ELF3and. FOXA2Loci, Probable Putative Trans-QTL for Abnormal Sperm Percentage in Cattle: An In- silicoanalysis

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1 ELF3and FOXA2Loci, Probable Putative Trans-QTL for Abnormal Sperm Percentage in Cattle: An In- silicoanalysis Suneel Kumar Onteru, Varij Nayan & Dheer Singh Animal Biochemistry Division National Dairy Research Institute Karnal

2 Outline oal for Animal Genomics ed for In-SilicoTrans QTL Identification thodology sults nclusion & Future Directions

3 A Goal of Farm Animal Genomics To select the farm animals for better Semen quality traits by genetic markers quirements Low to moderate for the heritability selection traits of can animals be improved by Variability in marker reproductive assisted traits selection (Different fertility rate) Heritabilities ( ) QTLs or chromosomal regions that explain the variation in phenotypes Breeding strategies Evaluation of selection response t et al., 2009

4 Identified QTL for Semen Quality Traits in Cattle No. of QTL Chromosom entage of abnormal sperm (ABSP) 3 20, 21, 2 ntage of live sperm after osmotic stress 1 23 ntage of livesperm after thawing 2 23, 27 Need to identify Further possible QTL & nvolume 4 4, 11, 15, 22 Functional annotation of genes in the QTL acrosome integrity rate 1 2 average path velocity 1 25 counts 3 4,11, 27 motility 6 4,7, 11, 27 A total of only 21 QTL in AnimalQTLdb

5 Methodology: In-Silico Analysis for Cattle ABSP d QTL (n = 3) nimalqtldb Extraction of chromosomal coordinates for the QTL from Cattle tracks from UMD3.1 Genome Extraction of annotated from Ensembl Cattle UM genome ne name if homology NCBI BLAST for the Protein/ nucleotide sequence Total Gene List (n = 134) NO Gene names known YES ctional annotation Panther 9.0 based n cattle database Single Site Analysis (SSA) by opposum 3.0 for common orthologous transcription factors (o based on human genome (Z score > 10; Fisher scor ative regulative (Trans) QTL based on significant otf gene co-ordinates in cattle genome ABSP: Percentage of Abnormal Sperm

6 ults: Molecular Functions of Genes at reported QT Cattle ABSP 36.6% 1% 32.7% 5.0% 6.9% 5.0% 4.0% Category 6.9%

7 sults: Protein Class of Genes at reported QTL for Ca ABSP 1.0% 4.0% 2.0% 3.0%5.0% Category 14.9% 2.0% 1.0% 8.9% 3.0% 3.0% 2.0% 7.9% 7.9% 8.9% 1.0% 6.9% 2.0% 4.0% 5.0% 4.0% 15.8%

8 ults: Biological Functions of Genes at Reported QTL Cattle ABSP 3.0% 5.9% 16.8% Category 41.6% 13.9% 11.9% 13.9% % 8.9% 2.0% 9.9%

9 sults: Cellular Locations of Genes at reported QTL Cattle ABSP 7.9% 3.0% 4.0% 2.0% 2.0% Category 4.0%

10 sults: Pathway Analysis of Genes at reported QTL f Cattle ABSP 2.0% 1.0% 4.0% Category

11 sults: Orthologous Transcription Factors for Genes 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. Target gene hits Target gene non hits Backgrou nd gene hits Backgro und gene non hits Target TFBS hits Target TFBS nucleoti de rate Backgro und TFBS hits Backgro und TFBS nucleoti de rate Z-Score

12 Results: Functional Role of ELF3 r Location Molecular Function Biological Function Blastocyst Development Transcription From RNA Polymerase II Promoter Inflammatory Response Multicellular Organismal Developmen us Sequence-specific DNA Binding olus Transcription Factor Activity lasm Transcription Coactivator Activity Apparatus Protein Binding Sequence-specific DNA Binding ELF3 in abnormal sperm percentage Epidermis Developmentin First in-slilico report for possible role of cattle Extracellular Matrix Organization Epithelial Cell Differentiation Negative Regulation Of Transcription, templated Positive Regulation Of Transcription, D templated Positive Regulation Of Transcription F RNA Polymerase II Promoter Mammary Gland Involution

13 s iption Complex sm Results: Functional Role of FOXA2 r Location Molecular Function Biological Functio 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 cattle Enhancer Sequencespecific DNA Binding Transcription Factor Activity Transcription Factor Binding DNA Binding, Bending Protein Domain Specific Binding Transcription Regulatory Region DNA Binding In Utero Embryonic Development Ectoderm Formation Regulation Of Transcript From RNA Polymerase II Promoter Pattern Specification Pro Adult Locomotory Beha Regulation Of Blood Coagulation Glucose Homeostasis First in-slilico report for possible role of FOXA2 in abnormal sperm percentage in

14 sults: Orthologous Transcription Factors for Genes Reported QTL for Cattle ABSP Earlier reported QTL for ABSP were on Chr 20, 21 and 27

15 Conclusions any genes in the reported QTL for ABSP are involved in cellular (42 etabolic(53), binding(37) and catalytic(33) activities. Pathway analysis found that 25 genes are involved in 21 pathway ith a major contribution of GnRH pathway members (follistati TGER4, ISL1 and MAP3K1) ELF3 (Chr 16 at 70.8 Mb) and FOXA2 (Chr 13 at 41.9 Mb) loci could probable putative trans QTL for abnormal sperm percentage in cattl In-silico analysis.

16 Future Directions Association studies have to be conducted for abnorma sperm percentage by using the genetic markers located a ELF3 and FOXA2 gene loci to confirm their possibl association with this ABSP in cattle.

17 Thank you!!