scholarly journals Attenuating homologous recombination stimulates an AID-induced antileukemic effect

2013 ◽  
Vol 210 (5) ◽  
pp. 1021-1033 ◽  
Author(s):  
Kristin R. Lamont ◽  
Muneer G. Hasham ◽  
Nina M. Donghia ◽  
Jane Branca ◽  
Margaret Chavaree ◽  
...  
Keyword(s):  
Homologous Recombination ◽  
Somatic Hypermutation ◽  
Cytidine Deaminase ◽  
Lymphocytic Leukemia ◽  
Disulfonic Acid ◽  
Dna Breaks ◽  
New Paradigm ◽  
Lymphoid Leukemia ◽  
Antileukemic Effect ◽  
B Lymphoid

Activation-induced cytidine deaminase (AID) is critical in normal B cells to initiate somatic hypermutation and immunoglobulin class switch recombination. Accumulating evidence suggests that AID is also prooncogenic, inducing cancer-promoting mutations or chromosome rearrangements. In this context, we find that AID is expressed in >40% of primary human chronic lymphocytic leukemia (CLL) cases, consistent with other reports. Using a combination of human B lymphoid leukemia cells and mouse models, we now show that AID expression can be harnessed for antileukemic effect, after inhibition of the RAD51 homologous recombination (HR) factor with 4,4′-diisothiocyanatostilbene-2-2′-disulfonic acid (DIDS). As a proof of principle, we show that DIDS treatment inhibits repair of AID-initiated DNA breaks, induces apoptosis, and promotes cytotoxicity preferentially in AID-expressing human CLL. This reveals a novel antineoplastic role of AID that can be triggered by inhibition of HR, suggesting a potential new paradigm to treat AID-expressing tumors. Given the growing list of tumor types with aberrant AID expression, this novel therapeutic approach has potential to impact a significant patient population.

scholarly journals Induction of homologous recombination between sequence repeats by the activation induced cytidine deaminase (AID) protein

eLife ◽  
2014 ◽  
Vol 3 ◽  
Author(s):  
Jean-Marie Buerstedde ◽  
Noel Lowndes ◽  
David G Schatz
Keyword(s):  
Homologous Recombination ◽  
Somatic Hypermutation ◽  
Cytidine Deaminase ◽  
Holliday Junctions ◽  
Mutagenic Potential ◽  
Activation Induced Cytidine Deaminase ◽  
Invasion Analysis ◽  
Dna End Resection ◽  
Cytidine Deamination ◽  
End Resection

The activation induced cytidine deaminase (AID) protein is known to initiate somatic hypermutation, gene conversion or switch recombination by cytidine deamination within the immunoglobulin loci. Using chromosomally integrated fluorescence reporter transgenes, we demonstrate a new recombinogenic activity of AID leading to intra- and intergenic deletions via homologous recombination of sequence repeats. Repeat recombination occurs at high frequencies even when the homologous sequences are hundreds of bases away from the positions of AID-mediated cytidine deamination, suggesting DNA end resection before strand invasion. Analysis of recombinants between homeologous repeats yielded evidence for heteroduplex formation and preferential migration of the Holliday junctions to the boundaries of sequence homology. These findings broaden the target and off-target mutagenic potential of AID and establish a novel system to study induced homologous recombination in vertebrate cells.

scholarly journals Regulation of class switch recombination and somatic mutation by AID phosphorylation

2008 ◽  
Vol 205 (11) ◽  
pp. 2585-2594 ◽  
Author(s):  
Kevin M. McBride ◽  
Anna Gazumyan ◽  
Eileen M. Woo ◽  
Tanja A. Schwickert ◽  
Brian T. Chait ◽  
...  
Keyword(s):  
Somatic Mutation ◽  
Somatic Hypermutation ◽  
Cytidine Deaminase ◽  
Point Mutations ◽  
Class Switch Recombination ◽  
Variable Region ◽  
Dna Breaks ◽  
Class Switching ◽  
Class Switch

