In present investigation, six hundred and seventy seven (677) RNA-guided recombinase target sites were identified in bovine genome (Table 1). On mapping these sequences against RefSeq Genome Database of
Bos Taurus (taxid: 9913) for highly similar sequences through Blast, it was found that 247 RGR target sites lie in genic region and 430 in intergenic region. Location and features of important target Sites identified in genic region of bovine genome for hyperactivated recombinase gin with spacer length six has been shown in Table 2.
In present study, whole genome of bovine (
Bos taurus) was analyzed to identify the location of target sites for hyperactivated recombinase Gin with spacer length six. It was found that RGR target sites are located in several important protein coding genes which regulates important biological proteins like Mab-21 domain containing 2, integrin beta-6 isoform X1, fibulin-1 precursor, thyroglobulin precursor, kinesin-like protein KIF22, Raftlin protein, inactive phospholipase C-like protein 1, docking protein 6, growth hormone receptor precursor, lysine-specific demethylase 4C, TRMT1-like protein, GRB2-associated-binding protein 1 isoform X1, cadherin-8 isoform X1, hyaluronan-binding protein 2 precursor, claudin-10 isoform b, nuclear receptor ROR-alpha, neuroblastoma-amplified sequence, protein NDRG1, anoctamin-3, NEDD4-like E3 ubiquitin-protein ligase WWP2, catenin delta-2 isoform X1
etc.
A RGR site in gene MB21D2 coding for Mab-21 domain containing 2 was located on chromosome 1. Olivieri
et al. (2016) studied to identify genomic regions and metabolic pathways associated with dry matter intake, average daily gain, feed efficiency and residual feed intake in an experimental Nellore cattle population and found the association of MB21D2 gene with average daily gain.
An et al. (2018) identified loci and candidate genes for internal organ weights in Simmental beef cattle by genome-wide association study. They found that polymorphism in MB21D2 gene is associated with kidney weight.
A RGR site in gene ITGB6 coding for integrin beta-6 was located on chromosome 2. The encoded macromolecule forms a dimer with alpha v chain and this heterodimer may bind to ligands like fibronectin and transforming growth factor beta one. These can function as receptors for foot-and-mouth disease virus (FMDV) in epithelium.
Berryman et al. (2005) have shown that ‘foot-and-mouth disease virus (FMDV) infection mediated by the integrin takes place through clathrin-dependent endocytosis however not caveolae or alternative endocytic pathways that rely upon lipoid rafts. A RGR site in gene coding for fibulin-1 precursor was located on chromosome 5. Fibulin-1 may play a role in cell adhesion and migration along protein fibers within the extracellular matrix (ECM). It might be vital for development related processes and contribute to the supramolecular organization of basement membranes.
Debber et al. (2002) reported that fibulin-1 gene (FBLN1) is disrupted in a reciprocal chromosomal translocation t (12;22) which is associated with a complex type of synpolydactyly.
A RGR site in gene coding for thyroglobulin precursor was located on chromosome 14. Thyroid hormones regulates metabolism and affect the homeostasis of fat deposition. The sequence coding iodoprotein (TG), producing the precursor for thyroid hormones, has been proposed as a positional and functional candidate gene for a QTL with an effect on fat deposition.
Gan et al. (2008) identified 6 novel SNPs at the 3’ flanking region of the TG gene. The SNP marker association analysis indicated that the SNP markers were significantly associated with marbling score. They prompt that theTG-gene-specific SNP is also a helpful marker for meat quality traits in future marker-assisted selection programmes in
Bos taurus. A RGR site in gene KIF22 encoding kinesin-like protein KIF22 was located on chromosome 25. It is involved in spindle formation and the movements of chromosomes during mitosis and meiosis. It binds to microtubules and to DNA. A RGR site in gene coding for raftlin isoform X3 was located on chromosome 1. Raftlin protein is pivotal for maintenance of lipid rafts and may be involved in regulation of B cell antigen receptor mediated signaling
(Saeki et al., 2003). Raftlin promotes binding of double stranded RNA, activations of B cell receptors and toll like receptor 3 signaling pathways and is involved in IL 17 production to release pro inflammatory cytokines.
