Dio3os allelic expression
Document Type
Oral Presentation
Department
Biological Sciences
Abstract
Abstract Background: Genomic imprinting is an epigenetic event that results in different expression levels for the alleles of a given gene, depending on the parental origin. Alterations in genomic imprinting lead to abnormal expression of imprinted genes and to subsequent developmental, neurological and metabolic disease [1]. Imprinting genes are often found in genomic clusters that feature coordinated mechanisms regulating allelic expression. One such cluster is the Dlk- Dio3 domain. Aberrant imprinting in this domain causes severe pathologies in mice and humans. This domain features Dio3os, a long, non-coding RNA of unknown function located close to paternally-expressed Dio3 and transcribed from the opposite DNA strand. However, the degree and status of Dio3os imprinting have not been defined. Our Aim is to determine the allelic expression of Dio3os across different tissues and investigate whether Dio3os influences the allelic expression of neighboring gene Dio3. Methods: We used novel transgenic mouse models with a deletion of Dio3os (Dio3os-/- mice) or with a MYC tag in the Dio3 gene (Dio3myc/myc mice). We crossed Dio3os-/- males with Dio3myc/myc females and vice versa to distinguish Dio3os and Dio3 alleles in the offspring. We then euthanized the double heterozygous, postnatal day 20 (P20) offspring and harvested tissues with abundant Dio3os expression levels ,including adrenal gland, liver, ovary, uterus, testis and white adipose tissue). We then used standard methods for RNA isolation, cDNA synthesis, and real-time quantitative PCR to determine the allele-specific expression levels of Dio3os and Dio3. Results: We observed that Dio3os expression was ~3-fold higher in the liver of animals with a Dio3os deletion in the paternal allele compared to those with a Dio3os deletion in the maternal allele. No significant difference in Dio3os expression was observed in the adrenal glands and the testes of animals with different parental origin of the Dio3os deletion. Based on qPCR reactions specific for the myc-tagged or untagged Dio3 allele, we observed that the adrenal gland of animals with deletion of Dio3os in the paternal allele showed a 2.5-fold increase in the paternal to maternal allele expression ratio of Dio3, when compared to those with a Dio3os deletion in the maternal allele. Conclusions: Dio3os is an imprinted gene preferentially expressed from the maternal allele in neonatal liver, but its imprinting is tissue-specific and may not be present in other tissues. Dio3os plays an allele-specific regulatory role in the allelic expression of Dio3 in the adrenal gland, a finding that could potentially extend to other tissues. Singer-Sam, J. (2010). When only one allele of a gene is actively transcribed, gene expression is termed monoallelic. What does monoallelic gene expression have to do with cats, twins, and genetic diseases? Nature Education 3(3):1. Retrieved February 26, 2022, from https://www.nature.com/scitable/topicpage/monoallelic-expression-8813275/
Dio3os allelic expression
Abstract Background: Genomic imprinting is an epigenetic event that results in different expression levels for the alleles of a given gene, depending on the parental origin. Alterations in genomic imprinting lead to abnormal expression of imprinted genes and to subsequent developmental, neurological and metabolic disease [1]. Imprinting genes are often found in genomic clusters that feature coordinated mechanisms regulating allelic expression. One such cluster is the Dlk- Dio3 domain. Aberrant imprinting in this domain causes severe pathologies in mice and humans. This domain features Dio3os, a long, non-coding RNA of unknown function located close to paternally-expressed Dio3 and transcribed from the opposite DNA strand. However, the degree and status of Dio3os imprinting have not been defined. Our Aim is to determine the allelic expression of Dio3os across different tissues and investigate whether Dio3os influences the allelic expression of neighboring gene Dio3. Methods: We used novel transgenic mouse models with a deletion of Dio3os (Dio3os-/- mice) or with a MYC tag in the Dio3 gene (Dio3myc/myc mice). We crossed Dio3os-/- males with Dio3myc/myc females and vice versa to distinguish Dio3os and Dio3 alleles in the offspring. We then euthanized the double heterozygous, postnatal day 20 (P20) offspring and harvested tissues with abundant Dio3os expression levels ,including adrenal gland, liver, ovary, uterus, testis and white adipose tissue). We then used standard methods for RNA isolation, cDNA synthesis, and real-time quantitative PCR to determine the allele-specific expression levels of Dio3os and Dio3. Results: We observed that Dio3os expression was ~3-fold higher in the liver of animals with a Dio3os deletion in the paternal allele compared to those with a Dio3os deletion in the maternal allele. No significant difference in Dio3os expression was observed in the adrenal glands and the testes of animals with different parental origin of the Dio3os deletion. Based on qPCR reactions specific for the myc-tagged or untagged Dio3 allele, we observed that the adrenal gland of animals with deletion of Dio3os in the paternal allele showed a 2.5-fold increase in the paternal to maternal allele expression ratio of Dio3, when compared to those with a Dio3os deletion in the maternal allele. Conclusions: Dio3os is an imprinted gene preferentially expressed from the maternal allele in neonatal liver, but its imprinting is tissue-specific and may not be present in other tissues. Dio3os plays an allele-specific regulatory role in the allelic expression of Dio3 in the adrenal gland, a finding that could potentially extend to other tissues. Singer-Sam, J. (2010). When only one allele of a gene is actively transcribed, gene expression is termed monoallelic. What does monoallelic gene expression have to do with cats, twins, and genetic diseases? Nature Education 3(3):1. Retrieved February 26, 2022, from https://www.nature.com/scitable/topicpage/monoallelic-expression-8813275/