Activation-induced cytidine deaminase (AID) is a mutator enzyme that initiates somatic mutation and class switch recombination in B lymphocytes by introducing uracil:guanine mismatches into DNA. Repair pathways process these mismatches to produce point mutations in the Ig variable region or double-stranded DNA breaks in the switch region DNA. However, AID can also produce off-target DNA damage, including mutations in oncogenes. Therefore, stringent regulation of AID is required for maintaining genomic stability during maturation of the antibody response. It has been proposed that AID phosphorylation at serine 38 (S38) regulates its activity, but this has not been tested in vivo. Using a combination of mass spectrometry and immunochemical approaches, we found that in addition to S38, AID is also phosphorylated at position threonine 140 (T140). Mutation of either S38 or T140 to alanine does not impact catalytic activity, but interferes with class switching and somatic hypermutation in vivo. This effect is particularly pronounced in haploinsufficient mice where AID levels are limited. Although S38 is equally important for both processes, T140 phosphorylation preferentially affects somatic mutation, suggesting that posttranslational modification might contribute to the choice between hypermutation and class switching.

Clonal lineage analysis of CLL research samples by IGH chain sequencing to reveal novel CLL subgroups defined by ongoing class-switch recombination and somatic hypermutation.

2020 ◽  
Vol 38 (15_suppl) ◽  
pp. e20001-e20001
Author(s):  
Timothy Looney ◽  
Graeme Quest ◽  
Harriet Feilotter ◽  
Zadie Davis
Keyword(s):  
Somatic Hypermutation ◽  
Class Switch Recombination ◽  
Lymphocytic Leukemia ◽  
Splenic Marginal Zone Lymphoma ◽  
Clonal Lineage ◽  
Sequencing Data ◽  
Dna Breaks ◽  
Class Switch ◽  
Repertoire Sequencing ◽  
Lineage Analysis

e20001 Background: B cell somatic hypermutation (SHM) and class switch recombination (CSR) are mechanistically related but distinct processes requiring precisely targeted generation and repair of single and double strand DNA breaks. Within the context of chronic lymphocytic leukemia (CLL), the presence of ongoing SHM or CSR may reveal the functionality of DNA damage repair pathways, with potential relevance to therapeutic strategies involving the generation of DNA breaks or inhibition of DNA repair machinery. Here we apply clonal lineage analysis of IGH chain sequencing data to evaluate CSR and SHM in a cohort of CLL and splenic marginal zone lymphoma (SMZL) research samples. We present evidence of ongoing CSR and SHM in a significant subset of samples. Methods: Multiplex primers targeting the IGH framework 1 region and isotype region (Oncomine IGH-LR assay; detection of all nine isotypes) were used for IGH repertoire sequencing via the Ion Gene Studio S5 from 25ng peripheral blood total RNA derived from 63 individuals with CLL and 4 individuals with SMZL. Clonotyping and clonal lineage analysis was performed by Ion Reporter, whereby clonal lineages are defined as sets of unique rearrangements having a shared variable and joining gene, the same CDR3 length, and a minimum CDR3 nucleotide similarity of 85%. Ongoing CSR was defined as the presence of IgM/IgD and at least one switched isotype (IgG, IgA, or IgE), or a combination of switched isotypes, within the same lineage. Ongoing SHM was defined as the presence of subclones that differ within the VDJ region sequence compared to other clonal lineage members. Results: 11/68 cases showed evidence of ongoing CSR or SHM. Of the 57 cases showing no evidence of ongoing CSR or SHM, variable gene mutation analysis revealed the presence of three distinct subgroups having either no SHM, intermediate SHM (average 98% sequence identity) or high SHM < 94% identity). 3 of 4 SMZL cases showed evidence of ongoing CSR or SHM. Conclusions: These results reveal previously underappreciated heterogeneity within CLL and suggest the subdivision of CLL based on a combination of IGHV mutation level and presence of ongoing SHM or CSR. The described heterogeneity may serve as a valuable criterion for stratifying CLL patients in the future.

scholarly journals RNA-binding motifs of hnRNP K are critical for induction of antibody diversification by activation-induced cytidine deaminase

2020 ◽  
Vol 117 (21) ◽  
pp. 11624-11635
Author(s):  
Ziwei Yin ◽  
Maki Kobayashi ◽  
Wenjun Hu ◽  
Koichi Higashi ◽  
Nasim A. Begum ◽  
...  
Keyword(s):  
Rna Binding ◽  
Somatic Hypermutation ◽  
Cytidine Deaminase ◽  
Binding Capacity ◽  
Dependent Manner ◽  
Dna Breaks ◽  
Nuclear Retention ◽  
Binding Motifs ◽  
Hnrnp K ◽  
Activation Induced Cytidine Deaminase

