Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “DNA editing”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 37 records · Page 2Linked to original sources
PubMed2026

DNA-guided CRISPR-Cas12 for cellular RNA targeting.

Here, we present ΨDNA, a DNA-based guide that enables RNA targeting by Cas12 nucleases, overcoming the traditional reliance on RNA-guided systems. We engineer ΨDNA to mimic a CRISPR RNA (crRNA) scaffold in reverse orientation, allowing AsCas12a and Cas12i1 to recognize RNA and trigger strong single-stranded DNA trans-cleavage for sensitive detection of diverse RNA species, including 100% accurate hepatitis C virus RNA detection in clinical samples. ΨDNA also achieves 70-95% multiplex knockdown of endogenous intracellular RNA transcripts through ribosome stalling across multiple human cell lines. Mechanistic studies reveal that activity depends on a stem loop that stabilizes a catalytically competent Cas12-ΨDNA-RNA complex. Lastly, codelivery of crRNA and ΨDNA enables simultaneous DNA editing and RNA knockdown with a single effector and modular fusions of different enzymes to AsCas12a extend ΨDNA to RNase H-mediated RNA degradation and METTL3-based epitranscriptomic editing. Together, ΨDNA guides constitute an adaptable toolkit that extends Cas12 systems beyond genome editing and diagnostics to enable precise, programmable control of cellular transcriptomes and their epitranscriptomic marks.

Journal Article↗
PubMed2001

Editors and editing of anti-DNA receptors.

Receptor editing is a means by which immature bone marrow B cells can become self-tolerant. Rearrangements of heavy (H) and/or light (L) chain genes are induced by encounter with autoantigens to change the specificity from self to nonself. We have developed site-directed transgenic mice (sd-tg) whose transgenes code for the H chain of antibodies that bind DNA. B cells that express the transgenic H chain associate mainly with four of the 93 functional Vkappa genes of the mouse. Numerous aspartate residues that might inhibit DNA binding by the V(H) domain distinguish these L chain Vkappa sequences, but engaging these Vkappa editors often requires multiple rearrangements. Among the edited B cells is a subset of multispecific cells that express multiple receptors. One consequence of multispecificity is partial autoreactivity; these multispecific B cells may contribute to autoimmunity.

Amino Acid Sequence↗
PubMed1990

A model for RNA editing in kinetoplastid mitochondria: "guide" RNA molecules transcribed from maxicircle DNA provide the edited information.

A class of small RNA molecules possibly involved in RNA editing is present in the mitochondrion of Leishmania tarentolae. These "guide" RNA (gRNA) molecules are encoded in intergenic regions of the mitochondrial maxicircle DNA and contain sequences that represent precise complementary versions of the mature mRNAs within the edited regions. In addition, the 5' portions of several gRNAs can form hybrids with mRNAs just 3' of the preedited region. A model is presented in which a partial hybrid formed between the gRNA and preedited mRNA is substrate for multiple cycles of cleavage, addition or deletion of uridylates, and religation, eventually resulting in a complete hybrid between the gRNA and the mature edited mRNA.

Animals↗
PubMed1996

Developmental- and tissue-specificity of RNA editing in mitochondria of suspension-cultured maize cells and seedlings.

C to U editing of apt9, nad3, and cox2 mRNAs was investigated in maize seedlings at various developmental stages as well as in suspension-cultured cells. Heterogeneity of mRNAs that result from incomplete editing was analyzed for each gene and from five tissues or developmental conditions. The editing status of approximately 30 cDNA clones was determined by digestion with a restriction enzyme that discriminates between unedited and edited DNA sequences. The atp9 and spliced cox2 cDNAs were essentially completely edited in all samples examined. Analysis of three editing sites of nad3 cDNAs indicated that incompletely edited cDNAs were detected in all tissues and treatments with a temporal increase in the overall editing status, from 50% at 3 days to about 75% at 7 days. These results indicate that incompletely edited mRNAs are prevalent for some plant mitochondrial genes, and can change with developmental or growth conditions.

Amino Acid Sequence↗
PubMed2005

Protecting APOBEC3G: a potential new target for HIV drug discovery.

