کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
8320329 | 1539361 | 2018 | 13 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Persistent 3â²-phosphate termini and increased cytotoxicity of radiomimetic DNA double-strand breaks in cells lacking polynucleotide kinase/phosphatase despite presence of an alternative 3â²-phosphatase
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کلمات کلیدی
موضوعات مرتبط
علوم زیستی و بیوفناوری
بیوشیمی، ژنتیک و زیست شناسی مولکولی
زیست شیمی
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چکیده انگلیسی
Polynucleotide kinase/phosphatase (PNKP) has been implicated in non-homologous end joining (NHEJ) of DNA double-strand breaks (DSBs). To assess the consequences of PNKP deficiency for NHEJ of 3â²-phosphate-ended DSBs, PNKP-deficient derivatives of HCT116 and of HeLa cells were generated using CRISPR/CAS9. For both cell lines, PNKP deficiency conferred sensitivity to ionizing radiation as well as to neocarzinostatin (NCS), which specifically induces DSBs bearing protruding 3â²-phosphate termini. Moreover, NCS-induced DSBs, detected as 53BP1 foci, were more persistent in PNKP â/â HCT116 cells compared to their wild-type (WT) counterparts. Surprisingly, PNKP-deficient whole-cell and nuclear extracts were biochemically competent in removing both protruding and recessed 3â²-phosphates from synthetic DSB substrates, albeit much less efficiently than WT extracts, suggesting an alternative 3â²-phosphatase. Measurements by ligation-mediated PCR showed that PNKP-deficient HeLa cells contained significantly more 3â²-phosphate-terminated and fewer 3â²-hydroxyl-terminated DSBs than parental cells 5-15â¯min after NCS treatment, but this difference disappeared by 1â¯h. These results suggest that, despite presence of an alternative 3â²-phosphatase, loss of PNKP significantly sensitizes cells to 3â²-phosphate-terminated DSBs, due to a 3â²-dephosphorylation defect.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: DNA Repair - Volume 68, August 2018, Pages 12-24
Journal: DNA Repair - Volume 68, August 2018, Pages 12-24
نویسندگان
Sri Lakshmi Chalasani, Ajinkya S. Kawale, Konstantin Akopiants, Yaping Yu, Mesfin Fanta, Michael Weinfeld, Lawrence F. Povirk,