DEVELOPMENT OF INTEGRATED CATTLE GENOMICS KNOWLEDGE BASE

Systems biology approaches being applied to animal breeding represent an opportunity to derive greater benefits from animal production systems. The increasingly detailed investigations in systems biology have led to a large amount of data dispersed over various sources; therefore, a centralized knowledge base is in demand. In this study, we have integrated cattle genomics data of heterogeneous sources and types and developed a bioinformatics tool to study genotype-phenotype associations in cattle: http://integromics-time.com/integromics-database/. The tool enables revealing genomic overlaps within trait-associated loci and identification of potential functional candidates. It might be also used as a tool for planning genotype– phenotype research in cattle

Pivotal role of the muscle-contraction pathway in cryptorchidism and evidence for genomic connections with cardiomyopathy pathways in RASopathies

BACKGROUND: Cryptorchidism is the most frequent congenital disorder in male children; however the genetic causes of cryptorchidism remain poorly investigated. Comparative integratomics combined with systems biology approach was employed to elucidate genetic factors and molecular pathways underlying testis descent. METHODS: Literature mining was performed to collect genomic loci associated with cryptorchidism in seven mammalian species. Information regarding the collected candidate genes was stored in MySQL relational database. Genomic view of the loci was presented using Flash GViewer web tool (http://gmod.org/wiki/Flashgviewer/). DAVID Bioinformatics Resources 6.7 was used for pathway enrichment analysis. Cytoscape plug-in PiNGO 1.11 was employed for protein-network-based prediction of novel candidate genes. Relevant protein-protein interactions were confirmed and visualized using the STRING database (version 9.0). RESULTS: The developed cryptorchidism gene atlas includes 217 candidate loci (genes, regions involved in chromosomal mutations, and copy number variations) identified at the genomic, transcriptomic, and proteomic level. Human orthologs of the collected candidate loci were presented using a genomic map viewer. The cryptorchidism gene atlas is freely available online: http://www.integratomics-time.com/cryptorchidism/. Pathway analysis suggested the presence of twelve enriched pathways associated with the list of 179 literature-derived candidate genes. Additionally, a list of 43 network-predicted novel candidate genes was significantly associated with four enriched pathways. Joint pathway analysis of the collected and predicted candidate genes revealed the pivotal importance of the muscle-contraction pathway in cryptorchidism and evidence for genomic associations with cardiomyopathy pathways in RASopathies. CONCLUSIONS: The developed gene atlas represents an important resource for the scientific community researching genetics of cryptorchidism. The collected data will further facilitate development of novel genetic markers and could be of interest for functional studies in animals and human. The proposed network-based systems biology approach elucidates molecular mechanisms underlying co-presence of cryptorchidism and cardiomyopathy in RASopathies. Such approach could also aid in molecular explanation of co-presence of diverse and apparently unrelated clinical manifestations in other syndromes

Genome-wide screening for genetic variants in polyadenylation signal (PAS) sites in mouse selection lines for fatness and leanness

Alternative polyadenylation (APA) determines mRNA stability, localisation, translation and protein function. Several diseases, including obesity, have been linked to APA. Studies have shown that single nucleotide polymorphisms in polyadenylation signals (PAS-SNPs) can influence APA and affect phenotype and disease susceptibility. However, these studies focussed on associations between single PAS-SNP alleles with very large effects and phenotype. Therefore, we performed a genome-wide screening for PAS-SNPs in the polygenic mouse selection lines for fatness and leanness by whole-genome sequencing. The genetic variants identified in the two lines were overlapped with locations of PAS sites obtained from the PolyASite 2.0 database. Expression data for selected genes were extracted from the microarray expression experiment performed on multiple tissue samples. In total, 682 PAS-SNPs were identified within 583 genes involved in various biological processes, including transport, protein modifications and degradation, cell adhesion and immune response. Moreover, 63 of the 583 orthologous genes in human have been previously associated with human diseases, such as nervous system and physical disorders, and immune, endocrine, and metabolic diseases. In both lines, PAS-SNPs have also been identified in genes broadly involved in APA, such as Polr2c, Eif3e and Ints11. Five PAS-SNPs within 5 genes (Car, Col4a1, Itga7, Lat, Nmnat1) were prioritised as potential functional variants and could contribute to the phenotypic disparity between the two selection lines. The developed PAS-SNPs catalogue presents a key resource for planning functional studies to uncover the role of PAS-SNPs in APA, disease susceptibility and fat deposition. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s00335-022-09967-8

