Characterizing the normal proteome of
human optic nerve
Presenter
Roopna Raveendran
Ph.D. Student
YU-IOB Center for Systems Biology and
Molecular Medicine
Guide
Dr. T. S. Keshava Prasad
Professor and Deputy Director
YU-IOB Center for Systems Biology and
Molecular Medicine
December 08, 2016
1
FUNDING AGENCY
Ministry of Science and Technology
Department of Science and Technology
WOMEN SCIENTISTS SCHEME (WOS-A)
2
DETAILS
Principle Investigator:
Dr. Sneha M. Pinto
DST-INSPIRE faculty and Assistant Professor
YU-IOB Center for Systems Biology and Molecular
Medicine
Co- investigator:
Roopna Raveendran
Ph.D. Student
YU-IOB Center for Systems Biology and Molecular
Medicine
3
CONTENT
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Introduction
Literature review
Lacunae observed
Aims and objectives
Methodology
Expected outcome
Budget
Timeline
References
4
INTRODUCTION
5
http://www.wisegeek.com/what-is-optic-nerve-regeneration.htm
OPTIC NERVE
• Second cranial nerve, lied at the back
of the eye
• Made up of nerve cells and consists
of over one million nerve fibers
• Connects the eye to the brain
• Carries the impulses formed by the
retina to the brain, which interprets
them as images
https://12cranialnerves.wordpress.com/cranial-nerve-2-optic-nerve/
LITERATURE REVIEW
Published article
Eye tissue
A comprehensive proteomics analysis of the human iris tissue: ready Iris
to embrace postgenomics precision medicine in ophthalmology?,
Murthy et al., OMICS: A Journal of Integrative Biology, 2016
Oct;16(19):2592-2596
Proteomics of human aqueous humor, Murthy et al., OMICS: A
Journal of Integrative Biology, 2015 May;19(5):283-93
Aqueous Humor
A draft map of the human proteome, Kim et al., Nature, 2014 May
29;509(7502):575-81
Retina
Proteomic analysis of human vitreous humor, Murthy et al., Clinical
Proteomics, 2014 Jul 14;11(1):29
Vitreous humor
Characterizing the normal proteome of human ciliary body, Goel et
al., Clinical Proteomics, 2013 Aug 1;10(1):9
Ciliary body
7
The draft map of the human proteome
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Kim et al., Nature, 2014
LACUNAE OBSERVED
• Till date there is no reference proteome dataset present
• The comprehensive dataset in this study will serve as
the basis for pursuing biomarkers for diseases
pertaining to human optic nerve
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AIM
To better understand the molecular mechanisms
behind the ocular diseases
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OBJECTIVES
•
•
Specific objective 1: To carry out proteomic profiling of
optic nerve
Specific objective 2: To carry out proteogenomic
analysis of human optic nerve
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IMPORTANCE OF THE WORK
• Proteomic investigations can provide insights into the
signaling pathways operating in the ocular disorders,
further help to better understand and diagnosis of ocular
diseases
STUDY DESIGN
• Ethical clearance
The study will be conducted after obtaining the
approval from the institutional ethical committee. The
informed consent form from the donors will be obtained.
• Inclusion criteria for subject’s recruitment
- Both male and female donors
- No medical history of glaucoma, other eye diseases or
malignancy
- Enucleation within 3–4 hours post mortem
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STUDY DESIGN
• Exclusion criteria for subject’s recruitment
- Enucleation not more than 3–4 hours post mortem
- With any eye abnormality will be excluded
• Clinical data collection
Samples for the proteomic analysis will be obtained at
autopsy after obtaining approval from institutional review
board of Vittala International Institute of Ophthalmology,
Bangalore
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METHODOLOGY
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PROTEOGENOMICS WORKFLOW
Renuse et al., Proteomics, 2011
Peptide identification using proteogenomics
VSTPNVSVR
DLLLQWCWENK
ECDVVSNTIIK
Customized
sequence DB
MS/MS spectra
Map peptides to the genome
Intergenic region
Protein-coding gene
Pseudogene
Novel protein-coding
locus
Exon 1
Exon 2
Exon 3
intron
3’ UTR
5’ UTR
Translational
start
Alt splicing
Translational
stop
Alexey I Nesvizhskii, Nature, 2014
Peptide identification using proteogenomics
Exon 1
Exon 2
intron
3’ UTR
Exon 1
Known
Exon 3
5’ UTR
Exon 2
Exon 3
Exon
Junction
--------------
--------------
5’ UTR
Novel
3’ UTR
Alt junction
v
----------------------------------------
Intron
Alt frame
Alexey I Nesvizhskii, Nature, 2014
PROTEOGENOMIC ANALYSIS
• Identification of several protein-coding regions/genes in
the human genome that have not been reported
previously
• Correction of the existing gene model
• Confirmation of the existing gene model
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Comparison with the human proteome
• 17,294 nonredundant proteins identified by Kim et al.
from 30 histologically normal human samples
• To identify proteins uniquely reported in this study,
comparison of the obtained protein list will be carried out
with those identified by Kim et .al
• Further, their role in the regulation of mechanism in optic
nerve will be studied based by carrying out gene
ontology and literature review
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EXPECTED OUTCOME
• Comprehensive proteome reference set for optic nerve
• Improved genome annotation
• Baseline to compare against proteome of ocular
disorders and for pursuing biomarkers for diseases
pertaining to human eye
BUDGET
S.NO.
1ST YEAR
2ND YEAR
3RD YEAR
TOTAL
(INR)
(INR)
(INR)
(INR)
HEAD
1.
Research fellowship
3,60,000
3,60,000
3,60,000
10,80,000
2.
Consumables
1,90,000
95,000
50,000
3,35,000
3.
Instrument
2,00,000
55,000
55,000
3,10,000
4.
Travel (within India)
10,000
10,000
10,000
30,000
5.
Contingencies
20,000
20,000
20,000
60,000
6.
Overhead charges
78,000
54,000
49,500
1,81,500
7.
GRAND TOTAL (INR)
8,58,000
5,94,000
5,44,500
19,96,500
22
TIMELINE
1ST YEAR
Work
1-6 months
7-12
months
2ND YEAR
13-18
months
19-24
months
3RD YEAR
25-30
months
31-36
months
Review of literature
Standardization of protocol
Ethical clearance
Objective 1
Objective 2
Publication and thesis
submission
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REFERENCES
• A comprehensive proteomics analysis of the human iris tissue:
ready to embrace postgenomics precision medicine in
ophthalmology?, Murthy et al., OMICS: A Journal of Integrative
Biology, 2016 Oct;16(19):2592-2596
• Murthy et al., Proteomics of human aqueous humor. OMICS: A
Journal of Integrative Biology. 2015 May;19(5):283-93
• Kim et al., A draft map of the human proteome. Nature. 2014 May
29;509(7502):575-81
• Murthy et al., Proteomic analysis of human vitreous humor. Clinical
Proteomics. 2014 Jul 14;11(1):29
• Goel et al., Characterizing the normal proteome of human ciliary
body. Clinical Proteomics. 2013 Aug 1;10(1):9
24
THANK YOU
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