Activation-induced cytidine deaminase (AID) is the key enzyme for class switch recombination (CSR) and somatic hypermutation (SHM) to generate antibody memory. Previously, heterogeneous nuclear ribonucleoprotein K (hnRNP K) was shown to be required for AID-dependent DNA breaks. Here, we defined the function of major RNA-binding motifs of hnRNP K, GXXGs and RGGs in the K-homology (KH) and the K-protein-interaction (KI) domains, respectively. Mutation of GXXG, RGG, or both impaired CSR, SHM, andcMyc/IgHtranslocation equally, showing that these motifs were necessary for AID-dependent DNA breaks. AID–hnRNP K interaction is dependent on RNA; hence, mutation of these RNA-binding motifs abolished the interaction with AID, as expected. Some of the polypyrimidine sequence-carrying prototypical hnRNP K-binding RNAs, which participate in DNA breaks or repair bound to hnRNP K in a GXXG and RGG motif-dependent manner. Mutation of the GXXG and RGG motifs decreased nuclear retention of hnRNP K. Together with the previous finding that nuclear localization of AID is necessary for its function, lower nuclear retention of these mutants may worsen their functional deficiency, which is also caused by their decreased RNA-binding capacity. In summary, hnRNP K contributed to AID-dependent DNA breaks with all of its major RNA-binding motifs.

Chronic lymphocytic leukemia B cells expressing AID display dissociation between class switch recombination and somatic hypermutation

Blood ◽  
2003 ◽  
Vol 101 (10) ◽  
pp. 4029-4032 ◽  
Author(s):  
Pablo Oppezzo ◽  
Françoise Vuillier ◽  
Yuri Vasconcelos ◽  
Gérard Dumas ◽  
Christian Magnac ◽  
...  
Keyword(s):  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
Gene Product ◽  
Mode Of Action ◽  
Somatic Hypermutation ◽  
Cytidine Deaminase ◽  
Class Switch Recombination ◽  
Lymphocytic Leukemia ◽  
Class Switch ◽  
Activation Induced Cytidine Deaminase

Abstract In B cells, somatic hypermutation (SHM) and class switch recombination (CSR) depend on the activation-induced cytidine deaminase (AID) gene product, although the precise mode of action of AID remains unknown. Because some chronic lymphocytic leukemia (CLL) B cells can undergo CSR without SHM, it constitutes a useful model to dissect AID function. In this work, we have studied AID expression, the presence of mutations in the preswitch μ DNA region, CSR, and the SHM in 65 CLL patients. Our results demonstrate that unmutated CLL B cells can constitutively express AID and that AID expression is associated with the presence of mutations in the preswitch region and in clonally related isotype-switched transcripts. They also demonstrate that in CLL without constitutive AID expression, AID induction on stimulation results in preswitch mutations and the CSR process. Our results show a dissociation between SHM and CSR in CLL and suggest that, in this disease, AID would require additional help for carrying out the SHM process.

scholarly journals Genetic loss of the ubiquitin ligase RNF126, an AID interacting partner and modifier, affects strand targeting during somatic hypermutation of antibody genes

2018 ◽  
Author(s):  
Nicholas Economos ◽  
Rebecca K Delker ◽  
Pete Stavropoulos ◽  
F. Nina Papavasiliou
Keyword(s):  
Somatic Hypermutation ◽  
Cytidine Deaminase ◽  
Point Mutations ◽  
Variable Region ◽  
Dna Breaks ◽  
Switch Region ◽  
Post Translational Modifications ◽  
Class Switch ◽  
Double Stranded Dna ◽  
Activation Induced Cytidine Deaminase