Molecular Mechanisms of Syndromic Cryptorchidism: Data Synthesis of 50 Studies and Visualization of Gene-Disease Network

Background: Cryptorchidism is one of the most frequent congenital birth defects in male children and is present in 2–4% of full-term male births. It has several possible health effects including reduced fertility, increased risk for testicular neoplasia, testicular torsion, and psychological consequences. Cryptorchidism is often diagnosed as comorbid; copresent with other diseases. It is also present in clinical picture of several syndromes. However, this field has not been systematically studied. The aim of the present study was to catalog published cases of syndromes which include cryptorchidism in the clinical picture and associated genomic information.Methods: The literature was extracted from Public/Publisher MEDLINE and Web of Science databases, using the keywords including: syndrome, cryptorchidism, undescended testes, loci, and gene. The obtained data was organized in a table according to the previously proposed standardized data format. The results of the study were visually represented using Gephi and karyotype view.Results: Fifty publications had sufficient data for analysis. Literature analysis resulted in 60 genomic loci, associated with 44 syndromes that have cryptorchidism in clinical picture. Genomic loci included 38 protein-coding genes and 22 structural variations containing microdeletions and microduplications. Loci, associated with syndromic cryptorchidism are located on 16 chromosomes. Visualization of retrieved data is presented in a gene-disease network.Conclusions: The study is ongoing and further studies will be needed to develop a complete catalog with the data from upcoming publications. Additional studies will also be needed for revealing of molecular mechanisms associated with syndromic cryptorchidism and revealing complete diseasome network

Transcriptomic signatures for human male infertility

Introduction: Male infertility is a common, complex disorder. A better understanding of pathogenesis and etiology is needed for timely diagnosis and treatment. The aim of this study, therefore, was to identify genes involved in the pathogenesis of idiopathic male infertility based on data from transcriptomic level supported with data from genomic level.Materials and methods: First, we performed whole gene expression analysis in 20 testis biopsy samples of patients with severely impaired (10) and normal spermatogenesis (10). Further, we have performed systematic review of comparable male infertility studies and overlapped the most significantly expressed genes identified in our study with the most differentially expressed genes from selected studies. Gene Ontology analysis and KEGG functional enrichment have been performed with Enrichr analysis tool. Additionally, we have overlapped these genes with the genes where rare variants have been identified previously.Results: In 10 patients with severely impaired spermatogenesis and 10 controls, we identified more than 1,800 differentially expressed genes (p < 0.001). With the systematic review of three previously performed microarray studies that have met inclusion criteria we identified 257 overlapped differentialy expressed genes (144 downregulated and 113 upregulated). Intersection of genes from transcriptomic studies with genes with identified rare variants revealed a total of 7 genes linked with male infertility phenotype (CYP11A1, CYP17A1, RSPH3, TSGA10, AKAP4, CCIN, NDNF).Conclusion: Our comprehensive study highlighted the role of four genes in pathogenesis of male infertility and provided supporting evidence for three promising candidate genes which dysfunction may result in a male infertility disorder

