Assessing the Pattern of Methylation at the lit1 Control Region in Developing Mouse Oocytes

Posted May 26th, 2010 at 2:38 pm.

Lilah Rahn-Lee

Imprinting is the differential expression of alleles based on parental origin. 70 imprinted genes have been identified in mammals which are either expressed maternally and silenced paternally, or expressed paternally and silenced maternally. Imprinted genes are marked as coming from one parent or another by differential methylation of the DNA along or near the gene. In early embryonic development, the DNA of the germline must be reprogrammed so that all alleles bear the maternal mark for developing oocytes, and the paternal mark for developing sperm. Understanding the patterns of methylation along imprinted genes in these early stages of the development of sex cells can lead us to a better understanding of imprinted control in mammals and of mechanisms by which gene expression can be regulated.

Imprinted genes occur in clusters on mammalian chromosomes. These clusters of genes are frequently regulated by one control region, whose methylation state dictates the expression or repression of the surrounding imprinted genes. One such control region is found at the promoter of the lit1 gene, which is methylated maternally and expressed paternally. Methylation at this location controls expression of several adjacent genes, including nap2, ipi, impt1 , and p57kip2 . I will be studying the establishment of methylation of the lit1 gene in developing oocytes from mouse embryos, in order to determine how the lit1 promoter controls expression of this region of the chromosome.

We are able to study DNA methylation through bisulfite mutagenesis of genomic DNA, a process which deaminates cytosine bases to uracil in non-methylated DNA only. The altered lit1 gene may then be amplified and sequenced to determine its methylation state at the time of mutagenesis. By using F1 hybrid embryos from a cross between the mouse strains black 6 and castaneus, which contain polymorphisms along the lit1 gene we are able to identify the parental origin of alleles and correlate that with their methylation state of the developing germ line DNA. Through the mutagenesis and sequencing of the lit1 gene across a spectrum of developmental stages, I will be able to determine how the methylation of the lit1 control region is established.

Filed under: 2004,Davis, Dr. Tamara,Rhan-Lee, Lilah by Ann Dixon

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