Research output: Contribution to journal › Article › peer-review
Double-Stranded DNA Fragments Bearing Unrepairable Lesions and Their Internalization into Mouse Krebs-2 Carcinoma Cells. / Dolgova, Evgeniya V.; Evdokimov, Alexey N.; Proskurina, Anastasia S. et al.
In: Nucleic Acid Therapeutics, Vol. 29, No. 5, 01.10.2019, p. 278-290.Research output: Contribution to journal › Article › peer-review
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TY - JOUR
T1 - Double-Stranded DNA Fragments Bearing Unrepairable Lesions and Their Internalization into Mouse Krebs-2 Carcinoma Cells
AU - Dolgova, Evgeniya V.
AU - Evdokimov, Alexey N.
AU - Proskurina, Anastasia S.
AU - Efremov, Yaroslav R.
AU - Bayborodin, Sergey I.
AU - Potter, Ekaterina A.
AU - Popov, Alexey A.
AU - Petruseva, Irina O.
AU - Lavrik, Olga I.
AU - Bogachev, Sergey S.
PY - 2019/10/1
Y1 - 2019/10/1
N2 - Murine Krebs-2 tumor-initiating stem cells are known to natively internalize extracellular double-stranded DNA fragments. Being internalized, these fragments interfere in the repair of chemically induced interstrand cross-links. In the current investigation, 756 bp polymerase chain reaction (PCR) product containing bulky photoreactive dC adduct was used as extracellular DNA. This adduct was shown to inhibit the cellular system of nucleotide excision repair while being resistant to excision by this DNA repair system. The basic parameters for this DNA probe internalization by the murine Krebs-2 tumor cells were characterized. Being incubated under regular conditions (60 min, 24°C, 500 μL of the incubation medium, in the dark), 0.35% ± 0.18% of the Krebs-2 ascites cells were shown to natively internalize modified DNA. The saturating amount of the modified DNA was detected to be 0.37 μg per 106 cells. For the similar unmodified DNA fragments, this ratio is 0.73 μg per 106 cells. Krebs-2 tumor cells were shown to be saturated internalizing either (190 ± 40) × 103 molecules of modified DNA or (1,000 ± 100) × 103 molecules of native DNA. On internalization, the fragments of DNA undergo partial and nonuniform hydrolysis of 3′ ends followed by circularization. The degree of hydrolysis, assessed by sequencing of several clones with the insertion of specific PCR product, was 30-60 nucleotides.
AB - Murine Krebs-2 tumor-initiating stem cells are known to natively internalize extracellular double-stranded DNA fragments. Being internalized, these fragments interfere in the repair of chemically induced interstrand cross-links. In the current investigation, 756 bp polymerase chain reaction (PCR) product containing bulky photoreactive dC adduct was used as extracellular DNA. This adduct was shown to inhibit the cellular system of nucleotide excision repair while being resistant to excision by this DNA repair system. The basic parameters for this DNA probe internalization by the murine Krebs-2 tumor cells were characterized. Being incubated under regular conditions (60 min, 24°C, 500 μL of the incubation medium, in the dark), 0.35% ± 0.18% of the Krebs-2 ascites cells were shown to natively internalize modified DNA. The saturating amount of the modified DNA was detected to be 0.37 μg per 106 cells. For the similar unmodified DNA fragments, this ratio is 0.73 μg per 106 cells. Krebs-2 tumor cells were shown to be saturated internalizing either (190 ± 40) × 103 molecules of modified DNA or (1,000 ± 100) × 103 molecules of native DNA. On internalization, the fragments of DNA undergo partial and nonuniform hydrolysis of 3′ ends followed by circularization. The degree of hydrolysis, assessed by sequencing of several clones with the insertion of specific PCR product, was 30-60 nucleotides.
KW - DNA internalization
KW - Fap-dC photo-adduct
KW - Krebs-2 cells
KW - TAMRA fluorochrome
KW - unrepairable DNA substrate
KW - HORIZONTAL TRANSFER
KW - STEM-CELLS
KW - PHOTOACTIVATED DNA
KW - HIGH-AFFINITY
KW - MAMMALIAN-CELLS
KW - NUCLEOTIDE EXCISION-REPAIR
KW - OLIGONUCLEOTIDE UPTAKE
KW - DAMAGE RECOGNITION
KW - PROTEINS
KW - INTRACELLULAR TRAFFICKING
UR - http://www.scopus.com/inward/record.url?scp=85072705516&partnerID=8YFLogxK
U2 - 10.1089/nat.2019.0786
DO - 10.1089/nat.2019.0786
M3 - Article
C2 - 31194620
AN - SCOPUS:85072705516
VL - 29
SP - 278
EP - 290
JO - Nucleic Acid Therapeutics
JF - Nucleic Acid Therapeutics
SN - 2159-3337
IS - 5
ER -
ID: 21741464