Year: 2014
Type: dissertation
Abstract: B cells undergo secondary antibody diversification to elicit an effective immune response. Secondary antibody diversification generates antibodies with high affinity for the cognate antigen, and antibodies of different classes. Different classes of antibodies mediate different effector functions, enabling the humoral response to be fine-tuned to the infection at hand. The different classes are generated by a process called class switch recombination (CSR), which requires the generation and repair of double-stranded DNA breaks (DSBs) at the antibody heavy chain locus, and relies on the 53BP1-dependent DNA damage response (DDR). However, the molecular mechanism and function of DDR in CSR is not fully understood. To characterize the role of DDR in CSR, we first established an in vitro CH12F3-2 mouse B cell line/ RNA interference system and used this system to demonstrate that the DDR factors RNF8 and RN168, play a role in CSR. Using this system, we then developed and conducted a genome-wide screen to identify novel CSR factors, followed by a secondary screen to identify DDR factors among the CSR candidates. Based on this study, we selected one candidate, the Drosophila enhancer of yellow 2 homolog (Eny2), for further characterization. Eny2 is a structural component of the H2B-deubiquitination module of the SAGA complex. We found that Eny2 likely acts downstream of the DSBs generated for CSR, and that Eny2 might act with the SAGA complex in mediating H2B-deubiquitination to facilitate CSR. We also provide evidence that, in addition to its role in CSR, Eny2 is important for H2B-deubiquitination upon irradiation-induced DNA damage, and may be required for optimal DNAPK activation.
Source: TSpace (University of Toronto)
Author: Shaliny Ramachandran
Language: English
Topic: DNA Repair Mechanisms
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W2955793849