Abstract
Defining protein-protein interactions (PPIs) is central to the biological sciences. Here, we present a novel platform - Affinity Capture of Polyribosomes followed by RNA sequencing (ACAPseq) - for identifying PPIs. ACAPseq harnesses the power of massively parallel RNA sequencing (RNAseq) to quantify the enrichment of polyribosomes based on the affinity of their associated nascent polypeptides for an immobilized protein 'bait'. This method was developed and tested using neonatal mouse brain polyribosomes and a variety of extracellular domains as baits. Of 92 baits tested, 25 identified one or more binding partners that appear to be biologically relevant; additional candidate partners remain to be validated. ACAPseq can detect binding to targets that are present at less than 1 part in 100,000 in the starting polyribosome preparation. One of the observed PPIs was analyzed in detail, revealing the mode of homophilic binding for Protocadherin-9 (PCDH9), a non-clustered Protocadherin family member.
| Original language | English (US) |
|---|---|
| Journal | eLife |
| Volume | 7 |
| DOIs | |
| State | Published - Oct 22 2018 |
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Keywords
- biochemistry
- chemical biology
- extracellular interactome
- mouse
- protocadherin
- RNAseq
- secretome
ASJC Scopus subject areas
- Neuroscience(all)
- Biochemistry, Genetics and Molecular Biology(all)
- Immunology and Microbiology(all)
Cite this
Affinity capture of polyribosomes followed by RNAseq (ACAPseq), a discovery platform for protein-protein interactions. / Peng, Xi; Emiliani, Francesco; Smallwood, Philip M.; Rattner, Amir; Lei, Hong; Sabbagh, Mark F.; Nathans, Jeremy.
In: eLife, Vol. 7, 22.10.2018.Research output: Contribution to journal › Article
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TY - JOUR
T1 - Affinity capture of polyribosomes followed by RNAseq (ACAPseq), a discovery platform for protein-protein interactions
AU - Peng, Xi
AU - Emiliani, Francesco
AU - Smallwood, Philip M.
AU - Rattner, Amir
AU - Lei, Hong
AU - Sabbagh, Mark F.
AU - Nathans, Jeremy
PY - 2018/10/22
Y1 - 2018/10/22
N2 - Defining protein-protein interactions (PPIs) is central to the biological sciences. Here, we present a novel platform - Affinity Capture of Polyribosomes followed by RNA sequencing (ACAPseq) - for identifying PPIs. ACAPseq harnesses the power of massively parallel RNA sequencing (RNAseq) to quantify the enrichment of polyribosomes based on the affinity of their associated nascent polypeptides for an immobilized protein 'bait'. This method was developed and tested using neonatal mouse brain polyribosomes and a variety of extracellular domains as baits. Of 92 baits tested, 25 identified one or more binding partners that appear to be biologically relevant; additional candidate partners remain to be validated. ACAPseq can detect binding to targets that are present at less than 1 part in 100,000 in the starting polyribosome preparation. One of the observed PPIs was analyzed in detail, revealing the mode of homophilic binding for Protocadherin-9 (PCDH9), a non-clustered Protocadherin family member.
AB - Defining protein-protein interactions (PPIs) is central to the biological sciences. Here, we present a novel platform - Affinity Capture of Polyribosomes followed by RNA sequencing (ACAPseq) - for identifying PPIs. ACAPseq harnesses the power of massively parallel RNA sequencing (RNAseq) to quantify the enrichment of polyribosomes based on the affinity of their associated nascent polypeptides for an immobilized protein 'bait'. This method was developed and tested using neonatal mouse brain polyribosomes and a variety of extracellular domains as baits. Of 92 baits tested, 25 identified one or more binding partners that appear to be biologically relevant; additional candidate partners remain to be validated. ACAPseq can detect binding to targets that are present at less than 1 part in 100,000 in the starting polyribosome preparation. One of the observed PPIs was analyzed in detail, revealing the mode of homophilic binding for Protocadherin-9 (PCDH9), a non-clustered Protocadherin family member.
KW - biochemistry
KW - chemical biology
KW - extracellular interactome
KW - mouse
KW - protocadherin
KW - RNAseq
KW - secretome
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UR - http://www.scopus.com/inward/citedby.url?scp=85055071382&partnerID=8YFLogxK
U2 - 10.7554/eLife.40982
DO - 10.7554/eLife.40982
M3 - Article
C2 - 30345971
AN - SCOPUS:85055071382
VL - 7
JO - eLife
JF - eLife
SN - 2050-084X
ER -