Transcript Genomic approaches to improve functional milk

ELF3 and FOXA2 Loci, Probable Putative Trans-QTL
for Abnormal Sperm Percentage in Cattle: An Insilico Analysis
Suneel Kumar Onteru, Varij Nayan & Dheer Singh
Animal Biochemistry Division
National Dairy Research Institute
Karnal-132001
Outline
• A Goal for Animal Genomics
• Need for In-Silico Trans QTL Identification
• Methodology
• Results
• Conclusion & Future Directions
A Goal of Farm Animal Genomics
“To select the farm animals for better Semen
quality traits by genetic markers”
Low to moderate
heritability
traits
be improved by
❉Requirements
for the
selection
of can
animals
assisted
selection
① Variability in marker
reproductive
traits
(Different fertility rate)
② Heritabilities (0.15-0.301)
③ QTLs or chromosomal regions that explain the variation in
phenotypes
④ Breeding strategies
⑤ Evaluation of selection response
1. Druet et al., 2009
Identified QTL for Semen Quality Traits in Cattle
Trait
No. of QTL
Chromosome
Percentage of abnormal sperm (ABSP)
3
20, 21, 27
Percentage of live sperm after osmotic stress
1
23
Percentage of livesperm after thawing
2
23, 27
Sperm average path velocity
1
25
Sperm counts
3
4,11, 27
Sperm motility
6
4,7, 11, 27
Need to identify Further possible
QTL
&
4
4, 11, 15, 22
Sperm acrosome
integrity rate
1
2 QTL
Functional
annotation of genes
in the
Semen Volume
A total of only 21 QTL in AnimalQTLdb
www.animalgenome.org/cgi-bin/QTLdb/BT/summary
Methodology: In-Silico Analysis for Cattle ABSP
Reported QTL (n = 3)
from AnimalQTLdb
Get gene name if
> 80% homology
Extraction of chromosomal coordinates
for the QTL from Cattle tracks from
UMD3.1 Genome
NCBI BLAST for the Protein/
nucleotide sequence
Extraction of annotated genes
from Ensembl Cattle UMD 3.1
genome
NO
Gene names known
YES
Total Gene List (n = 134)
Functional annotation
by Panther 9.0 based
on cattle database
Single Site Analysis (SSA) by oPPOSUM 3.0
for common orthologous transcription factors (oTF)
based on human genome (Z score > 10; Fisher score > 7)
Putative regulative (Trans) QTL based on significant oTF gene co-ordinates in cattle genome
ABSP: Percentage of Abnormal Sperm
Results: Molecular Functions of Genes at reported QTL for
Cattle ABSP
1%
36.6%
32.7%
No. of Genes
5.0%
6.9%
6.9%
5.0%
4.0%
Category
6.9%
Results: Protein Class of Genes at reported QTL for Cattle
ABSP
No. of Genes
1.0%
4.0%
2.0%
3.0%
5.0%
8.9%
3.0%
14.9%
2.0%
1.0%
4.0%
3.0%
2.0%
7.9%
7.9%
8.9%
1.0%
6.9%
2.0%
4.0%
Category
5.0%
15.8%
1.0%
Results: Biological Functions of Genes at Reported QTL for
Cattle ABSP
3.0%
5.9%
16.8%
4.0%
No. of Genes
41.6%
13.9%
11.9%
13.9%
52.50
%
2.0%
Category
9.9%
8.9%
Results: Cellular Locations of Genes at reported QTL for
Cattle ABSP
7.9%
3.0%
No. of Genes
4.0%
2.0%
2.0%
4.0%
Category
Results: Pathway Analysis of Genes at reported QTL for
Cattle ABSP
1.0%
No. of Genes
2.0%
1.0%
4.0%
1.0%
Category
Results: Orthologous Transcription Factors for Genes at
Reported QTL for Cattle ABSP
A total of 178 TF were predicted to be regulating at least 2
genes within 5000 bp up- or down-stream of a gene.
TF
Target Target
Backgrou Backgro
gene gene
nd gene und
hits
non hits hits
gene
non hits
ELF3 66
FOXA2 45
Target
TFBS
hits
Target
TFBS
nucleoti
de rate
Backgro
und
TFBS
hits
Backgro Z-Score Fisher
und TFBS
Score
nucleoti
de rate
33
12278
12474
246
0.0186
40428
0.0146
13.93 7.684
54
6534
18218
94
0.00931
14127
0.0067
13.42 10.23
Results: Functional Role of ELF3
Cellular Location
•
•
•
•
Nucleus
Nucleolus
Cytoplasm
Golgi Apparatus
Molecular Function
• Sequence-specific DNA Binding
Transcription Factor Activity
• Transcription Coactivator Activity
• Protein Binding
• Sequence-specific DNA Binding
Biological Function
• Blastocyst Development
• Transcription From RNA Polymerase II
Promoter
• Inflammatory Response
• Multicellular Organismal Development
• Epidermis Development
• Extracellular Matrix Organization
• Epithelial Cell Differentiation
• Negative Regulation Of Transcription, DNAtemplated
• Positive Regulation Of Transcription, DNAtemplated
• Positive Regulation Of Transcription From
RNA Polymerase II Promoter
• Mammary Gland Involution
First in-slilico report for possible role of
ELF3 in abnormal sperm percentage in
cattle
http://severus.dbmi.pitt.edu/wiki-pi/index.php/gene/view/3170
Results: Functional Role of FOXA2
Cellular Location
• Nucleus
• Transcription
Factor Complex
• Cytoplasm
Molecular Function
• RNA Polymerase II Core Promoter Proximal
Region Sequence-specific DNA Binding
• DNA Binding
• Double-stranded DNA Binding
• Sequence-specific DNA Binding Transcription
Factor Activity
• RNA Polymerase II Distal Enhancer Sequencespecific DNA Binding Transcription Factor
Activity
• Transcription Factor Binding
• DNA Binding, Bending
• Protein Domain Specific Binding
• Transcription Regulatory Region DNA Binding
Biological Function
• In Utero Embryonic
Development
• Ectoderm Formation
• Regulation Of Transcription
From RNA Polymerase II
Promoter
• Pattern Specification Process
• Adult Locomotory Behavior
• Regulation Of Blood
Coagulation
• Glucose Homeostasis
First in-slilico report for possible role of
FOXA2 in abnormal sperm percentage in
cattle
http://severus.dbmi.pitt.edu/wiki-pi/index.php/gene/view/3170
Results: Orthologous Transcription Factors for Genes at
Reported QTL for Cattle ABSP
Earlier reported QTL for ABSP were on Chr 20, 21 and 27
Conclusions
❶Many genes in the reported QTL for ABSP are involved in cellular (42),
metabolic (53), binding (37) and catalytic (33) activities.
❷Pathway analysis found that 25 genes are involved in 21 pathways
with a major contribution of GnRH pathway members (follistatin,
PTGER4, ISL1 and MAP3K1)
❸ELF3 (Chr 16 at 70.8 Mb) and FOXA2 (Chr 13 at 41.9 Mb) loci could be
probable putative trans QTL for abnormal sperm percentage in cattle by
In-silico analysis.
Future Directions
“Association studies have to be conducted for abnormal
sperm percentage by using the genetic markers located at
ELF3 and FOXA2 gene loci to confirm their possible
association with this ABSP in cattle.”
Thank you!!