AbstractActivation-induced cytidine deaminase (AID) initiates somatic hypermutation (SHM) and class switch recombination (CSR) in B lymphocytes by catalyzing the introduction of deoxyuracil: deoxyguanine mismatches into the DNA of the transcribed Ig locus. Repair pathways then process these mismatches to produce point mutations in the Ig variable region or double-stranded DNA breaks in the switch region followed by deletional recombination. It has been suggested that post-translational modifications on AID mediate a number of these different decisions, ranging from global targeting (Ig vs the genome), local targeting (variable vs switch region; transcribed vs non-transcribed strand) as well as process-appropriate DNA repair. Here we demonstrate that absence of RNF126, an E3 ligase shown to mono-ubiquitylate AID, results in a specific strand targeting defect in SHM, producing substantial G>C bias; strickingly, loss of RNF126 was also associated with tandem indels within the variable region (JH4 intron) but only a slight increase in the types of chromosomal translocations that are characteristic of deregulated AID. Conversely, these findings suggest that mono-ubiquitination of AID, likely in situ, is necessary for the proper removal of the protein from the non-transcribed strand, thus producing both optimal patterns of SHM and also limiting the number of indels within the target locus.

Activation-induced cytidine deaminase in chronic lymphocytic leukemia B cells: expression as multiple forms in a dynamic, variably sized fraction of the clone

Blood ◽  
2003 ◽  
Vol 102 (9) ◽  
pp. 3333-3339 ◽  
Author(s):  
Emilia Albesiano ◽  
Bradley T. Messmer ◽  
Rajendra N. Damle ◽  
Steven L. Allen ◽  
Kanti R. Rai ◽  
...  
Keyword(s):  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
Somatic Hypermutation ◽  
Splice Variants ◽  
Cytidine Deaminase ◽  
Prognostic Indicator ◽  
Lymphocytic Leukemia ◽  
Leukemic Cells ◽  
Activation Induced Cytidine Deaminase

AbstractThe degree of somatic mutation of immunoglobulin variable (Ig V) region genes is an important prognostic indicator of clinical course and outcome in B-cell chronic lymphocytic leukemia (B-CLL), although the reason for this association remains unclear. Furthermore, some B-CLL cells continue to acquire Ig V gene mutations after the transforming event. Because activation-induced cytidine deaminase (AID) is an essential component of the canonical somatic hypermutation process in healthy B cells, its expression in B-CLL is potentially relevant to the disease. We detected full-length AID transcripts and 3 splice variants by conventional reverse transcription polymerase chain reaction (RT-PCR) in approximately 40% of the cases examined. More sensitive real-time quantitative PCR detected AID transcripts in virtually all B-CLL samples tested, although the range of transcript levels was very large between different cases and varied within individual cases over time. Limiting dilution assays revealed that AID expression was restricted to a small fraction of the leukemic cells in the blood. However, this small fraction is not unique in its ability to express AID, because in vitro stimulation of B-CLL cells with appropriate stimuli significantly increased the fraction of AID-expressing cells. These data suggest that AID-mediated DNA alterations may occur in a variably sized, minor subset of B-CLL cells at any given time.

scholarly journals A primary immunodeficiency characterized by defective immunoglobulin class switch recombination and impaired DNA repair

2007 ◽  
Vol 204 (5) ◽  
pp. 1207-1216 ◽  
Author(s):  
Sophie Péron ◽  
Qiang Pan-Hammarström ◽  
Kohsuke Imai ◽  
Likun Du ◽  
Nadine Taubenheim ◽  
...  
Keyword(s):  
Dna Repair ◽  
Somatic Hypermutation ◽  
Cytidine Deaminase ◽  
Class Switch Recombination ◽  
Cd40 Ligand ◽  
Dna Breaks ◽  
Immunoglobulin Class ◽  
Class Switch ◽  
Double Stranded Dna ◽  
Cd40 Activation

Immunoglobulin class switch recombination (CSR) deficiencies are rare primary immunodeficiencies, characterized by a lack of switched isotype (IgG, IgA, or IgE) production, variably associated with abnormal somatic hypermutation (SHM). Deficiencies in CD40 ligand, CD40, activation-induced cytidine deaminase, and uracil-N-glycosylase may account for this syndrome. We previously described another Ig CSR deficiency condition, characterized by a defect in CSR downstream of the generation of double-stranded DNA breaks in switch (S) μ regions. Further analysis performed with the cells of five affected patients showed that the Ig CSR deficiency was associated with an abnormal formation of the S junctions characterized by microhomology and with increased cell radiosensitivity. In addition, SHM was skewed toward transitions at G/C residues. Overall, these findings suggest that a unique Ig CSR deficiency phenotype could be related to an as-yet-uncharacterized defect in a DNA repair pathway involved in both CSR and SHM events.