Human immune cells possess a built-in mechanism that could potentially block the replication of retroviruses such as HIV-1. This protective mechanism centers on apolipoprotein B mRNA-editing enzyme, catalytic polypeptide-like 3G (APOBEC3G), a DNA-editing enzyme produced by host cells infected by certain retroviruses that is then incorporated into virions. Due to its cytidine deaminase activity, APOBEC3G is able to mutate the minus-strand DNA formed during reverse transcription. These events ultimately halt completion of the HIV life cycle. Unfortunately, HIV-1 encodes a protein termed virion infectivity factor (Vif) that specifically suppresses the activity of APOBEC3G. Vif achieves this effect by depleting the intracellular stores of APOBEC3G, thus making this antiviral enzyme unavailable for incorporation into budding virions. APOBEC3G depletion involves the recruitment of a specific E3 ligase complex by Vif leading to the polyubiquitylation and proteasome-mediated degradation of this enzyme. The potent activity of APOBEC3G has led to considerable interest in the identification of small molecules that interrupt the Vif-induced degradative process.

APOBEC-3G Deaminase↗
PubMed2004

Editing anti-DNA B cells by Vlambdax.

Receptor editing is performed by replacement of Vkappa genes that contribute to autoreactivity. In addition, the Ckappa locus can be deleted by Vkappa rearrangement to intronic or 3' of Ckappa RS sequences (also referred to as kappa deletion elements). B cells that delete the Ckappa can then express lambda light chains. However, the lambda locus, either of man or mouse, does not allow V gene replacement. Nor does it appear to be deleted. Therefore, editing of autoreactive lambda B cells may require alternative pathways. We have found that in anti-DNA heavy chain transgenic mice (tgs) VH3H9/56R, B cells that express anti-DNA receptors comprised of lambda1 in association with an anti-DNA heavy chain often coexpress a kappa chain that prevents DNA binding. We speculate that such isotypically included cells may have low anti-DNA receptor densities, a feature that may lead to self-tolerance. Here we describe a mechanism of preventing DNA binding by expression of a rarely used member of the Vlambda family, Vlambdax. The lambdax B cells of the tgs also express CD25 and may represent B cells that have exhausted light chain editing possibilities.

Animals↗
PubMed2006

Functional central polypurine tract provides downstream protection of the human immunodeficiency virus type 1 genome from editing by APOBEC3G and APOBEC3B.

Lentiviruses utilize two polypurine tracts for initiation of plus-strand viral DNA synthesis. We have examined to what extent human immunodeficiency virus type 1 plus-strand initiation at the central polypurine tract (cPPT) could protect the viral genome from DNA editing by APOBEC3G and APOBEC3B. The presence of a functional cPPT, but not of a mutated cPPT, extensively reduced editing by both APOBEC3G and APOBEC3B of sequences downstream, but not upstream, of the cPPT, with significant protection observed as far as 400 bp downstream. Thus, in addition to other potential functions, the cPPT could help protect lentiviruses from editing by cytidine deaminases of the APOBEC family.

APOBEC-3G Deaminase↗
PubMed2006

HIV accomplices and adversaries in macrophage infection.

Cell surface and intracellular proteins in macrophages influence various steps in the life cycle of lentiviruses. Characterization of these restriction and/or cofactors is essential to understanding how macrophages become unwitting HIV hosts and in fact, can coexist with a heavy viral burden. Although many of the cellular pathways co-opted by HIV in macrophages mimic those seen in CD4+ T cells, emerging evidence reveals cellular constituents of the macrophage, which may be uniquely usurped by HIV. For example, in addition to CD4 and CCR5, membrane annexin II facilitates early steps in infection of macrophages, but not in T cells. Blockade of this pathway effectively diminishes macrophage infection. Viral binding engages a macrophage-centric signaling pathway and a transcriptional profile, including genes such as p21, which benefit the virus. Once inside the cell, multiple host cell molecules are engaged to facilitate virus replication and assembly. Although the macrophage is an enabler, it also possesses innate antiviral mechanisms, including apolipoprotein B mRNA-editing enzyme-catalytic polypeptide-like 3G (APOBEC3) family DNA-editing enzymes to inhibit replication of HIV. Differential expression of these enzymes, which are largely neutralized by HIV to protect its rebirth, is associated with resistance or susceptibility to the virus. Higher levels of the cytidine deaminases endow potential HIV targets with a viral shield, and IFN-alpha, a natural inducer of macrophage APOBEC expression, renders macrophages tougher combatants to HIV infection. These and other manipulatable pathways may give the macrophage a fighting chance in its battle against the virus.

HIV Infections↗
PubMed2006

Twin gradients in APOBEC3 edited HIV-1 DNA reflect the dynamics of lentiviral replication.

