Simian virus 40 (SV40) is a monkey virus with tumorigenic potential in rodents and is associated with several types of human cancers, including lymphomas. A related Merkel cell polyomavirus causes carcinoma in humans by expressing truncated large tumor antigen (LT), with truncations caused by APOBEC family of cytidine deaminase-induced mutations. AID (activation-induced cytidine deaminase), a member of the APOBEC family, is the initiator of the antibody diversification process known as somatic hypermutation and its aberrant expression and targeting is a frequent source of lymphomagenesis. In this study, we investigated whether AID could cause mutations in SV40 LT. We demonstrate that the SV40 enhancer has strong somatic hypermutation targeting activity in several cell types and that AID-induced mutations accumulate in SV40 LT in B cells and kidney cells and cause truncated LT expression in B cells. Our results argue that the ability of the SV40 enhancer to target somatic hypermutation to LT is a potential source of LT truncation events that could contribute to tumorigenesis in various cell types, thereby linking SV40 infection with malignant development through a novel mutagenic pathway.
- MeSH
- antigeny transformující polyomavirové genetika metabolismus MeSH
- antigeny virové nádorové genetika metabolismus MeSH
- B-lymfocyty virologie metabolismus imunologie MeSH
- buněčné linie MeSH
- cytidindeaminasa * genetika metabolismus MeSH
- infekce onkogenními viry genetika virologie MeSH
- karcinogeneze genetika MeSH
- lidé MeSH
- mutace MeSH
- opičí virus SV40 * genetika MeSH
- polyomavirové infekce genetika virologie MeSH
- somatická hypermutace imunoglobulinových genů genetika MeSH
- zesilovače transkripce * genetika MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Somatic hypermutation (SHM) drives the genetic diversity of Ig genes in activated B cells and supports the generation of Abs with increased affinity for Ag. SHM is targeted to Ig genes by their enhancers (diversification activators [DIVACs]), but how the enhancers mediate this activity is unknown. We show using chicken DT40 B cells that highly active DIVACs increase the phosphorylation of RNA polymerase II (Pol II) and Pol II occupancy in the mutating gene with little or no accompanying increase in elongation-competent Pol II or production of full-length transcripts, indicating accumulation of stalled Pol II. DIVAC has similar effect also in human Ramos Burkitt lymphoma cells. The DIVAC-induced stalling is weakly associated with an increase in the detection of ssDNA bubbles in the mutating target gene. We did not find evidence for antisense transcription, or that DIVAC functions by altering levels of H3K27ac or the histone variant H3.3 in the mutating gene. These findings argue for a connection between Pol II stalling and cis-acting targeting elements in the context of SHM and thus define a mechanistic basis for locus-specific targeting of SHM in the genome. Our results suggest that DIVAC elements render the target gene a suitable platform for AID-mediated mutation without a requirement for increasing transcriptional output.
- MeSH
- aktivace lymfocytů MeSH
- Burkittův lymfom genetika imunologie MeSH
- cytidindeaminasa genetika MeSH
- genetická transkripce MeSH
- imunoglobuliny genetika metabolismus MeSH
- kur domácí MeSH
- lidé MeSH
- mutace genetika MeSH
- mutageneze cílená MeSH
- podskupiny B-lymfocytů imunologie MeSH
- ptačí proteiny genetika metabolismus MeSH
- RNA-polymerasa II genetika metabolismus MeSH
- rozmanitost protilátek MeSH
- somatická hypermutace imunoglobulinových genů MeSH
- zesilovače transkripce genetika MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, N.I.H., Extramural MeSH
Squamous cell carcinoma of the penis (PSC) is a rare disease with limited information on the molecular events leading to malignant transformation. In a third of PSC cases, presence of human papilloma virus (HPV) is found. The APOBEC3 family of proteins is known to play a significant role in defense against HPV infection, but their role in PSC is largely unknown. In this study, we aim to assess mRNA expression levels of APOBEC3 family members in HPV+ and HPV- PSC to get insight into their association with clinicopathological features and to evaluate their prognostic impact. Expression levels of six APOBEC3 family members in tissue from 50 patients with PSC were determined by RT-PCR and correlated with clinical and histopathological features. Lower expression of APOBEC3A, APOBEC3B, and APOBEC3C was observed in advanced PSC stages. Except for APOBEC3D, HPV+ samples showed higher expression of APOBEC3s compared to HPV- samples. In univariate analyses, APOBEC3A and APOBEC3C expression tended to be associated with disease-free survival and APOBEC3A expression with overall survival; however, multivariable analyses failed to confirm these associations with outcome. More extensive external validation and functional laboratory studies are needed to evaluate further their role in PSC development and progression.