Qu et al. (2017) investigated differentially expressed membrane proteins of pulmonary alveolar macrophages infected with highly pathogenic porcine reproductive and respiratory syndrome virus and its attenuated strain by label free quantitative proteomic analysis and found higher abundance of raftlin in the HP PRRSV group compared to the AP PRRSV and control groups which explain the more severe inflammation triggered by HP PRRSV.
A RGR site in gene PLCL1 coding for inactive phospholipase C-like protein 1 was located on chromosome 2. It is involved in an inositol phospholipid-based intracellular signaling cascade. It regulates the turnover of receptors and thus contributes to the maintenance of GABA-mediated synaptic inhibition. Its aberrant expression might contribute to the genesis and progression of respiratory organ malignant neoplastic disease.
Asano et al. (2014) investigated the role of phospholipase C-related catalytically inactive protein (PRIP) in insulin granule exocytosis using Prip-knockdown mouse insulinoma (MIN6) cells and demonstrated that PRIP regulate KIF5B-mediated insulin secretion. It was found that insulin release from Prip-knockdown MIN6 cells was higher than that from control cells.
A RGR site in gene KDM4C coding for lysine-specific demethylase 4C was located on chromosome 8. The encoded super molecule could be a trimethylation-specific demethylase and converts specific trimethylated histone residues to the dimethylated type. This enzymatic action regulates gene expression and chromosome segregation. KDM4C is induced during adipocyte differentiation and depletion of KDM4C is sufficient to block this kind of cellular differentiation
(Lu et al., 2012). KDM4C is amplified or upregulated in several cell lines derived from esophageal squamous carcinomas, medulloblastoma and breast cancer
(Yang et al., 2000, Cloos et al., 2006, Ehrbrecht et al., 2006, Liu et al., 2009 and
Uimonn et al., 2014). In addition, in agreement with a contribution of KDM4C to tumor development, inhibition of KDM4C expression reduces cell proliferation
(Cloos et al., 2006).
A RGR site in gene coding for growth hormone receptor precursor was located on chromosome 20. Growth hormone receptor (GHR) is receptor for pituitary gland growth hormone involved in regulating postnatal body growth. On ligand binding, it activates the JAK2/STAT5 pathway.
Rahmatalla et al. (2011) studied effect of non-synonymous single nucleotide polymorphism (SNP) in exon 8 which leads to a phenylalanine to tyrosine amino acid substitution (F279Y) in the receptor. They predicted the consequences of the F279Y mutation on milk yield, fat, protein, casein, lactose yield and content as well as somatic cell score (SCS), in a German Holstein dairy cattle population. They found that tyrosine variant occurred as the minor allele (16.5%) but its substitution effects were 320 kg (305 d), 0.02 kg per day and 0.07 kg per day for milk, casein and lactose yields, respectively. The same allelomorph had negative effects on fat, protein and casein contents. Besides this, the high-milk-yield tyrosine allele was also associated with lower SCS (p < 0.05). They suggested that F279Y polymorphism possess high potential as a marker for the improvement of milk traits in selection programs.
Qin et al. (2007) identified polymorphism in exon 10 of GHR in three cattle breeds (Nanyang cattle, limousin and galloway) and found correlations between GHR gene polymorphism and body size indexes in cattle. A RGR site in DOK6 gene coding for docking protein 6 was located on chromosome 24. DOK proteins are enzymatically inert scaffolding proteins. They provide a tying up platform for the assembly of multimolecular signal complexes. DOK6 promotes Ret-mediated neurite growth and may affect brain development and maintenance.
RNA-guided recombinase target sites for hyperactivated recombinase beta in bovine genome has been also explored previously in which 436 RGR target sites were identified in bovine genome for recombinase Beta with spacer length five
(Pathak et al., 2019).