The human immunodeficiency virus (HIV) Vif protein blocks incorporation of two host cell cytidine deaminases, APOBEC3F and 3G, into the budding virion. Not surprisingly, on a vif background nascent minus strand DNA can be extensively edited leaving multiple uracil residues. Editing occurs preferentially in the context of TC (GA on the plus strand) and CC (GG) depending on the enzyme. To explore the distribution of APOBEC3F and -3G editing across the genome, a product/substrate ratio (AA + AG)/(GA + GG) was computed for a series of 30 edited genomes present in the data bases. Two highly polarized gradients were noted each with maxima just 5' to the central polypurine tract (cPPT) and LTR proximal polypurine tract (3'PPT). The gradients are in remarkable agreement with the time the minus strand DNA remains single stranded. In vitro analyses of APOBEC3G deamination of nascent cDNA spanning the two PPTs showed no pronounced dependence on the PPT RNA:DNA heteroduplex ruling out the competing hypothesis of a PPT orientation effect. The degree of hypermutation varied smoothly among genomes indicating that the number of APOBEC3 molecules packaged varied considerably.

APOBEC Deaminases↗
PubMed1993

An experimentally derived data set constructed for testing large-scale DNA sequence assembly algorithms.

A data set consisting of DNA sequences from a large-scale shotgun DNA cloning and sequencing project has been collected and posted for public release. The purpose is to propose a standard genomic DNA sequencing data set by which various algorithms and implementations can be tested. This set of data is divided into two subsets, one containing raw DNA sequence data (1023 clones) and the other consisting of the corresponding partially refined or edited DNA sequence data (820 clones). Suggested criteria or guidelines for this data refinement are presented so that algorithms for preprocessing and screening raw sequences may be developed. Development of such preprocessing, screening, aligning, and assembling algorithms will expedite large-scale DNA sequencing projects so that the complete unambiguous consensus DNA sequences will be made available to the general research community in a quicker manner. Smaller scale routine DNA sequencing projects will also be greatly aided by such computational efforts.

Algorithms↗
PubMed2005

Hyper-immunoglobulin M syndromes caused by intrinsic B-lymphocyte defects.

Hyper-immunoglobulin M (IgM) syndromes are primary immunodeficiencies characterized by normal or elevated serum IgM levels with the absence of other isotypes, pinpointing to a defect in the Ig class switch recombination (CSR). The delineation of hyper-IgM syndromes made it possible to better define the mechanisms underlying the two major events of antibody maturation in humans, CSR and introduction of somatic hypermutation (SHM) in the variable region of immunoglobulins. The description of the activation-induced cytidine deaminase (AID) deficiency, characterized by a defect in both CSR and SHM, demonstrated for the first time that this molecule acts as a master player in the antigen-induced Ig gene-modification events responsible for both CSR and SHM. However, deleterious mutations located in the C-terminus lead to a CSR defect without affecting SHM, providing evidence for a role of AID in CSR distinct from the cytidine deaminase activity, likely by binding to a specific CSR cofactor. Molecular causes of two other hyper-IgM conditions have not yet been defined. However, they may be caused by either a defect in AID targeting on S regions or a CSR-specific DNA-repair defect. The mechanism of action of AID remains somewhat debated, but the observation that uracil-DNA-glycosylase deficiency leads to a severe hyper-IgM syndrome strongly argues in favor of a DNA-editing activity of AID.

B-Lymphocytes↗
PubMed2001

Oxygen free radical damage to DNA. Translesion synthesis by human DNA polymerase eta and resistance to exonuclease action at cyclopurine deoxynucleoside residues.

Cyclopurine deoxynucleosides are common DNA lesions generated by exposure to reactive oxygen species under hypoxic conditions. The S and R diastereoisomers of cyclodeoxyadenosine on DNA were investigated separately for their ability to block 3' to 5' exonucleases. The mammalian DNA-editing enzyme DNase III (TREX1) was blocked by both diastereoisomers, whereas only the S diastereoisomer was highly efficient in preventing digestion by the exonuclease function of T4 DNA polymerase. Digestion in both cases was frequently blocked one residue before the modified base. Oligodeoxyribonucleotides containing a cyclodeoxyadenosine residue were further employed as templates for synthesis by human DNA polymerase eta (pol eta). pol eta could catalyze translesion synthesis on the R diastereoisomer of cyclodeoxyadenosine. On the S diastereoisomer, pol eta could catalyze the incorporation of one nucleotide opposite the lesion but could not continue elongation. Although pol eta preferentially incorporated dAMP opposite the R diastereoisomer, elongation continued only when dTMP was incorporated, suggesting bypass of this lesion by pol eta with reasonable fidelity. With the S diastereoisomer, pol eta mainly incorporated dAMP or dTMP opposite the lesion but could not elongate even after incorporating a correct nucleotide. These data suggest that the S diastereoisomer may be a more cytotoxic DNA lesion than the R diastereoisomer.