- MeSH
- cytidindeaminasa genetika MeSH
- deaminasy APOBEC MeSH
- infekce papilomavirem * komplikace genetika MeSH
- lidé MeSH
- Papillomaviridae genetika MeSH
- penis patologie MeSH
- prognóza MeSH
- spinocelulární karcinom * patologie MeSH
- vedlejší histokompatibilní antigeny genetika MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Somatic hypermutation (SHM) introduces point mutations into immunoglobulin (Ig) genes but also causes mutations in other parts of the genome. We have used lentiviral SHM reporter vectors to identify regions of the genome that are susceptible ("hot") and resistant ("cold") to SHM, revealing that SHM susceptibility and resistance are often properties of entire topologically associated domains (TADs). Comparison of hot and cold TADs reveals that while levels of transcription are equivalent, hot TADs are enriched for the cohesin loader NIPBL, super-enhancers, markers of paused/stalled RNA polymerase 2, and multiple important B cell transcription factors. We demonstrate that at least some hot TADs contain enhancers that possess SHM targeting activity and that insertion of a strong Ig SHM-targeting element into a cold TAD renders it hot. Our findings lead to a model for SHM susceptibility involving the cooperative action of cis-acting SHM targeting elements and the dynamic and architectural properties of TADs.
- MeSH
- cytidindeaminasa genetika metabolismus MeSH
- HEK293 buňky MeSH
- Lentivirus MeSH
- lidé MeSH
- mutace genetika MeSH
- nádorové buněčné linie MeSH
- plazmidy genetika MeSH
- RNA-polymerasa II genetika metabolismus MeSH
- somatická hypermutace imunoglobulinových genů genetika MeSH
- zesilovače transkripce genetika MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, N.I.H., Extramural MeSH
Activation-induced cytidine deaminase (AID) is a mutator enzyme essential for somatic hypermutation (SHM) and class switch recombination (CSR) during effective adaptive immune responses. Its aberrant expression and activity have been detected in lymphomas, leukemias, and solid tumors. In chronic lymphocytic leukemia (CLL) increased expression of alternatively spliced AID variants has been documented. We used real-time RT-PCR to quantify the expression of AID and its alternatively spliced transcripts (AIDΔE4a, AIDΔE4, AIDivs3, and AIDΔE3E4) in 149 CLL patients and correlated this expression to prognostic markers including recurrent chromosomal aberrations, the presence of complex karyotype, mutation status of the immunoglobulin heavy chain variable gene, and recurrent mutations. We report a previously unappreciated association between higher AID transcript levels and trisomy of chromosome 12. Functional analysis of AID splice variants revealed loss of their activity with respect to SHM, CSR, and induction of double-strand DNA breaks. In silico modeling provided insight into the molecular interactions and structural dynamics of wild-type AID and a shortened AID variant closely resembling AIDΔE4, confirming its loss-of-function phenotype.
