The role of RuvBL2 in the development, maturation, and aging of antibody producing B-cells

The human body encounters a limitless number of potential foreign antigens on a daily basis. How, then, does our body recognize and fend off such foreign invaders before having had any prior exposure to them? An important component of the immune system are B-cells, which produce high affinity antibodies that are highly specific for a given antigen. Instead of encoding individual genes to produce individual antibodies, B-cells evolved the ability to have one specific gene undergo programmed mutagenesis, which leads to large scale genomic rearrangements combined with an extraordinarily high rate of point mutations, where a single base of the DNA is altered, reaching a million-fold above background levels. Paradoxically, however, the same machinery that normally protects our genome from DNA damage is hijacked by B-cells in order to induce these mutagenic alterations. Therefore, unlike all other somatic cells, B-cells are unique in that their development and maturation is dependent on programmed genomic instability. As one may expect, this mutagenic process sometimes results in off-target mutations at non-antibody genes, which can lead to chromosomal instability and compromises the health and well-being of many organisms. One such example is the accumulation of mutations in B-cells leading to the aging of the immune compartment, which has been acquiring increased emphasis as an important driver for organismal ageing. The aim of my research proposal is to understand how DNA repair processes affect the development, maturation, and ageing of B-cells. Our results will have broad implications for the mechanisms underlying antibody diversification, genomic instability, tumorigenesis, and how DNA damage affects cellular and organismal aging.

An inadequate antibody response is a public health issue because it compromises our ability to fend off infections and maintain our well-being. This is exacerbated by natural aging, which renders our antibody response even weaker and less specific towards foreign antigens. In this proposal, we will take advantage of two genome wide screens for DNA repair and antibody diversification to understand the contribution of RuvBL2 ATPase/Helicase protein in controlling B cell development, maturation, and aging. Aside from commonly used molecular and immunological techniques we will continue developing our bioinformatic tools published previously (PNAS 2015), which will facilitate the analysis of large sets of low complexity next generation sequencing data required for the high resolution analyses of antibody hypermutation in our first two objectives. We will employ a combination of conventional and CRISPR-mediated genome editing tools to examine the role of the active transcription histone marks on DNA repair fidelity in B cells and its contribution to antibody diversification. We will examine the potential crosstalk between DNA damage repair mechanisms and histone modifications on antibody diversification throughout the life course of mice. Our studies will provide better understanding of antibody development and how to potentially preserve the efficiency of our immune system through old age.

This basic science project is data-driven, discovery-based, medically focused, and commercially exploitable. The timescale to impact in the clinical or industrial setting is by necessity long term for basic science advances, but training advances and benefits to the scientific community will be on a shorter timescale. Thus, this work may have a positive impact on a number of important beneficiaries:

1) Scientific community: The wider scientific community in the fields of humoural immunity, antibody development, genomic stability, and aging will benefit. This project will improve understanding of the DNA repair mechanisms involved in the development and progression of antibody-related immunodeficiency and a greater understanding of the biological function of DNA mismatch repair in tumorigenesis. In the short term, this project will also have an impact on the scientific community through the development of an automated bioinformatics pipeline for the analysis of mutations in low complexity reads for next generation sequencing; which will facilitate the identification and cloning of therapeutic monoclonal antibodies. We have a track record of freely sharing reagents and bioinformatic tools we develop (http://scb.aecom.yu.edu/cgi-bin/p1 and PNAS 2015)

2) Researcher working on the project: The researcher undertaking the project will benefit significantly. The project involves a wide range of scientific techniques; both in vitro and in vivo and so will provide extensive training and experience and thus progress personal development and future employability.

3) Biopharmaceutical industry: Our research has the potential to lead to the development of new modalities for: i) better design and cloning of therapeutic monoclonal antibodies and ii) counteracting the effects of aging. Understanding antibody diversification has unlimited potential for exploitation in drug design, especially for the generation of experimental, diagnostic, and therapeutic monoclonal antibodies. Our work will potentially allow us to mimic antibody development ex vivo, thereby allowing us to produce antibodies quickly, more economically, and efficiently. This could be particularly useful for companies developing drugs against immune aging and companies attempting to optimize antibody cloning (for use as vaccines). Knowledge gained from this study will help identify the key components that could ultimately be commercially exploited for large-scale ex vivo antibody production. This would be of benefit to the UK and European pharmaceutical sectors; as well as in the diagnosis of pathophysiological conditions and disorders in the public health sector.

4) Society and public engagement; Aging is steadily becoming a public health issue in the UK and worldwide, representing an increasingly significant cost to society. It has been predicted that by 2050 there will be ~19 million people in the UK over 65 years of age. . We will work closely with press officers at Exeter and the BBSRC to communicate aspects of this research to the general public and to policy-makers and to explain its potential importance for society and health. The potential to develop new modalities to delay or conform aging consequences will engage public interest. The University of Exeter has exceptional links with the wider public including regular interactive visits of the public.

5) Local economy. Our tools may be translated in the mid-to-long term into therapeutic entities and generate patentable or commercially exploitable tools which will benefit the UK economy. The research knowledge transfer at the University of Exeter has experience in promoting and assisting scientists to commercialize their basic research.