Mentor: Dr. Tamara Davis
For most genes, both parents’ copies of the gene are expressed. For about 85 mammalian genes, however, one parent’s copy is silenced. These “imprinted genes” must gain their differential expression not because of changes in the actual genetic code; instead there must be structural changes in the DNA which silence or activate imprinted genes. In Gtl2, a maternally expressed imprinted gene, the mechanism controlling expression is differential methylation. Methyl groups attach to cytosines in areas rich in 5’-CG-3’, known as CpG islands, on the paternal allele, silencing its expression. The maternal copy of the gene, on the other hand, remains unmethylated and is therefore expressed.
For some areas of Gtl2, the paternal allele-specific methylation is acquired during spermatogenesis and retained throughout development. However, the CpG island located at the Gtl2 promoter is not methylated in sperm. My research hopes to determine when methylation of the Gtl2 promoter is acquired on the paternal copy of the chromosome during mouse embryonic development. I am analyzing methylation in mouse blastocysts (3.5 days post conception) and embryos (6.5 days post conception) using bisulfite mutagenesis. I will set up crosses between different strains of mice, which exhibit DNA sequence variation at three distinct bases in the Gtl2 promoter region, in order to identify which copy of the gene came from the mother and which from the father. I will then compare methylation patterns over development on both the maternal and paternal alleles, to determine when the methylation of relevant cytosines occurs on the paternal allele.
Determining the timeframe of methylation can potentially shed some light on the mechanism for differential methylation. The way the paternal gene “remembers” its origin without methylation is still unclear, as is the catalyst for the gene to gain its methylation during development. Methylation is most likely activated by a certain chemical signal, but without knowing when methylation occurs, it is impossible to identify the specific chemical switch. My research attempts to take the first step towards solving this problem.