved in specific biological functions like protein binding, nucleoside binding, neuron projection, membrane organization and cell adhesion. In the end, the total number of human transcripts with protein-coding potential was estimated to be at least 204,950.

No MeSH data available.


Related in: MedlinePlus

A diagrammatic representation of transcript prediction using ALTSCAN and validation pipeline based on RNA-seq datasets.The upper part shows the pipeline of alternative transcript prediction and the MIXTURE dataset construction. The lower part shows the pipeline for transcript validation with RNA-seq data. The grey blocks indicate raw public data. Candidate gene regions were extracted from various public annotations and then ASs were predicted by ALTSCAN for these regions. Together with the well-annotated KNOWN transcripts, ALTSCAN transcripts were validated with a large number of RNA-seq data. TC is short for transcript coverage and JC is short for junction coverage. The NIJ (novel internal junction) filter was used to check if novel internal junction(s) existed in transcripts (Figure S3). The novel transcript datasets VHC, VMC and VLC were defined as in the figure.
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f1: A diagrammatic representation of transcript prediction using ALTSCAN and validation pipeline based on RNA-seq datasets.The upper part shows the pipeline of alternative transcript prediction and the MIXTURE dataset construction. The lower part shows the pipeline for transcript validation with RNA-seq data. The grey blocks indicate raw public data. Candidate gene regions were extracted from various public annotations and then ASs were predicted by ALTSCAN for these regions. Together with the well-annotated KNOWN transcripts, ALTSCAN transcripts were validated with a large number of RNA-seq data. TC is short for transcript coverage and JC is short for junction coverage. The NIJ (novel internal junction) filter was used to check if novel internal junction(s) existed in transcripts (Figure S3). The novel transcript datasets VHC, VMC and VLC were defined as in the figure.

Mentions: ALTSCAN was developed by extending Viterbi algorithm to predict the most probable N paths (transcripts) for each gene region from the genomic sequence only (see Methods and Figure S1 for details) and applied to human genome sequences (upper part of Fig. 1). As a result, 320,784 transcripts with complete ORFs from 33,945 loci were predicted. Among them, 298,454 transcripts were from 22,606 loci in GENCODE or Refseq gene regions; 8,331 transcripts were from 2,721 loci overlapped with pseudogenes; and almost all remaining transcripts located in repeat-rich regions. Notably, 9,682 transcripts from 7,663 loci overlapped more than 50% (of each transcript) with L1 elements.


Revealing Missing Human Protein Isoforms Based on Ab Initio Prediction, RNA-seq and Proteomics.

Hu Z, Scott HS, Qin G, Zheng G, Chu X, Xie L, Adelson DL, Oftedal BE, Venugopal P, Babic M, Hahn CN, Zhang B, Wang X, Li N, Wei C - Sci Rep (2015)

A diagrammatic representation of transcript prediction using ALTSCAN and validation pipeline based on RNA-seq datasets.The upper part shows the pipeline of alternative transcript prediction and the MIXTURE dataset construction. The lower part shows the pipeline for transcript validation with RNA-seq data. The grey blocks indicate raw public data. Candidate gene regions were extracted from various public annotations and then ASs were predicted by ALTSCAN for these regions. Together with the well-annotated KNOWN transcripts, ALTSCAN transcripts were validated with a large number of RNA-seq data. TC is short for transcript coverage and JC is short for junction coverage. The NIJ (novel internal junction) filter was used to check if novel internal junction(s) existed in transcripts (Figure S3). The novel transcript datasets VHC, VMC and VLC were defined as in the figure.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4496727&req=5

f1: A diagrammatic representation of transcript prediction using ALTSCAN and validation pipeline based on RNA-seq datasets.The upper part shows the pipeline of alternative transcript prediction and the MIXTURE dataset construction. The lower part shows the pipeline for transcript validation with RNA-seq data. The grey blocks indicate raw public data. Candidate gene regions were extracted from various public annotations and then ASs were predicted by ALTSCAN for these regions. Together with the well-annotated KNOWN transcripts, ALTSCAN transcripts were validated with a large number of RNA-seq data. TC is short for transcript coverage and JC is short for junction coverage. The NIJ (novel internal junction) filter was used to check if novel internal junction(s) existed in transcripts (Figure S3). The novel transcript datasets VHC, VMC and VLC were defined as in the figure.
Mentions: ALTSCAN was developed by extending Viterbi algorithm to predict the most probable N paths (transcripts) for each gene region from the genomic sequence only (see Methods and Figure S1 for details) and applied to human genome sequences (upper part of Fig. 1). As a result, 320,784 transcripts with complete ORFs from 33,945 loci were predicted. Among them, 298,454 transcripts were from 22,606 loci in GENCODE or Refseq gene regions; 8,331 transcripts were from 2,721 loci overlapped with pseudogenes; and almost all remaining transcripts located in repeat-rich regions. Notably, 9,682 transcripts from 7,663 loci overlapped more than 50% (of each transcript) with L1 elements.

Bottom Line: Based on these novel transcripts, at least 36 novel proteins were detected from shotgun proteomics data of 41 breast samples.We also showed L1 retrotransposons have a more significant impact on the origin of new transcripts/genes than previously thought.In the end, the total number of human transcripts with protein-coding potential was estimated to be at least 204,950.

View Article: PubMed Central - PubMed

Affiliation: 1] School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, 800 Dongchuan Road, Shanghai 200240, China [2] Shanghai Center for Bioinformation Technology, 1278 Keyuan Road, Pudong District, Shanghai 201203, China.

ABSTRACT
Biological and biomedical research relies on comprehensive understanding of protein-coding transcripts. However, the total number of human proteins is still unknown due to the prevalence of alternative splicing. In this paper, we detected 31,566 novel transcripts with coding potential by filtering our ab initio predictions with 50 RNA-seq datasets from diverse tissues/cell lines. PCR followed by MiSeq sequencing showed that at least 84.1% of these predicted novel splice sites could be validated. In contrast to known transcripts, the expression of these novel transcripts were highly tissue-specific. Based on these novel transcripts, at least 36 novel proteins were detected from shotgun proteomics data of 41 breast samples. We also showed L1 retrotransposons have a more significant impact on the origin of new transcripts/genes than previously thought. Furthermore, we found that alternative splicing is extraordinarily widespread for genes involved in specific biological functions like protein binding, nucleoside binding, neuron projection, membrane organization and cell adhesion. In the end, the total number of human transcripts with protein-coding potential was estimated to be at least 204,950.

No MeSH data available.


Related in: MedlinePlus