- MeSH
- alternativní sestřih * MeSH
- biologické modely * MeSH
- chronická lymfatická leukemie * enzymologie genetika patologie MeSH
- cytidindeaminasa * biosyntéza chemie genetika MeSH
- lidé středního věku MeSH
- lidé MeSH
- lidské chromozomy, pár 12 enzymologie genetika MeSH
- myši knockoutované MeSH
- myši MeSH
- nádorové proteiny * biosyntéza chemie genetika MeSH
- počítačová simulace MeSH
- regulace genové exprese enzymů * MeSH
- regulace genové exprese u nádorů * MeSH
- senioři MeSH
- simulace molekulární dynamiky MeSH
- trizomie * genetika patologie MeSH
- zvířata MeSH
- Check Tag
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- myši MeSH
- senioři MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
PURPOSE: To describe concentration versus time profiles of capecitabine and its metabolites 5'-DFUR, 5'-DFCR and 5-FU, depending on tablet formulation and on frequent and/or relevant genetic polymorphisms of cytidine deaminase, dihydropyrimidine dehydrogenase, thymidylate synthase and methylenetetrahydrofolate reductase (MTHFR). METHODS: In 46 cancer patients on chronic capecitabine treatment, who voluntarily participated in the study, individual therapeutic doses were replaced on four consecutive mornings by the study medication. The appropriate number of 500 mg test (T) or reference (R) capecitabine tablets was given in randomly allocated sequences TRTR or RTRT (replicate design). Average bioavailability was assessed by ANOVA. RESULTS: Thirty female and 16 male patients suffering from gastrointestinal or breast cancer (mean age 53.4 years; mean dose 1739 mg) were included. The T/R ratios for AUC0-t(last) and C max were 96.7 % (98 % CI 90.7-103.2 %) and 87.2 % (98 % CI 74.9-101.5 %), respectively. Within-subject variability for AUC0-t(last) and C max (coefficient of variation for R) was 16.5 and 30.2 %, respectively. Similar results were seen for all metabolites. No serious adverse events occurred. For the MTHFR C677T (rs1801133) genotype, an increasing number of 677C alleles showed borderline correlation with an increasing elimination half-life of capecitabine (p = 0.043). CONCLUSIONS: The extent of absorption was similar for T and R, but the rate of absorption was slightly lower for T. While such differences are not considered as clinically relevant, formal bioequivalence criteria were missed. A possible, probably indirect role of the MTHFR genotype in pharmacokinetics of capecitabine and/or 5-FU should be investigated in further studies.
- MeSH
- alely MeSH
- antimetabolity antitumorózní aplikace a dávkování farmakokinetika MeSH
- aplikace orální MeSH
- capecitabinum aplikace a dávkování farmakokinetika MeSH
- cytidindeaminasa genetika metabolismus MeSH
- deoxycytidin analogy a deriváty metabolismus MeSH
- dihydrouracildehydrogenasa (NADP) genetika metabolismus MeSH
- dospělí MeSH
- floxuridin metabolismus MeSH
- fluorouracil metabolismus MeSH
- genotyp MeSH
- játra enzymologie MeSH
- jednonukleotidový polymorfismus MeSH
- karboxylesterasa metabolismus MeSH
- lidé středního věku MeSH
- lidé MeSH
- metabolická aktivace genetika MeSH
- methylentetrahydrofolátreduktasa (NADPH2) genetika metabolismus MeSH
- nádorové proteiny metabolismus MeSH
- plocha pod křivkou MeSH
- prekurzory léčiv aplikace a dávkování farmakokinetika MeSH
- senioři MeSH
- tablety MeSH
- terapeutická ekvivalence MeSH
- thymidinfosforylasa metabolismus MeSH
- thymidylátsynthasa genetika metabolismus MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- multicentrická studie MeSH
- práce podpořená grantem MeSH
- randomizované kontrolované studie MeSH
- srovnávací studie MeSH
The motif DGYW/WRCH (Mh) and its frequently discussed simplified derivative GYW/WRC (Mhs) are involved in immunoglobulin (Ig) hypermutation. Both these motifs appear to be markedly shorter than the corresponding conventionally predicted minima of valid sequence lengths (MVSL). The same conclusion concerning both Mh and Mhs can also be obtained in the combined case including a less strict semi-empirically defined w-value and one nucleotide length tolerance related to MVSL. Such disagreement indicates considerably low information content in Mh and Mhs when evaluating these motifs as alphabetical structures (words). This fact raises a question of actually recognized structures (presumably longer than Mh and Mhs). Interestingly, both Mh and Mhs dimers or pairs of closely located Mh or Mhs achieve confirmation of length validity in the case of w=0.05, suggesting thus double-motif recognition as one of statistically consistent explanations. This possibility is also in agreement with the results of our model sequence study of mRNA derived from variable Ig gene sequences (rIgV) with respect to the most frequently occurring structures formed by motif overlaps in all model sequence sets. On the other hand, additional superior occurrence of motif pairs at a structurally important distance of a single DNA thread was found in the conserved domain (cd00099) related sequences of Elasmobranchii origin and less markedly in the corresponding human rIgV, but not in a randomly selected human subset of rIgV. The data are discussed with respect to statistical evaluation and structural properties of hypermutation motifs or the competent enzyme, i.e. activation-induced cytidine deaminase.
