Background:
Systemic sclerosis (SSc) is a serious autoimmune disease characterized by fibrosis of the skin and internal organs. There is currently no established treatment for the disease, but recent clinical trials have highlighted deep B-cell depletion therapy as a promising treatment for SSc and several other autoimmune diseases.
B cells play an important role in immune responses against foreign intruders such as viruses and bacteria through production of antibodies, secretion of cytokines and presentation of antigen to T cells. In autoimmunity, normal tolerance mechanisms are broken, leading to activation of B cells reactive with self-antigens. The result is generation of disease-specific autoantibodies. In SSc, there is formation of specific autoantibodies targeting centromere proteins (CENPs), in particular CENP-B, in the cell nucleus. These autoantibodies depend on interactions between B cells and T cells, possibly via complex formation between CENP-B and bacterial DNA (Fig. 1).
Aim:
Work performed in the group has identified CENP-B-specific B cells in blood samples of SSc patients. These cells have been isolated, and their antibody genes have been cloned for generation of recombinant monoclonal antibodies (mAbs). Interestingly, we have observed strong selection for certain antibody gene segments across patients, indicating targeting of a dominant epitope. To understand the mechanisms of autoantibody formation, we aim to characterize binding between recombinant CENP-B and anti-CENP-B mAbs in detail.
The goal of this MSc project is to generate length variants of CENP-B that will be used to set up binding assays with anti-CENP-B mAbs as well as functional assays to evaluate binding to DNA. It will thereby be possible to locate antibody target epitopes and assess formation of ternary complexes between CENP-B, mAb and DNA. Further, the generated CENP-B proteins and anti-CENP-B mAbs can be used to perform structural studies employing techniques such as X-ray crystallography or electron microscopy to get detailed insight into the antibody-antigen interaction. Ultimately, if we understand the rules of autoantibody formation, it might be possible to devise new treatment strategies aiming to prevent pathogenic T cell-B cell interactions.
Key methods:
- Molecular cloning
- Expression and purification of recombinant protein in eukaryotic cells
- Enzyme-linked immunosorbent assay (ELISA)
- Quantitative PCR
Workplace:
The work will be carried out in Rasmus Iversen’s research group at the Department of Immunology, Oslo University Hospital – Rikshospitalet. We are a relatively new research group focusing on B-cell biology - in particular in relation to autoimmune diseases. To gain insight into disease mechanisms, we employ a combination of biochemical studies and high-throughput sequencing analyses of single cells isolated from patient samples.
Supervisors:
- Rasmus Iversen (rasmus.iversen@medisin.uio.no)
- Christine Skagen (postdoc in the group)
- Finn-Eirik Johansen (internal supervisor)