New Research Strategies in Lactation Biology

Different approaches have been used to study milk related traits in farm animals, reaching from statistical dissection of phenotypic variation to the search of candidate genes with major phenotypic effects. The aim of this study was to develop a new research tool devoted to in vitro studies of physiological pathways responsible for mammary gland development, lactation, remodeling and immune response, supported by a user friendly map based bioinformatics tool for integration of different types of data. We established goat mammary gland derived primary epithelial cell line with predominantly epithelial morphology, responsive to lactogenic hormones and exhibiting regeneration potential in heterologous mouse system. The response of primary epithelial cells to pathogenic bacterium Mycoplasma agalactiae was studied using RNA sequencing approach and 1553 differentially expressed genes were detected 24h post infection. The majority of differentially expressed genes belonged to cell cycle regulating genes, pro-inflammatory cytokines, chemokines and genes involved in lipid metabolism. Bioinformatic analysis of 359 putative target sites for mammary gland expressed miRNAs revealed polymorphic miRNA target sites for bta-miR-199b, -199a-5p, and -361 in the IL1B gene and for -miR-126 in the CYP11B1 gene. Graphical integration of different types of data to DairyVis platform allowed identification of genomic regions with higher number of potential functional elements that deserve further experimental analysis. The newly developed MEC line and integration of bioinformatics tools into DairyVis database represent a promising methodological support for further research in the field of lactation biology

Vascular endothelial growth factor (VEGF)-related polymorphisms rs10738760 and rs6921438 are not risk factors for proliferative diabetic retinopathy (PDR) in patients with type 2 diabetes mellitus (T2DM)

Vascular endothelial growth factor (VEGF) is an important regulator of angiogenesis and has been investigated as a candidate gene in a number of conditions, including diabetes and its microvascular complications (e.g., retinopathy and nephropathy). Several VEGF-related polymorphisms have been shown to contribute to nearly half of the variability in circulating VEGF levels in healthy individuals. Our aim was to assess the association between VEGF-related rs10738760 and rs6921438 polymorphisms and proliferative diabetic retinopathy (PDR) in Slovenian patients with type 2 diabetes mellitus (T2DM). We also investigated the effect of these polymorphisms on VEGF receptor 2 (VEGFR-2) expression in fibrovascular membranes (FVMs) from patients with PDR. This case-control study enrolled 505 unrelated patients with T2DM: 143 diabetic patients with PDR as a study group, and 362 patients with T2DM of >10 years duration and with no clinical signs of PDR as a control group. Patient clinical and laboratory data were obtained from their medical records. rs10738760 and rs6921438 polymorphisms were genotyped using TaqMan SNP Genotyping assay. VEGFR-2 expression was assessed by immunohistochemistry in 20 FVMs from patients with PDR, and numerical areal density of VEGFR-2-positive cells was calculated. The occurrence of PDR was 1.7 times higher in diabetic patients carrying GA genotype of rs6921438 compared to patients with GG genotype, with a borderline statistical significance (OR = 1.7, 95% CI = 1.00 – 2.86, p = 0.05). In addition, A allele of rs6921438 was associated with increased VEGFR-2 expression in FVMs from PDR patients. However, we observed no association between AA genotype of rs6921438 nor between rs10738760 variants and PDR, indicating that the two polymorphisms are not genetic risk factors for PDR

Use of SNP Markers Within the Fat Mass and Obesity-associated (FTO) Gene to Verify Pedigrees and Determine Haplotypes in Paternal Half-sib Families of Slovenian Simmental Cattle

The objective of this preliminary study was to identify SNP markers within the FTO gene for evaluation of pedigree data accuracy and determination of haplotypes in paternal half-sib families of Slovenian Simmental cattle. Out of 23 polymorphic SNPs identified ten most informative SNPs for genotyping 31 sires and 56 half-sib progeny were used. The ATLAS program was used for paternity testing. Haplotype analysis revealed three haplotype blocks. The effect of SNPs “ex2 T>C” and “int2 indel*>T” was significant on three correlated carcass traits: live weight at slaughter (P= 0.03), carcass weight (P= 0.038), and lean weight (P= 0.048). The FTO gene can thus be regarded as a candidate for the marker assisted selection programs in our and possibly other populations of cattle. Future studies in cattle might also reveal novel roles of the FTO gene in carcass traits on livestock species as well as fatness control in other mammals