B lymphocytes are the blood cells responsible for immunoglobulin (Ig, or antibody) production and secretion, a fundamental process for the humoral immune response to infections and vaccines. To become antibody factories, B lymphocytes are activated and then differentiate into plasma cells. This requires huge changes in the cell’s secretory membranes and particularly involves rapid growth of the endoplasmic reticulum (ER), the site of protein synthesis, as well as changes to the downstream pathway control points.
In mammalian cells, the zones of protein export from the ER are termed the ER exit site (ERES). Transport of immunoglobulins (as well as thousands of other newly synthesised proteins) from the ER to the Golgi apparatus is mediated by the COPII coat complex, which remodels membranes into coated carriers while concentrating cargo into them. While the basic functions of COPII components have been well characterised in yeast, its regulation in the more complex mammalian systems (where multiple COPII paralogues are present) and particularly in specialised secretory cells remains poorly understood.
Antibodies can come in different varieties known as isotypes or classes of which there are 5 in mammalian cells; IgA, IgD, IgE, IgG, IgM. The Ig class secreted by a B cell is dependent on B cell maturation and performs different roles during the immune response. IgG is secreted as a monomer while IgM is much larger, forming a pentameric structure of 45-50 nm diameter, which has implications for cargo export and transport carriers.
This project aims to understand the mechanisms behind secretion of different types of immunoglobulins from B-cells. Preliminary data from our labs suggest differentiating B cells express specific COPII paralogues. This project will build on our observations to investigate outstanding questions:
1. Is paralogue specificity associated with specific changes in secretion, for example cargo load, or expression of certain Ig isotype that have very different sizes?
2. Does COPII composition differ between naïve undifferentiated B cells and plasma cells?
3. What are the morphological changes in ERES organisation upon differentiation?
This project will have important implications both in terms of human health and response to infection, and also for optimised antibody production in biomedical/pharma applications.
You will use a combination of cell biology, biochemistry, light microscopy and cryo-electron tomography to reveal the role of the COPII coat complex in export of different classes of Igs from the endoplasmic reticulum (ER). This will include B cell in vitro activation assays, confocal microscopy, real-time PCR, mathematical modelling, CRISPR gene editing, cryo electron microscopy and tomography, cryo FIB/SEM, cryo-correlative microscopy.
B cells, antibody secretion, COPII complex, membrane trafficking, fluorescence microscopy, cryo-electron microscopy.