Project description:The aim of this study is to discover new genes and cells involved in life-long tooth replacement. Here we study the adult dentition of the leopard gecko (Eublepharis macularius). Bulk RNAseq was used to compare teeth that are in function versus unerupted, developing teeth and single cell RNA-seq was carried out on jaw segments containing the dental forming tissues. In bulk RNAseq data, we found that functional teeth expressed genes involved in bone and tooth resorption. Indeed, we found expression of these markers in multinucleated odontoclasts within resorbing functional teeth. Chemotaxis genes SEMA3A and SEMA3E, were expressed within odontoblasts and in adjacent mesenchyme using RNAscope. Semaphorins could be involved in regulating odontoclast formation, recruitment or repulsion from developing teeth. The scRNA-seq experiment successfully isolated dental mesenchyme and several epithelial clusters. We confirmed that some of these genes are expressed in the earliest tooth buds within the tooth forming field and in erupting teeth. This work will lead to discovery of genes and cell populations that may have been gained or lost during evolution of amniotes. Moreover, gene differences may lead to dental therapies to prevent tooth loss from disease or injury.
Project description:The aim of this study is to discover new genes and cells involved in life-long tooth replacement. Here we study the adult dentition of the leopard gecko (Eublepharis macularius). Bulk RNAseq was used to compare teeth that are in function versus unerupted, developing teeth and single cell RNA-seq was carried out on jaw segments containing the dental forming tissues. In bulk RNAseq data, we found that functional teeth expressed genes involved in bone and tooth resorption. Indeed, we found expression of these markers in multinucleated odontoclasts within resorbing functional teeth. Chemotaxis genes SEMA3A and SEMA3E, were expressed within odontoblasts and in adjacent mesenchyme using RNAscope. Semaphorins could be involved in regulating odontoclast formation, recruitment or repulsion from developing teeth. The scRNA-seq experiment successfully isolated dental mesenchyme and several epithelial clusters. We confirmed that some of these genes are expressed in the earliest tooth buds within the tooth forming field and in erupting teeth. This work will lead to discovery of genes and cell populations that may have been gained or lost during evolution of amniotes. Moreover, gene differences may lead to dental therapies to prevent tooth loss from disease or injury.
Project description:BACKGROUND:Geckos are among the most species-rich reptile groups and the sister clade to all other lizards and snakes. Geckos possess a suite of distinctive characteristics, including adhesive digits, nocturnal activity, hard, calcareous eggshells, and a lack of eyelids. However, one gecko clade, the Eublepharidae, appears to be the exception to most of these 'rules' and lacks adhesive toe pads, has eyelids, and lays eggs with soft, leathery eggshells. These differences make eublepharids an important component of any investigation into the underlying genomic innovations contributing to the distinctive phenotypes in 'typical' geckos. FINDINGS:We report high-depth genome sequencing, assembly, and annotation for a male leopard gecko, Eublepharis macularius (Eublepharidae). Illumina sequence data were generated from seven insert libraries (ranging from 170 to 20 kb), representing a raw sequencing depth of 136X from 303 Gb of data, reduced to 84X and 187 Gb after filtering. The assembled genome of 2.02 Gb was close to the 2.23 Gb estimated by k-mer analysis. Scaffold and contig N50 sizes of 664 and 20 kb, respectively, were comparable to the previously published Gekko japonicus genome. Repetitive elements accounted for 42 % of the genome. Gene annotation yielded 24,755 protein-coding genes, of which 93 % were functionally annotated. CEGMA and BUSCO assessment showed that our assembly captured 91 % (225 of 248) of the core eukaryotic genes, and 76 % of vertebrate universal single-copy orthologs. CONCLUSIONS:Assembly of the leopard gecko genome provides a valuable resource for future comparative genomic studies of geckos and other squamate reptiles.