Identification of genetic variants associated with navigation ability using analysis of hippocampus transcriptome data in racing homer and rock dove pigeons

Introduction: The racing homer pigeon is one of the most popular ecotypes that have a remarkable navigation ability, compared to the Rock dove pigeon, which has an undesirable navigation ability. Therefore, in this research, these two ecotypes were used for comparative genomics and identification of genetic variants associated with navigation traits and flight ability. Functional examination of related variants and genes can help identify putative genes associated with navigation mechanisms and elaboration of the relevant biological pathways. On the other hand, these pigeons can be used as an animal model to investigate biological processes related to memory and learning ability also identifying key genes and biological processes related to memory and learning processes. This study aimed to identify the genetic variants related to the navigation ability in two ecotypes of pigeons.
Materials and methods: Transcriptome data (eight samples) related to the hippocampus of Rock dove pigeons (weak routing ability) and Racing homer (good routing and long-distance homing ability) were downloaded from the EBI database with the accession number PRJNA532674. The reference genome and all available annotations related to pigeons were downloaded from the Ensembl database (ftp://ftp.ensembl.org). Data quality control was performed based on read length, GC content, unread bases, duplicate reads, and Phred index. To identify the variants, the fixed ploidy algorithm, and the following parameters were used. The score of the base variant was 30, the nighber base score of the variant was 20, and the frequency of the base variant was set at 30%. Quality control of reads, mapping, and identification of different genetic variants between Rock and Homing groups were performed with CLC Genomics workbench software. The functional analysis of different variants was performed by gene ontology analysis by the VEP tool in the Ensembl database with a significance level of 5%. Gene ontology analysis was performed at three levels: molecular function (MF), biological processes (BP), and cellular components (CC).
Results and discussion: Around two million genetic variants, were identified for the eight samples (944325 for Racing homer and 1055277 for Rock dove pigeon), of which 2800 different genetic variants were reported between the two groups. The variants were classified into four categories including single nucleotide polymorphisms (71.1%), multiple nucleotide variants (16.8%), insertions (6.5%), and short deletions (5.5%) for Racing homer pigeons. The frequencies of corresponding variants for the Rock dove pigeon were 73.2%, 12.5%, 7.5%, and 68%, respectively. All reads showed an alignment of above 90% with the reference genome which indicated a good alignment. The results of the gene ontology analysis showed that different variants are related to the LDHA gene. This gene is important in the flight power and anaerobic metabolism of cells by coding the lactate dehydrogenase A enzyme. Birds with a mutated version of the LDHA gene perform better in competitions for flying power and navigation. Another finding of this study was the association of genetic variants with the DRD4 gene, which is important in controlling behavioral patterns, memory, and learning. Protein metabolism pathways were also one of the important results of this research, due to their essential role in the differentiation of nervous system cells. For example, the Ubiquitin protein acts as a messenger for proteases by marking other proteins and facilitates the digestion of ineffective proteins in the cell. Also, this protein has a regulatory role in the activity of other proteins. One of the results of gene ontology analysis at the molecular function level was related to protein Serine/threonine kinase activity, which confirms the results of biological process analysis. Also, the PIK3C2A candidate gene was suggested for this molecular effector. This gene, by encoding Phosphatidylinositol 4-phosphate 3-kinase C2 domain-containing subunit alpha, plays a fundamental role in regulating cell activity such as the transfer of glucose to plasma, the secretion of insulin and the response to UV or other stresses.
Conclusions: The results of this study confirmed that the homing pigeon and rock pigeon ecotypes had different genetic potential in terms of navigation ability. Differences in routing ability between two studied ecotypes should be related to mutations and changes in genomic regions during evolution. However, the identified genetic variants must be confirmed under laboratory conditions using PCR or other techniques. The identified variants can be used as a model for further research into human diseases and the identification of paralog genes related to human memory and learning ability.