Bahjat, M.

English

B cells generate a vast repertoire of B-cell receptors (BCR), which are generated by immunoglobulin (Ig) gene diversification processes. Ig diversification is initiated in the bone marrow, through the process of Variable (V), Diversity (D) and Joining (J) gene segment recombination (VDJ recombination), which is an antigen-independent and highly ordered process that requires the recombination activating gene products 1 &amp; 2 (RAG1/2). Upon encountering antigen in secondary lymphoid organs, B cells undergo additional Ig diversifications during two processes termed somatic hypermutation (SHM) and class switch recombination (CSR), which require activation-induced cytidine deaminase (AID). Since the enzymatic activity of both the RAG1/2 complex and AID results in DNA damage, the regulation of these enzymes is crucial to ensure genomic integrity. The mechanistic aspects of this regulation are the central topics of this thesis. The results of our findings are presented in 7 chapters of this thesis

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UvA-DARE is a service provided by the library of the University of Amsterdam (https://dare.uva.nl)UvA-DARE (Digital Academic Repository)The delicate balance between DNA damage and repair in B cellsBahjat, M.Publication date2019Document VersionOther versionLicenseOtherLink to publicationCitation for published version (APA):Bahjat, M. (2019). The delicate balance between DNA damage and repair in B cells. [Thesis,fully internal, Universiteit van Amsterdam].General rightsIt is not permitted to download or to forward/distribute the text or part of it without the consent of the author(s)and/or copyright holder(s), other than for strictly personal, individual use, unless the work is under an opencontent license (like Creative Commons).Disclaimer/Complaints regulationsIf you believe that digital publication of certain material infringes any of your rights or (privacy) interests, pleaselet the Library know, stating your reasons. In case of a legitimate complaint, the Library will make the materialinaccessible and/or remove it from the website. Please Ask the Library: https://uba.uva.nl/en/contact, or a letterto: Library of the University of Amsterdam, Secretariat, Singel 425, 1012 WP Amsterdam, The Netherlands. Youwill be contacted as soon as possible.Download date:15 Apr 20238Outline of this thesisIn this thesis we aim to get a deeper understanding of the role of DNA damage response (DDR) and DNA repair mechanisms in B-cell development, and how these contribute to the prevention of B-cell transformation.In Chapter 1, we reviewed the current literature on the regulation of two mutagenic enzymes that shape B-cell development; recombination-activating gene 1 & 2 (RAG1/2) complex and activation-induced cytidine deaminase (AID).  More specifically, we discuss how these enzymes are regulated by the DDR. The RAG1/2 complex is of crucial importance for lymphocyte development by initiating the rearrangement of both the T-cell receptor (TCR) and B cell receptor (BCR) genes. For these rearrangements to occur, RAG1/2 induces DNA cleavage.  Off-target activity of RAG1/2 is involved in the generation of chromosomal aberrations that characterizes several types of B-cell lymphoma and leukemia.  Therefore, the proper regulation of its expression and activity is essential to prevent genomic instability.  In Chapter 2 we investigated the regulation of RAG1/2 in normal and transformed precursor-B cells. Mouse Abl transformed pre-B cells and human pre-B-cell acute lymphoblastic leukemia (B-ALL) cells were used as model systems.  The role of the nuclear factor kappa-B (NFKB) and phosphoinositol-3 kinase (PI3K)/AKT signaling pathways in the regulation of RAG1/2 expression and activity was studied in this chapter. The DDR plays an important role in maintaining genomic integrity.  In Chapter 3, we studied the contribution of this protective response in controling RAG1/2 expression and activity in cells that sustain RAG1/2-dependent and -independent DNA damage. The involvement of the ataxia telangiectasia mutated (ATM) kinase, as one of the main sensor of DNA damage, and its crosstalk with the forkhead box O1 (FOXO1) transcription factor in the regulation of RAG1/2 expression was evaluated in this chapter. Upon activation, B cells may undergo additional antigen-dependent Ig gene diversifications in secondary lymphoid organs.  In the germinal center (GC), B cells undergo somatic hypermutation (SHM) and class switch recombination (CSR).  During SHM, single nucleotide substitutions in the antigen-binding coding part of the Ig gene are introduced, which alter the affinity of the antibody.  Affinity maturation takes place by selection of B-cell clones that have acquired increased affinity for the antigen.  By CSR, the isotype of the expressed Ig is altered, which shapes the effector function of the humoral immune response.  Both SHM and CSR crucially depend on AID, which is responsible for MB PS 2019v12.indd   8 01-04-19   10:419the introduction of mutations in the IgV genes during SHM and DNA double stranded breaks in the Ig switch regions (DSBs) during CSR.  Several DNA repair pathways are involved in the resolution of AID-instigated DNA lesions, which in part determine the fate of these lesions.  In Chapter 4 we studied the complex interplay between the base excision repair (BER) and the mismatch repair (MMR) pathways during AID-dependent mutagenesis. Targeting the DDR has become a bona fide therapeutic strategy for the treatment of a wide range of cancers.  In Chapter 5 we investigated the effects of the NEDD8-activating enzyme (NAE) inhibitor pevonedistat (MLN4924) on human BCR-ABL+ leukemia cells, focused on understanding the mechanism behind its anti-leukemic activity. In Chapter 6, we described the in situ proximity ligation assay (PLA) as a useful novel technique that allows the direct visualization of protein-protein interactions in suspension cells.  We used this approach to study DDR-related protein complexes that are formed upon the induction of DNA damage. In Chapter 7, we discuss our findings.  Outline of this thesisMB PS 2019v12.indd   9 01-04-19   10:41

The delicate balance between DNA damage and repair in B cells

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The delicate balance between DNA damage and repair in B cells

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