Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.7.7 (DNA polymerase)
 17,007 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

DNA damage was evaluated by flow cytometric (FCM) analysis of cells treated with L-phenylalanine mustard (L-PAM) and stained with anti-DNA monoclonal antibody (MAb) F7-26. DNA damage was rapidly repaired, as indicated by the loss of DNA immunoreactivity after removal of L-PAM. Two types of drug combinations were found to inhibit DNA repair. Combinations containing inhibitors of DNA polymerase (ara-C, aphidicolin) or these inhibitors and hydroxyurea inhibited DNA repair in A2780/PAM and A549 cells. The inhibition of DNA repair by combinations of DNA-damaging agents thioTEPA or cisplatin and DNA polymerase inhibitors is a novel observation based on the specificity of DNA damage assay with MAb F7-26. Combinations containing thioTEPA or cisplatin inhibited DNA repair in A549 but not in A2780/PAM cells. Drug combinations which inhibited DNA repair also significantly enhanced cell killing by L-PAM. Cell survival in cultures treated with L-PAM and efficient inhibitors was 2 to 3 orders of magnitude lower than was expected for additive survival. ThioTEPA and cisplatin play a dual role in combination chemotherapy by inducing DNA damage and inhibiting repair of DNA damage. FCM analysis of DNA repair may be a useful component of drug evaluation and could be applied to determine cell-type specific sensitivity to inhibitors of DNA repair.
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PMID:Inhibition of DNA repair and the enhancement of cytotoxicity of alkylating agents. 190 57

Harnessing CRISPR-Cas9 technology provides an unprecedented ability to modify genomic loci via DNA double-strand break (DSB) induction and repair. We analyzed nonhomologous end-joining (NHEJ) repair induced by Cas9 in budding yeast and found that the orientation of binding of Cas9 and its guide RNA (gRNA) profoundly influences the pattern of insertion/deletions (indels) at the site of cleavage. A common indel created by Cas9 is a 1-bp (+1) insertion that appears to result from Cas9 creating a 1-nt 5' overhang that is filled in by a DNA polymerase and ligated. The origin of +1 insertions was investigated by using two gRNAs with PAM sequences located on opposite DNA strands but designed to cleave the same sequence. These templated +1 insertions are dependent on the X-family DNA polymerase, Pol4. Deleting Pol4 also eliminated +2 and +3 insertions, which are biased toward homonucleotide insertions. Using inverted PAM sequences, we also found significant differences in overall NHEJ efficiency and repair profiles, suggesting that the binding of the Cas9:gRNA complex influences subsequent NHEJ processing. As with events induced by the site-specific HO endonuclease, CRISPR-Cas9-mediated NHEJ repair depends on the Ku heterodimer and DNA ligase 4. Cas9 events are highly dependent on the Mre11-Rad50-Xrs2 complex, independent of Mre11's nuclease activity. Inspection of the outcomes of a large number of Cas9 cleavage events in mammalian cells reveals a similar templated origin of +1 insertions in human cells, but also a significant frequency of similarly templated +2 insertions.
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PMID:CRISPR/Cas9 cleavages in budding yeast reveal templated insertions and strand-specific insertion/deletion profiles. 2978 87