Defining the Sequence Elements and Candidate Genes for the Coloboma Mutation

Abstract

The chicken coloboma mutation exhibits features similar to human congenital developmental malformations such as ocular coloboma, cleft-palate, dwarfism, and polydactyly. The coloboma-associated region and encoded genes were investigated using advanced genomic, genetic, and gene expression technologies. Initially, the mutation was linked to a 990 kb region encoding 11 genes; the application of the genetic and genomic tools led to a reduction of the linked region to 176 kb and the elimination of 7 genes. Furthermore, bioinformatics analyses of capture array-next generation sequence data identified genetic elements including SNPs, insertions, deletions, gaps, chromosomal rearrangements, and miRNA binding sites within the introgressed causative region relative to the reference genome sequence. Coloboma-specific variants within exons, UTRs, and splice sites were studied for their contribution to the mutant phenotype. Our compiled results suggest three genes for future studies. The three candidate genes, SLC30A5 (a zinc transporter), CENPH (a centromere protein), and CDK7 (a cyclin-dependent kinase), are differentially expressed (compared to normal embryos) at stages and in tissues affected by the coloboma mutation. Of these genes, two (SLC30A5 and CENPH) are considered high-priority candidate based upon studies in other vertebrate model systems.

The chick as an animal model of eye disease

“…The mutation has been mapped to a relatively short section of 176 kb on chromosome X. Three candidate genes (SLC30A5, CENPH and CDK7) have been identified on the basis of this location and differential expression compared with the normal embryos [24].…”

Two Proximally Close Priority Candidate Genes for diplopodia-1, an Autosomal Inherited Craniofacial-Limb Syndrome in the Chicken: MRE11 and GPR83

“…Full details of the array setup and methods can be found in Robb and Delany (2012b). (Note that the same array was utilized to sequence 2 other UCD-congenic developmental mutations, coloboma [co.003] and wingless-2 [wg-2.331]; Robb and Delany 2012b; Robb et al 2013; Webb et al 2018.) The other 2 congenic lines, which mapped to different chromosomes (GGA Z and 12), were utilized as controls for GGA 1 comparisons (Robb et al 2011; Robb 2012; Robb and Delany 2012b).…”

“…The discrimination of non-causative polymorphic elements (naturally occurring in the introgressed region) from the potentially causative polymorphic elements was accomplished by a comparative approach. Polymorphisms were identified by comparing the Dp-1.003 causative (sequenced) region to variants identified in other chicken genetic lines including: the other inbred congenic lines ( wg-2 , Webb et al 2018, and co , Robb et al 2013) utilized on the capture array, the chicken reference genome (UCD-001), or any other previously reported polymorphism (e.g., within NCBI [ex. dbSNP] and the UCSC genome browser).…”

“…Variability in clefting (mild through severe) is also observed in UCD-Dp-1.003. Note that dp-1 mutant embryos displaying severe clefting, a phenotype not as common but yet observed over the course of this study, were not available on the day of photography and it was deemed inappropriate/unnecessary to set additional eggs for observations when the severe phenotype representation is available from other lines (e.g., coloboma—see Robb et al 2013) at same age (i.e., in terms of reducing the usage of animals).…”

The chick eye in vision research: An excellent model for the study of ocular disease

“…The clinical characteristics of these chicks are similar to those seen in human coloboma cases (Hocking and Guggenheim, 2014). Recent progress has been made toward identification of the responsible mutation, which is lethal for female embryos (Robb et al, 2013). Three candidate genes have been identified as targets for further investigation.…”