- MeSH
- aktivace enzymů MeSH
- aminokyselinové motivy MeSH
- cytidindeaminasa genetika MeSH
- geny pro imunoglobuliny MeSH
- lidé MeSH
- modely genetické MeSH
- mutační analýza DNA MeSH
- somatická hypermutace imunoglobulinových genů MeSH
- statistické modely MeSH
- terciární struktura proteinů MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
Abnormalities of the TP53 gene are associated with a particularly severe prognosis in patients with B-cell chronic lymphocytic leukemia (B-CLL). This tumor-suppressor is mostly inactivated by the deletion of one and point mutation of the other allele and has not been previously shown to be hypermutated in B-CLL. We identified two patients whose lymphocytes showed repeatedly an extensive proportion of TP53 mutated cells by FASAY analysis (the yeast functional assay) and harbored various TP53 mutations, mostly single-base substitutions, in individual cells. The mutation targeting exhibited characteristic traits of the somatic hypermutation process. In the first patient (harboring the unmutated IgVH locus) a significant bias to point mutations at CG pairs (21/25; P=0.009), their remarkable preference for the RGYW/WRCY motives (28%) and the highest expression of the activation-induced cytidine deaminase (AID) mRNA among the 34 tested B-CLL samples. In the second patient no CG bias was observed but the targeting of point mutations into the RGYW/WRCY motives was even more prominent here (7/16; 44%). Moreover, six out of eight point mutations affecting AT pairs were localized in the WA/TW motives, which are also characteristic for the somatic hypermutations. This patient, who was IgVH-mutated, already did not express any significant amount of the AID transcript. Our findings add a new aspect to the mosaic of the p53 mutability in B-CLL.
Hyper-IgM syndrom je zajímavou skupinou onemocnění, jejichž podkladem je porucha změny izotypu produkovaných protilátek. Studium poruch u jednotlivých typů hyper-IgM syndromu umožnilo velký rozvoj poznání principů class-switch rekombinace a somatické hypermutace. Přes obrovský pokrok v objasňování molekulárně genetické podstaty poruch zůstávají stále "bílá místa". Autoři prezentují souhrn posledních poznatků a doplňují kazuistiku jednoho pacienta s X-vázaným hyper-IgM syndromem v péči jejich pracoviště.
Hyper-IgM syndrome is a very interesting group of diseases, which have in common a class-switch defect of produced antibodies. The study of specific defects underlying different types of hyper-IgM syndrome allowed development of knowledge about class-switch and somatic hypermutation principles. Despite the tremendous advance in enlightening the molecular genetic basis of the defects, white areas still remain. The authors present an overview of latest knowledge together with a case-study of the single patient with X-linked hyper-IgM syndrome in the care of their department.
- MeSH
- agamaglobulinemie diagnóza etiologie farmakoterapie MeSH
- antigeny CD40 fyziologie MeSH
- bodová mutace MeSH
- cytidindeaminasa antagonisté a inhibitory fyziologie genetika MeSH
- DNA-glykosylasy fyziologie genetika chemie MeSH
- imunoglobulin M fyziologie genetika krev MeSH
- lidé MeSH
- přehledová literatura jako téma MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- Publikační typ
- kazuistiky MeSH