Expression of the AID protein in normal and neoplastic B cells

Blood ◽  
2004 ◽  
Vol 104 (10) ◽  
pp. 3318-3325 ◽  
Author(s):  
Laura Pasqualucci ◽  
Roberta Guglielmino ◽  
Jane Houldsworth ◽  
Jessica Mohr ◽  
Said Aoufouchi ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Somatic Hypermutation ◽  
Cytidine Deaminase ◽  
Variable Region ◽  
Affinity Maturation ◽  
Lymphocytic Leukemia ◽  
Antibody Affinity ◽  
Disease Subtype ◽  
Activation Induced Cytidine Deaminase

Abstract Somatic hypermutation (SHM) targets primarily the immunoglobulin variable region (IgV) genes in germinal center (GC) B cells, thereby allowing antibody affinity maturation. A malfunction of SHM, termed aberrant somatic hypermutation (ASHM), was found in about 50% of diffuse large B-cell lymphomas (DLBCLs), leading to mutations in the 5′ sequences of multiple genes, including oncogenes. Although the SHM mechanism is largely unknown, it was shown to require the activation-induced cytidine deaminase (AID) gene. AID mRNA is expressed in GC B cells and GC-derived lymphomas, but the pattern of expression of the AID protein is not known. Using 2 specific antibodies, here we show that the AID protein can be detected in GC centroblasts and their transformed counterpart (Burkitt lymphoma) but not in pre-GC B cells and post-GC neoplasms, including B-cell chronic lymphocytic leukemia and multiple myeloma. DLBCLs displayed variable levels of AID expression, which did not correlate with IgV ongoing hypermutation, ASHM, or disease subtype. Finally, both in normal and malignant B cells the AID protein appeared predominantly localized in the cytoplasm. These results indicate that the AID protein is specifically expressed in normal and transformed GC B cells; nonetheless, its predominantly cytoplasmic localization suggests that additional mechanisms may regulate its function and may be altered during lymphomagenesis. (Blood. 2004;104:3318-3325)

scholarly journals Alternative splicing regulates activation-induced cytidine deaminase (AID): implications for suppression of AID mutagenic activity in normal and malignant B cells

Blood ◽  
2008 ◽  
Vol 112 (12) ◽  
pp. 4675-4682 ◽  
Author(s):  
Xiaosheng Wu ◽  
Jaime R. Darce ◽  
Sook Kyung Chang ◽  
Grzegorz S. Nowakowski ◽  
Diane F. Jelinek
Keyword(s):  
B Cells ◽  
Somatic Hypermutation ◽  
Splice Variants ◽  
Cytidine Deaminase ◽  
Mutagenic Activity ◽  
Enzyme Activation ◽  
Lymphocytic Leukemia ◽  
Human B Cells ◽  
Activation Induced Cytidine Deaminase ◽  
The Individual

Abstract The mutagenic enzyme activation-induced cytidine deaminase (AID) is required for immunoglobulin class switch recombination (CSR) and somatic hypermutation (SHM) in germinal center (GC) B cells. Deregulated expression of AID is associated with various B-cell malignancies and, currently, it remains unclear how AID activity is extinguished to avoid illegitimate mutations. AID has also been shown to be alternatively spliced in malignant B cells, and there is limited evidence that this also occurs in normal blood B cells. The functional significance of these splice variants remains unknown. Here we show that normal GC human B cells and blood memory B cells similarly express AID splice variants and show for the first time that AID splicing variants are singly expressed in individual normal B cells as well as malignant B cells from chronic lymphocytic leukemia patients. We further demonstrate that the alternative AID splice variants display different activities ranging from inactivation of CSR to inactivation or heightened SHM activity. Our data therefore suggest that CSR and SHM are differentially switched off by varying the expression of splicing products of AID at the individual cell level. Most importantly, our findings suggest a novel tumor suppression mechanism by which unnecessary AID mutagenic activities are promptly contained for GC B cells.

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