DNA Damage↗
PubMed1992

DNA Translator and Aligner: HyperCard utilities to aid phylogenetic analysis of molecules.

DNA Translator and Aligner are molecular phylogenetics HyperCard stacks for Macintosh computers. They manipulate sequence data to provide graphical gene mapping, conversions, translations and manual multiple-sequence alignment editing. DNA Translator is able to convert documented GenBank or EMBL documented sequences into linearized, rescalable gene maps whose gene sequences are extractable by clicking on the corresponding map button or by selection from a scrolling list. Provided gene maps, complete with extractable sequences, consist of nine metazoan, one yeast, and one ciliate mitochondrial DNAs and three green plant chloroplast DNAs. Single or multiple sequences can be manipulated to aid in phylogenetic analysis. Sequences can be translated between nucleic acids and proteins in either direction with flexible support of alternate genetic codes and ambiguous nucleotide symbols. Multiple aligned sequence output from diverse sources can be converted to Nexus, Hennig86 or PHYLIP format for subsequent phylogenetic analysis. Input or output alignments can be examined with Aligner, a convenient accessory stack included in the DNA Translator package. Aligner is an editor for the manual alignment of up to 100 sequences that toggles between display of matched characters and normal unmatched sequences. DNA Translator also generates graphic displays of amino acid coding and codon usage frequency relative to all other, or only synonymous, codons for approximately 70 select organism-organelle combinations. Codon usage data is compatible with spreadsheet or UWGCG formats for incorporation of additional molecules of interest. The complete package is available via anonymous ftp and is free for non-commercial uses.

Amino Acid Sequence↗
PubMed2004

Crystallographic structure of the nuclease domain of 3'hExo, a DEDDh family member, bound to rAMP.

A human 3'-5'-exoribonuclease (3'hExo) has recently been identified and shown to be responsible for histone mRNA degradation. Functionally, 3'hExo and a stem-loop binding protein (SLBP) target opposite faces of a unique highly conserved stem-loop RNA scaffold towards the 3' end of histone mRNA, which is composed of a 6 bp stem and a 4 nt loop, followed by an ACCCA sequence. Its Caenorhabditis elegans homologue, ERI-1, has been shown to degrade small interfering RNA in vitro and to function as a negative regulator of RNA interference in neuronal cells. We have determined the structure of the nuclease domain (Nuc) of 3'hExo complexed with rAMP in the presence of Mg2+ at 1.6 A resolution. The Nuc domain adopts an alpha/beta globular fold, with four acidic residues coordinating a binuclear metal cluster within the active site, whose topology is related to DEDDh exonuclease family members, despite a very low level of primary sequence identity. The two magnesium cations in the Nuc active site are coordinated to D134, E136, D234 and D298, and together with H293, which can potentially act as a general base, provide a platform for hydrolytic cleavage of bound RNA in the 3' --> 5' direction. The bound rAMP is positioned within a deep active-site pocket, with its purine ring close-packed with the hydrophobic F185 and L189 side-chains and its sugar 2'-OH and 3'-OH groups hydrogen bonded to backbone atoms of Nuc. There are striking similarities between the active sites of Nuc and epsilon186, an Escherichia coli DNA polymerase III proofreading domain, providing a common hydrolytic cleavage mechanism for RNA degradation and DNA editing, respectively.

Adenosine Monophosphate↗
PubMed2003

Activation-induced cytidine deaminase: a dual role in class-switch recombination and somatic hypermutation.

Maturation of the antibody repertoire is mediated by two different mechanisms: class-switch recombination (CSR) and somatic hypermutation (SHM). These two processes are T cell dependent and occur in the germinal centers of secondary lymphoid organs. CSR leads to the production of antibodies of different isotypes whereas SHM leads to the selection of B cells expressing a BCR with high affinity for antigen. The activation-induced cytidine deaminase (AID) was recently shown to play a key role in these two mechanisms, demonstrating for the first time that these maturation processes share a common mechanism. There is evidence that AID is involved in the somatic DNA alterations required for CSR and SHM. The mechanism of action of AID is unclear. As AID and APOBEC-1 display a sequence similarity, AID may act as an RNA-editing enzyme. However, the immunological abnormalities observed in uracil-N glycosylase deficiency in a recent study indirectly suggest that AID may edit DNA directly.

