Activation Induced cytosine Deaminase(AID) and Somatic Hypermutation

Contents 1 Introduction 2 Section 1 Mechanism 3 Section 2 Regulation 4 Conclusion 5 References Introduction Deamination is a proposed mechanism of somatic hypermutation(SHM) which results in point mutations within the Immunoglobulin variable region(IgV) gene. Deamination involves the use of activation-induced deaminase(AID), a protein which converts cytidine(cytosine bound to ribose) to uridine(uracil bound to ribose). The completion of this process results in a miss match of bases and therefore initiates DNA repair. However, the repair process uses mutagenic mechanisms, such as error-prone DNA polymerases, which result in the creation of mutations in and rearrangement of IgV genes. These mutations code for a varying expression of the gene itself resulting in an antibody with either a higher or lower binding affinity. After hypermutation, higher binding affinity is selected for during affinity maturation resulting in a more effective antibody. Mutagenic processes like those induced by AID must be tightly regulated due to the disease related and oncogenic risks of mutating DNA. Therefore, the AID protein is controlled by a complex network of regulatory factors which are vital to the safety of SMH.

AID The AID protein itself is 198 amino acids long and is encoded by the AICDA region of the DNA. AID belongs to the APOBEC-1-related protein family which includes other DNA editing molecules. The molecule contains a cytosine deaminase domain as well as a leucine rich region around the c terminus, the latter of which is likely involved in a protein-protein interaction. Within B cells, AID initiates the processes of both SHM and class switch recombination(CSR), although different parts of the protein are involved in each process.The C terminus of the AID protein is associated with CSR. One study(https://onlinelibrary.wiley.com/doi/epdf/10.1002/humu.20414) show that the expression of C-terminal mutations in the AICDA gene lead to decreased levels of CSR as the level of SMH remains constant.Similarly, mutations of the N terminus led to the defective induction of SMH suggesting that the N terminus of AID plays some sort of role in initiating SHM.

Mechanism and regulation of AID AID carries a potential tumorigenic nature when mistargeted on to non-Ig loci.AID has been shown to promote B cell leukemia, lymphoma, and myeloma. Therefore the regulation of AID is as critical as the mechanism itself in terms of protecting the body. Regulation of AID starts with the AICDA gene and the transcription factors which manage its expression. AICDA is composed of 4 regions which include binding sites for at least 19 transcription factors. While transcription factors like NFκB, STAT6 and C/EBP bind to enhancer regions which promote transcription, transcription factors like PAX5 and E2A bind to and inactivate repressor regions of the gene therefore also promoting transcription. Most of these transcription factors are only upregulated during B cell activation, subjugating incidents of hypermutation as a response to B cell activation only. High transcription levels of AID are not desired in non-differentiating B cells. Excess levels of AID are linked to more frequent targeting of non IgV region genes inducing undesired mutations which may be associated with cancers. As a result, the level of AID is also heavily regulated on a post-transcription level using miRNAs. Two main miRNAs(miR-155 and miR-181b) target transcripts of the AICDA gene. Both miRNAs bind to the 3’ untranslated region of AID mRNA inducing a mechanism that ultimately results in lowered levels of SMH and CSR. Despite targeting the same sites, miR-181b differs in its expression pattern from miR-155. miR-181b levels are highest in unstimulated B cells and its levels decrease heavily as a result of activation. However, miR-181 levels gradually recover after activation.It should be noted that miR-181 and miR-155 have multiple target molecules besides AID mRNA and therefore the expression patterns of the two molecules may not be entirely based on the regulation of AID levels. AID protein is further regulated by the cell using protein localization. The concentration of AID in the nucleus is regulated by multiple mechanisms, ultimately resulting in a higher concentration of AID in the cytoplasm than in the nucleus. One mechanism involves an anchor sequence on the C-terminal region of AID which hitches the protein to the cytoplasm. Another mechanism involves the active transport of AID into the nucleus via the protein’s N-terminal NLS(nuclear localization signal). Finally, AID is transported out of the nucleus via the protein’s NES(nuclear export signal) which binds to the CRM1 export protein carrying the molecule into the cytoplasm. Concentration of AID is also regulated by the half life of the molecule within and outside of the nucleus. While AID has a half life of around 8 hours in the cytoplasm, nuclear AID has a half life of only about 2.5 hours due to higher levels of polyubiquitination by proteasomes. The reason being that high levels of AID degradation in the nucleus prevent non Ig-targeting by keeping AID concentrations low. AID is phosphorylated at multiple sites, inducing the molecule to associate with RPA(replication protein A) which targets AID to both the V and S regions of the immunoglobulin gene. However the mechanism by which AID binds specifically to the V region over other loci is still unknown. RPA can only bind to single stranded DNA segments and therefore relies on either transcription bubbles or single stranded bubbles formed by other means to bind AID to its DNA substrate. AID may move along with the transcription bubble as it moves down the DNA. Once bound, AID deaminates Cytidine, converting the molecule into uridine.

Bacterial infection of AID