Role of ncRNA Surveillance Complex "RNA Exosome" in Class Switch Recombination and Somatic Hypermutation

PROJECT SUMMARY
Background: Class switch recombination (CSR) and somatic hypermutation (SHM) are two B lymphocyte
specific processes that mediate antibody diversification. Activation Induced Cytidine Deaminase (AID) is
essential for initiating both of these processes by deaminating cytidine residues in immunoglobulin (Ig) loci
DNA. Despite its specific and indispensible function in the Ig loci, AID has been demonstrated to also
deaminate non-Ig locus genes, with the concomitant negative outcome of catalyzing various oncogenic
translocations that manifest in tumorigenesis. Recent work has indicated that AID's DNA deamination activity
requires its association with transcriptionally stalled RNA polymerase II and the RNA exosome complex. RNA
exosome is a cellular non-coding RNA (ncRNA) processing and/or degradation macromolecular complex. How
ncRNA and RNA exosome facilitate AID's DNA deamination activity is a question we plan to address.
Objective/Hypothesis: We will determine how RNA exosome activity on ncRNAs support AID activity at the
IgH locus and stabilizes 3-dimensional organization of the B cell genome. Specific aims: AIM 1: To understand
the chromatin associated function of noncoding RNAs and RNA exosome in controlling AID activity. AIM 2: To
understand the role of CTCF binding element (cbe) ncRNAs in TADIgH organization. AIM 3: To understand the
role of novel RNA exosome cofactors in regulating AID activity.
Study Design: Using mouse models that are deficient in RNA exosome-mediated RNA degradation, we have
identified regions in the B cell genome that express exosome sensitive ncRNAs and also are mutated by AID.
We will evaluate the mechanism of formation of single-strand DNA structures following localized chromatin
remodeling at these identified AID target DNA sequences. We have discovered a new family of exosome
sensitive ncRNAs that are associated with CTCF-binding elements (CBE), defined as cbeRNAs. We will
evaluate how cbeRNA processing regulates the 3 dimensional topologically associating domain structure of IgH
and influences SHM and CSR efficiencies. Finally, using B cells from TAP-tagged Exosome expressing mouse
model, we have purified and identified chromatin associated RNA exosome co-factors. We will evaluate the role
of RNA exosome co-factor(s) in stimulating AID/RNA exosome complex function.
Disease Relevance: The proposed studies will lead to a better understanding of the mechanisms initiating
AID dependent oncogenesis in B lymphocytes (particularly in the contexts of DLBCL and multiple myeloma) as
well as have direct implications in understanding B lymphocyte-based immunodeficiency syndromes like
Hyper-IgM syndrome type 2. RNA exosome is implicated in many diseases including numerous
neurodegenerative disorders and multiple myeloma and findings from this grant will lead to a better
understanding of the etiologies of the disorders and development of therapies.

Grant Number: 5R01AI099195-15
NIH Institute/Center: NIH
Principal Investigator: Uttiya Basu