What is genomic imprinting? How was GI discrovered? - Dr Helen crouse and the Dipteran Fly. When does genomic imprinting occur and where? It takes place before fertilization, in the egg and sperm cells What is hemizygosity? A chr in a diploid cell is hemizygous when only one copy of a gene is present or being expressed. This can be observed when one copy of a gene is deleted, or silenced. Hemizyogisty is also observed where heterogametic sex is located on a sex chromosome – ie, males. Human males – almost all X-linked genes are hemizygous in males with normal chrs because they only have one X chromosome and a few of the same genes are on the Y chromosome. Genomic imprinting is often described as being parent-of-origin specific, what does this mean? Parent-of-origin effects occur when the phenotypic effect of an allele depends on whether it was inherited from an individuals mother or father. The best characterised parent-of-origin dependent gene expression is associated with genomic imprinting. What is chromosome imprinting When an entire chromosome is silenced and a Barr Body is produced What is X inactivation? Calico Cat example What is the balance dosage? Describe the process of DNA methylation. Covalent addition of the methyl group at the 5-carbon of the cytosine ring resulting in 5methylcytosine 5mC. This methyl group projects into the major groove of the DNA and inhibits transcription. (S-adenosylmethionine – dna methyltransferases – S-adenosylhomocytosine) - Dna methyltransferase is also known as DNMT// DNMT3a+3b mediate establishment of dna meth DNMT1 maintains the pattern What are CpG sites? They are imprinting clusters. They are heavily methylated islands found near promoters of normal somatic cells. Gene silencing is initiated by abnormal CpG methylation near these promoters, DNA hypermethylation recruits a complex of proteins that include Methyl-CpG-Binding domains such as MeCP2 which brings co-repressors and Histone DeAcetylase to remove the acetyl group from histone proteins on the DNA which causes the compaction of chromatin, causing silencing of gene expression. What are the principles behind Nuclear Transfer Technologies Nuclear transfer is a form of cloning. The steps involve removing the DNA from an oocyte (unfertilised egg), and injecting the nucleus which contains the DNA to be cloned. How can one identify an imprinted gene? How does imprinting of genes occur? (epigenetics – dna meth and histone) What are imprinting clusters? The grouping of imprinted genes within clusters allows them to share common regulatory elements, such as non-coding RNAs and differentially methylated regions (DMRs). When these regulatory elements control the imprinting of one or more genes, they are known as imprinting control regions (ICR). What is the function of the gametic DMR? DNA is mostly methylated at a CpG site, which is a cytosine followed by a guanine. The “p” refers to the phosphate linker between them. DMR usually involves adjacent sites or a group of sites close together that have different methylation patterns between samples. CpG islands appear to be unmethylated in most of the normal tissues, however, are highly methylated in cancer tissues. What are control regions/elements? What is the kinship theory? What is the trophoblast defence theory? What is the role of lncRNAs in imprinting? Some evidence suggests that several of these lncRNAs may regulate their own transcription in cis, and that of nearby genes, by recruiting remodeling factors to local chromatin. Notably, lncRNAs are known to exist at many imprinted gene clusters. Genomic imprinting is a complex and highly regulated process resulting in the monoallelic silencing of certain genes, based on the parent-oforigin of the allele. It is thought that lncRNAs may regulate many imprinted loci, however, the mechanism by which they exert such influence is poorly understood. - One mode of lncRNA action is to recruit epigenetic silencing to target distant genes on the same chr. A well-characterised group of lncRNAs that acti this way to silence genes are imprinted lncRNAs such as the Airn silencing gene in the Igfr2 imprinted cluster. Airn expression blocks chr interactions between the promoter and the Airn gene locus. What are the functions of genomic imprinting? What are the disorders associated with maternal imprinting defects? Maternal chromosome 15 deletion along with a paternally imprinted chromosome 15 causes Angelman syndrome. AS is caused from a lack of function of part of Chr15 inherited from the offsprings mother – the fathers version of this gene are imprinted, therefore inactived – no functional version of the gene remains. What are the disorders associated with paternal imprinting defects? Paternal chr15 deletion, paired with a maternally imprinted chr15 causes Prader-willi syndrome. PWS is caused by a deletion in the fathers chr15, the chr15 gene in the mother is silenced therefore no working copy is available. Characterised by obesity and type II diabetes Describe a molecular genetic test for which imprinted disorders can be tested? Most diagnostic labs in Europe use MS-MLPA to detect the methylation status of a gene and if a deletion is present. If an abnormality is detected through MS-PCR, presence of a deletion is investigated using MS-MLPA. If abnormal methylation is detected but there is an accompanying deletion, it must be figured out which chr there is a deletion on. Therefore microsatellite analysis using polymorphic markers will distinguish between a UPD or another defect. If the microsatellite analysis shows UPD of chr 15, analysis of the IC region should be performed to distinguish between an isolated epimutation or a deletion in the IC domain.
