TY - JOUR
T1 - MS/MS in silico subtraction-based proteomic profiling as an approach to facilitate disease gene discovery
T2 - application to lens development and cataract
AU - Aryal, Sandeep
AU - Anand, Deepti
AU - Hernandez, Francisco G.
AU - Weatherbee, Bailey A.T.
AU - Huang, Hongzhan
AU - Reddy, Ashok P.
AU - Wilmarth, Phillip A.
AU - David, Larry L.
AU - Lachke, Salil A.
N1 - Funding Information:
The authors thank Drs. Melinda Duncan and Velia Fowler for helpful discussions. This work was supported by National Institutes of Health/National Eye Institute [R01 EY021505 to S.L.]. Support from the University of Delaware Core Imaging Facility and Proteomics and Mass Spectrometry Facility was made possible through the Institutional Development Award (IDeA) from the National Institutes of Health/National Institute of General Medical Sciences INBRE Program Grant [Grant Number P20 GM103446]. Acquisition of the confocal microscope used in this study was funded by the National Institutes of Health/National Center for Research Resources Grant [S10 RR027273]. Mass spectrometric analysis was performed by the OHSU Proteomics Shared Resource with partial support from NIH core Grants P30 EY010572 and P30 CA069533 and shared instrument Grant S10OD-012246. S.A. was supported by a Fight for Sight Summer Student Fellowship and Sigma Xi award.
Funding Information:
The authors thank Drs. Melinda Duncan and Velia Fowler for helpful discussions. This work was supported by National Institutes of Health/National Eye Institute [R01 EY021505 to S.L.]. Support from the University of Delaware Core Imaging Facility and Proteomics and Mass Spectrometry Facility was made possible through the Institutional Development Award (IDeA) from the National Institutes of Health/National Institute of General Medical Sciences INBRE Program Grant [Grant Number P20 GM103446]. Acquisition of the confocal microscope used in this study was funded by the National Institutes of Health/National Center for Research Resources Grant [S10 RR027273]. Mass spectrometric analysis was performed by the OHSU Proteomics Shared Resource with partial support from NIH core Grants P30 EY010572 and P30 CA069533 and shared instrument Grant S10OD-012246. S.A. was supported by a Fight for Sight Summer Student Fellowship and Sigma Xi award.
Publisher Copyright:
© 2019, Springer-Verlag GmbH Germany, part of Springer Nature.
PY - 2020/2/1
Y1 - 2020/2/1
N2 - While the bioinformatics resource-tool iSyTE (integrated Systems Tool for Eye gene discovery) effectively identifies human cataract-associated genes, it is currently based on just transcriptome data, and thus, it is necessary to include protein-level information to gain greater confidence in gene prioritization. Here, we expand iSyTE through development of a novel proteome-based resource on the lens and demonstrate its utility in cataract gene discovery. We applied high-throughput tandem mass spectrometry (MS/MS) to generate a global protein expression profile of mouse lens at embryonic day (E)14.5, which identified 2371 lens-expressed proteins. A major challenge of high-throughput expression profiling is identification of high-priority candidates among the thousands of expressed proteins. To address this problem, we generated new MS/MS proteome data on mouse whole embryonic body (WB). WB proteome was then used as a reference dataset for performing “in silico WB-subtraction” comparative analysis with the lens proteome, which effectively identified 422 proteins with lens-enriched expression at ≥ 2.5 average spectral counts, ≥ 2.0 fold enrichment (FDR < 0.01) cut-off. These top 20% candidates represent a rich pool of high-priority proteins in the lens including known human cataract-linked genes and many new potential regulators of lens development and homeostasis. This rich information is made publicly accessible through iSyTE (https://research.bioinformatics.udel.edu/iSyTE/), which enables user-friendly visualization of promising candidates, thus making iSyTE a comprehensive tool for cataract gene discovery.
AB - While the bioinformatics resource-tool iSyTE (integrated Systems Tool for Eye gene discovery) effectively identifies human cataract-associated genes, it is currently based on just transcriptome data, and thus, it is necessary to include protein-level information to gain greater confidence in gene prioritization. Here, we expand iSyTE through development of a novel proteome-based resource on the lens and demonstrate its utility in cataract gene discovery. We applied high-throughput tandem mass spectrometry (MS/MS) to generate a global protein expression profile of mouse lens at embryonic day (E)14.5, which identified 2371 lens-expressed proteins. A major challenge of high-throughput expression profiling is identification of high-priority candidates among the thousands of expressed proteins. To address this problem, we generated new MS/MS proteome data on mouse whole embryonic body (WB). WB proteome was then used as a reference dataset for performing “in silico WB-subtraction” comparative analysis with the lens proteome, which effectively identified 422 proteins with lens-enriched expression at ≥ 2.5 average spectral counts, ≥ 2.0 fold enrichment (FDR < 0.01) cut-off. These top 20% candidates represent a rich pool of high-priority proteins in the lens including known human cataract-linked genes and many new potential regulators of lens development and homeostasis. This rich information is made publicly accessible through iSyTE (https://research.bioinformatics.udel.edu/iSyTE/), which enables user-friendly visualization of promising candidates, thus making iSyTE a comprehensive tool for cataract gene discovery.
UR - http://www.scopus.com/inward/record.url?scp=85075922723&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85075922723&partnerID=8YFLogxK
U2 - 10.1007/s00439-019-02095-5
DO - 10.1007/s00439-019-02095-5
M3 - Article
C2 - 31797049
AN - SCOPUS:85075922723
SN - 0340-6717
VL - 139
SP - 151
EP - 184
JO - Human Genetics
JF - Human Genetics
IS - 2
ER -