Elucidating Epigenetic Roles of rDNA locus and nucleolus

Multi-copy rDNA locus is the primary determinant of nucleolus formation and mis-regulation of
its copy number is associated with enlarged and irregular nucleoli which is a hallmark of malignant cancer. rDNA copy loss is associated with epigenetic mis-regulation, increased DNA damage and massive gene expression changes and is frequently observed in cancer. In Drosophila, induced variation in rDNA copy number modifies heterochromatin-induced silencing and causes expression changes in hundreds to thousands of genes in an isogenic background. Several cancers including osteosarcoma, lymphoma, esophageal adenocarcinoma, lung adenocarcinoma & squamous cell carcinoma and kidney renal clear cell carcinoma were all observed to have reduced rDNA copies which is surprisingly associated with enlarged & irregular nucleoli. Somatic mutations that cause loss of function in P53 and mouse leukemia model with PTEN deletion both display rDNA copy loss that is associated with higher proliferation. In a MYC driven lymphoma model, malignant progression associates with differential rDNA chromatin interactions that contribute to upregulation of specific genes that regulate growth and differentiation. Thus, malformation of nucleolus and rDNA-chromatin interactions affect gene expression and essential biological processes beyond ribosome biogenesis.

Nucleolus anchors specific and yet diverse chromatin domains which are called nucleolar associated DNA domains (NADs). In addition to interacting with Pol-I and Pol-III transcribed genes, subtelomeric regions, repetitive DNA and centromeres, NADs surprisingly include distinct chromatin with protein coding loci. The regions harboring protein coding genes are classified into two categories, (1) constitutive heterochromatin such as transposable elements or pseudogenes–type-I NADs–(2) heterochromatin with relatively higher gene expression that is comprised of developmentally regulated genes–type-II NADs, as illustrated below.

We currently lack an understanding of the causality of functional dynamics and significance of nucleolus for heterochromatin maintenance and gene expression. Thus, systematic approaches to elucidate the functions of nucleolus-chromatin interactions is fundamentally important to understand the ribosome biogenesis independent roles of nucleolus in chromatin regulation and gene expression

The specific questions that we are working on: Does nucleolus play an active role in silencing type-I NADs heterochromatic regions? Does nucleolus play a role in fine-tuning type-II NAD expression through changes in their spatial distribution during development? We are in a unique position to study the epigenetic function of nucleolus during differentiation and tissue diversification for the following two reasons. First, C. elegans is amenable to tracking chromatin organization changes along the nucleolus axis in different cell types. Second, C. elegans is the only metazoan with a complete deletion of multi-copy rDNA locus (18S, 5.8S and 26S), which is required for nucleolar assembly. rDNA locus deleted embryos that we have generated can survive via maternal ribosome loading but lack a discernible nucleolus structure. Thus, this system will enable us to directly probe the ribosome biogenesis independent epigenetic role of nucleolus in a multi-cellular organism.

Elucidating the function of nucleolus for heterochromatin maintenance and gene expression during cell division and differentiation will be fundamentally valuable in understanding rDNA loss mediated epigenetic changes that happens disease systems such as cancer.