Animals↗
PubMed1989

Isolation and characterization of mutants with deletions in dnaQ, the gene for the editing subunit of DNA polymerase III in Salmonella typhimurium.

dnaQ (mutD) encodes the editing exonuclease subunit (epsilon) of DNA polymerase III. Previously described mutations in dnaQ include dominant and recessive mutator alleles as well as leaky temperature-sensitive alleles. We describe the properties of strains bearing null mutations (deletion-substitution alleles) of this gene. Null mutants exhibited a growth defect as well as elevated spontaneous mutation. As a consequence of the poor growth of dnaQ mutants and their high mutation rate, these strains were replaced within single colonies by derivatives carrying an extragenic suppressor mutation that compensated the growth defect but apparently not the mutator effect. Sixteen independently derived suppressors mapped in the vicinity of dnaE, the gene for the polymerization subunit (alpha) of DNA polymerase III, and one suppressor that was sequenced encoded an altered alpha polypeptide. Partially purified DNA polymerase III containing this altered alpha subunit was active in polymerization assays. In addition to their dependence on a suppressor mutation affecting alpha, dnaQ mutants strictly required DNA polymerase I for viability. We argue from these data that in the absence of epsilon, DNA replication falters unless secondary mechanisms, including genetically coded alteration in the intrinsic replication capacity of alpha and increased use of DNA polymerase I, come into play. Thus, epsilon plays a role in DNA replication distinct from its known role in controlling spontaneous mutation frequency.

Cloning, Molecular↗
PubMed2006

Hyper-IgM syndromes.

PURPOSE OF REVIEW: The recent elucidation of the molecular defects leading to hyper-IgM syndromes has provided considerable insight into the complex mechanisms that govern the antibody maturation in humans. RECENT FINDINGS: The study of a large cohort of patients revealed unexpected clinical, immunological and genetic findings, which have significant implications on the molecular basis of immunoglobulin class switch recombination and somatic hypermutation, as shown for hypomorphic mutations in the nuclear factor-kappaB essential modulator (NEMO) gene and peculiar activation-induced cytidine deaminase defects that differently affect class switch recombination and somatic hypermutation. The description of the hyper-IgM condition due to mutations in the gene encoding uracil-N glycosylase has been essential for defining the DNA-editing activity of activation-induced cytidine deaminase. Novel findings are awaited from the study of the yet genetically undefined hyper-IgM syndromes, leading to the identification of activation-induced cytidine deaminase cofactors and proteins involved in class switch recombination-induced DNA repair. In the genetically characterized hyper-IgM syndromes, the precise identification of the molecular defect allows the evaluation of hyper-IgM complications, and thus aids assessment of prognosis and proper survey and treatment. SUMMARY: The important contribution made by investigation of this condition improves our understanding of the physiology of the antibody response in humans.

Chromosomes, Human, X↗
PubMed1992

DNA replication defect in Salmonella typhimurium mutants lacking the editing (epsilon) subunit of DNA polymerase III.

In Salmonella typhimurium, dnaQ null mutants (encoding the epsilon editing subunit of DNA polymerase III [Pol III]) exhibit a severe growth defect when the genetic background is otherwise wild type. Suppression of the growth defect requires both a mutation affecting the alpha (polymerase) subunit of DNA polymerase III and adequate levels of DNA polymerase I. In the present paper, we report on studies that clarify the nature of the physiological defect imposed by the loss of epsilon and the mechanism of its suppression. Unsuppressed dnaQ mutants exhibited chronic SOS induction, indicating exposure of single-stranded DNA in vivo, most likely as gaps in double-stranded DNA. Suppression of the growth defect was associated with suppression of SOS induction. Thus, Pol I and the mutant Pol III combined to reduce the formation of single-stranded DNA or accelerate its maturation to double-stranded DNA. Studies with mutants in major DNA repair pathways supported the view that the defect in DNA metabolism in dnaQ mutants was at the level of DNA replication rather than of repair. The requirement for Pol I was satisfied by alleles of the gene for Pol I encoding polymerase activity or by rat DNA polymerase beta (which exhibits polymerase activity only). Consequently, normal growth is restored to dnaQ mutants when sufficient polymerase activity is provided and this compensatory polymerase activity can function independently of Pol III. The high level of Pol I polymerase activity may be required to satisfy the increased demand for residual DNA synthesis at regions of single-stranded DNA generated by epsilon-minus pol III. The emphasis on adequate polymerase activity in dnaQ mutants is also observed in the purified alpha subunit containing the suppressor mutation, which exhibits a modestly elevated intrinsic polymerase activity relative to that of wild-type alpha.

Base Sequence↗