Research Goals

  1. Elucidate molecular mechanisms regulating expression of H19 and Igf2 genes.
  2. Develop and characterize mouse models for loss of imprinting diseases.
  3. Develop and characterize mouse models to understand the role of H19 RNAs in normal development.
  4. Understand the role of cardiac calsequestrin in normal heart development and function.

Genomic imprinting is an unusual form of gene regulation in which an allele's parental origin restricts its expression. For example, almost all expression of the non-coding RNA tumor suppressor gene, H19, is from the maternal chromosome. In contrast, expression of the neighboring Insulin-like Growth Factor 2 gene (Igf2) is from the paternal chromosome. Imprinted genes are not randomly scattered throughout the genome but rather are localized in discrete clusters where monoallelic expression is determined by a cis-acting element called the Imprinting Control Region (ICR). We study a cluster of imprinted genes on the distal end of mouse chromosome 7. The syntenic region in humans (11p15.5) is highly conserved in gene organization and in expression patterns. Imprinting of H19 and of Igf2 is regulated by the H19ICR which is located just upstream of the H19 promoter. We have shown that the molecular function of the H19ICR is to organize the region into alternative 3D structures. Upon maternal inheritance, the H19ICR is not methylated, binds the transcriptional regulator, CTCF, and organizes the locus into loop structures that bring the H19 promoter into physical contact with share downstream enhancers but exclude the Igf2 promoter from physical contact with these enhancers. Upon paternal inheritance, the H19ICR is methylated and cannot bind CTCF. Therefore, alternative loop structures form which allow Igf2 promoters to interact with the shared enhancers. In current studies we are analyzing the role of the enhancer sequences and binding proteins in forming these alternative loop structures.

In humans, epigenetic mutations that disrupt H19ICR function result in loss of monoallelic expression. Lesions altering paternal H19ICR function result in loss of Igf2 expression and biallelic (2X) H19 expression and are associated with Russell-Silver syndrome. Lesions altering maternal H19ICR function result in biallelic (2X) Igf2 expression and in reduced  levels of H19 RNA and are associated with Beckwith Wiedemann syndrome and with several pediatric cancers. We have generated mouse models for BWS. We are using complex genetic strategies to understand which BWS phenotypes are associated with extra Igf2, which are associated with loss of H19, and which are dependent on their combined mis-regulation. Especially we are focused on understanding the etiology of cardiac dysfunctions.  Our data show that H19 lncRNA regulates endothelial cell fate.

The biochemical roles for H19 are highly controversial. For example, in human studies, H19 has been identified  both as a tumor suppressor and as an oncogene. The H19 gene encodes two RNAs: 1) a 2.3 kb long noncoding RNA and 2) miRNA 675. We have established mouse models that prevent expression of the lncRNA only, of the miRNA only, and of both together. We are analyzing development phenotypes associated with these three kinds of genetic lesions. Our results indicate that these studies will resolve some key controversies regarding H19 function.

Finally our lab has generated and is characterizing  mouse models that allow us to conditionally ablate or to conditionally rescue Casq2 function. Casq2 encodes cardiac calsequestrin, a protein that regulates opening of the RyR Ca2+ channel in cardomyocyte junctional sarcoplasmic reticulum (SR). Loss of function mutations in mice and humans cause Ca2+ leakage from the SR and thereby result in stress induced arrhythmias. We have developed our models as a means to understand the role of normal development in disease progression. One unexpected finding was that the slow heart rates in Casq2-deficient mice and humans are an excellent therapeutic target.  

Please see the NICHD Annual Report for a detailed description of our research.

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