Base editing with a Cpf1–cytidine deaminase fusion
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE110136 | GEO | 2018/03/18
REPOSITORIES: GEO
Similar Datasets
Project description:C-to-T base editing mediated by CRISPR/Cas9 base editors (BEs) needs a G/C-rich PAM and the editing fidelity is compromised by unwanted indels and non-C-to-T substitutions. We developed CRISPR/Cpf1-based BEs to recognize a T-rich PAM and induce efficient C-to-T editing with few indels and/or non-C-to-T substitutions. The requirement of editing fidelity in therapeutic-related trials necessitates the development of CRISPR/Cpf1-based BEs, which also facilitates base editing in A/T-rich regions.
2018-10-16 | GSE105002 | GEO
Project description:Base editing with a Cpf1–cytidine deaminase fusion
| PRJNA432936 | ENA
Project description:High-fidelity base editing mediated by Cpf1-cytidine deaminase fusion
| PRJNA414402 | ENA
Project description:Base editors (BEs) shed new light on correcting disease-related T-to-C mutations. However, current rat APOBEC1-based BEs are less efficient in editing cytosines in highly-methylated regions or in GpC context. By screening a variety of APOBEC/AID deaminases, we showed that human APOBEC3A-conjugated BE and its engineered forms can mediate efficient C-to-T base editing in all examined contexts, including regions with high-methylation levels and GpC dinucleotides, which extends base editing scope.
2018-08-20 | GSE114999 | GEO
Project description:The APOBEC3 proteins form a multigene family of cytidine deaminases with inhibitory activity against viruses and retrotransposons. In contrast to APOBEC3G (A3G), APOBEC3A (A3A) has no effect on lentiviruses but dramatically inhibits replication of the parvovirus adeno-associated virus (AAV). To study the contribution of deaminase activity to the antiviral activity of A3A, we performed a comprehensive mutational analysis of A3A. By mutation of non-conserved residues, we found that regions outside of the catalytic active site contribute to both deaminase and antiviral activities. Using A3A point mutants and A3A/A3G chimeras, we show that deaminase activity is not required for inhibition of recombinant AAV production. We also found that deaminase-deficient A3A mutants block replication of both wild-type AAV and the autonomous parvovirus minute virus of mice (MVM). In addition, we identify specific residues of A3A that confer activity against AAV when substituted into A3G. In summary, our results demonstrate that deaminase activity is not necessary for the antiviral activity of A3A against parvoviruses.
| S-EPMC2678267 | biostudies-literature
Project description:Activation-induced cytidine deaminase (AID) initiates class switch recombination (CSR) and somatic hypermutation (SHM) by deaminating cytosine residues in immunoglobulin genes (Igh, Igκ, and Igλ). At a lower frequency, AID also causes collateral DNA damage at non-Ig loci, including genes that are rearranged or mutated in B-cell lymphoma. Precisely how AID is recruited to these off-target sites is not entirely understood. To gain further insight into how AID selects its targets, we compared AID-mediated translocations in two different cell types, B cells and mouse embryonic fibroblasts (MEFs). AID targets a distinct set of hotspots in the two cell types. In both cases, hotspots are concentrated in highly transcribed but stalled genes. However, transcription alone is insufficient to recruit AID activity. Comparison of genes similarly transcribed in B cells and MEFs but targeted in only one of the two cell types reveals a common set of epigenetic features associated with AID recruitment in both cells. AID target genes are enriched in chromatin modifications associated with active enhancers (such as H3K27Ac) and marks of active transcription (such as H3K36me3) in both fibroblasts and B cells, indicating that these features are universal mediators of AID recruitment.
| S-EPMC4284578 | biostudies-literature
Project description:Precisely how AID is recruited to these off-target sites is not entirely understood. To gain further insight into how AID selects its targets we compared AID-mediated translocations in two different cell types, B cells and mouse embryonic fibroblasts (MEFs). AID targets a distinct set of hotspots in the two cell types. In both cases, hotspots are concentrated in highly transcribed but stalled genes. However, transcription alone is insufficient to recruit AID activity. Comparison of genes similarly transcribed in B cells and MEFs but targeted in only one of the two cell types reveals a common set of epigenetic features associated with AID recruitment in both cell types. AID target genes are enriched in chromatin modifications associated with active enhancers (such as H3K27Ac) and marks of active transcription (such as H3K36me3) in both fibroblasts and B cells, indicating that these features are universal mediators of AID recruitmen Analysis of total mRNA and PolII loading in activated B lymphocytes and Mouse Embryonic Fibroblasts (MEFs)
2015-01-30 | E-GEOD-61523 | biostudies-arrayexpress
Project description:We previously reported that replacing HIV-1 nucleocapsid (NC) domain with SARS-CoV nucleocapsid (N) residues 2-213, 215-421, or 234-421 results in efficient virus-like particle (VLP) production at a level comparable to that of wild-type HIV-1. In this study we demonstrate that these chimeras are capable of packaging large amounts of human APOBEC3G (hA3G), and that an HIV-1 Gag chimera containing the carboxyl-terminal half of human coronavirus 229E (HCoV-229E) N as a substitute for NC is capable of directing VLP assembly and efficiently packaging hA3G. When co-expressed with SARS-CoV N and M (membrane) proteins, hA3G was efficiently incorporated into SARS-CoV VLPs. Data from GST pull-down assays suggest that the N sequence involved in N-hA3G interactions is located between residues 86 and 302. Like HIV-1 NC, the SARS-CoV or HCoV-229E N-associated with hA3G depends on the presence of RNA, with the first linker region essential for hA3G packaging into both HIV-1 and SARS-CoV VLPs. The results raise the possibility that hA3G is capable of associating with different species of viral structural proteins through a potentially common, RNA-mediated mechanism.
| S-EPMC7103413 | biostudies-literature
Project description:The human genome encodes seven APOBEC3 (A3) cytidine deaminases with potential antiretroviral activity: A3A, A3B, A3C, A3DE, A3F, A3G, and A3H. A3G was the first identified to block replication of human immunodeficiency virus type 1 (HIV-1) and many other retroviruses. A3F, A3B, and A3DE were shown later to have similar activities. HIV-1 produces a protein called Vif that is able to neutralize the antiretroviral activities of A3DE, A3F, and A3G, but not A3B. Only the antiretroviral activity of A3H remains to be defined due to its poor expression in cell culture. Here, we studied the mechanism impairing A3H expression. When primate A3H sequences were compared, a premature termination codon was identified on the fifth exon of the human and chimpanzee A3H genes, which significantly decreased their protein expression. It causes a 29-residue deletion from the C terminus, and this truncation did not reduce human A3H protein stability. However, the mRNA levels of the truncated gene were significantly decreased. Human A3H protein expression could be restored to a normal level either by repairing this truncation or through expression from a vector containing an intron from human cytomegalovirus. Once expression was optimized, human A3H could reduce HIV-1 infectivity up to 150-fold. Importantly, HIV-1 Vif failed to neutralize A3H activity. Nevertheless, extensive sequence analysis could not detect any significant levels of G-to-A mutation in the HIV-1 genome by human A3H. Thus, A3H inhibits HIV-1 replication potently by a cytidine deamination-independent mechanism, and optimizing A3H expression in vivo should represent a novel therapeutic strategy for HIV-1 treatment.
| S-EPMC2430661 | biostudies-literature