scholarly journals DNA repair synthesis in human heterokaryons. III. The rapid and slow complementing varieties of xeroderma pigmentosum

1976 ◽  
Vol 20 (1) ◽  
pp. 207-213
Author(s):  
F. Giannelli ◽  
S.A. Pawsey
Keyword(s):  
Protein Synthesis ◽  
Xeroderma Pigmentosum ◽  
Excision Repair ◽  
Dependent Manner ◽  
Repair Synthesis ◽  
Normal Cells ◽  
Complementation Groups ◽  
Dna Repair Synthesis ◽  
And Control ◽  
The Relationship

Patients with Xeroderma pigmentosum and defective DNA excision repair can be distinguished as a rapid (r-XP) and slow (s-XP) complementing variety. When fused with normal cells, fibroblasts from the r-XP are complemented rapidly and in the absence of protein synthesis while those from the s-XP are complemented slowly by a process partly, but not entirely, dependent on protein synthesis. Heterokaryons with different ratios of r-XP to s-XP nuclei (i.e. 1:1-5 and 1-5:1) and control heterokaryons containing one normal and 1-5 r- or s-XP nuclei show that if cell fusion and incubation is conducted in medium preventing protein synthesis, the rXP cells do not complement the s-XP partner at all and, conversely, that the latter is not as effective as normal cells at complementing the rXP partner. On the contrary, if protein synthesis is permitted, the 2 types of XP cells complement each other in a gene dose-dependent manner and to an extent similar to that observed in the control heterokaryons. These findings indicate that the r- and s-XP varieties are caused by mutations at different loci and suggest that the products of these loci interact to produce a functional unit which is present in normal control cells but absent in the XP strains. The relationship between the complementation groups described here and those already reported in the literature being investigated.

Xeroderma pigmentosum D-HeLa hybrids with low and high ultraviolet sensitivity associated with normal and diminished DNA repair ability, respectively

1985 ◽  
Vol 76 (1) ◽  
pp. 115-133
Author(s):  
R.T. Johnson ◽  
S. Squires ◽  
G.C. Elliott ◽  
G.L. Koch ◽  
A.J. Rainbow
Keyword(s):  
Dna Repair ◽  
Xeroderma Pigmentosum ◽  
Excision Repair ◽  
Complementation Group ◽  
Repair Capacity ◽  
Repair Synthesis ◽  
Repair Ability ◽  
Dna Repair Synthesis ◽  
Host Cell Reactivation ◽  
Group D

Fusion between HeLa and fibroblasts from complementation group D xeroderma pigmentosum (XPD) followed by challenge with small doses of ultraviolet light (u.v.) results in the production of hybrid cells expressing either HeLa (HD1) or XPD-like (HD2) sensitivity to u.v. and related repair capacity. Assays used included unscheduled DNA synthesis (UDS), DNA break accumulation in the presence of inhibitors of DNA repair synthesis and host cell reactivation of irradiated adenovirus. Complementation assay in heterokaryons reveals limited ability of HD2 to restore UDS in XPD nuclei. We believe this complementation is more apparent than real since proliferating hybrids of HD2 and XPD parentage are without exception u.v.-sensitive and express limited excision repair. On the other hand hybrids between HD2 and XPC, XPE or XPF fibroblasts show true complementation resulting in a return to normal u.v. sensitivity and elevated repair ability.

Xeroderma pigmentosum complementation group C cells remove pyrimidine dimers selectively from the transcribed strand of active genes

1991 ◽  
Vol 11 (8) ◽  
pp. 4128-4134
Author(s):  
J Venema ◽  
A van Hoffen ◽  
V Karcagi ◽  
A T Natarajan ◽  
A A van Zeeland ◽  
...  
Keyword(s):  
Xeroderma Pigmentosum ◽  
Mammalian Cells ◽  
Excision Repair ◽  
Complementation Group ◽  
C Cells ◽  
Dhfr Gene ◽  
Normal Cells ◽  
Pyrimidine Dimers ◽  
Normal Human ◽  
Active Genes

We have measured the removal of UV-induced pyrimidine dimers from DNA fragments of the adenosine deaminase (ADA) and dihydrofolate reductase (DHFR) genes in primary normal human and xeroderma pigmentosum complementation group C (XP-C) cells. Using strand-specific probes, we show that in normal cells, preferential repair of the 5' part of the ADA gene is due to the rapid and efficient repair of the transcribed strand. Within 8 h after irradiation with UV at 10 J m-2, 70% of the pyrimidine dimers in this strand are removed. The nontranscribed strand is repaired at a much slower rate, with 30% dimers removed after 8 h. Repair of the transcribed strand in XP-C cells occurs at a rate indistinguishable from that in normal cells, but the nontranscribed strand is not repaired significantly in these cells. Similar results were obtained for the DHFR gene. In the 3' part of the ADA gene, however, both normal and XP-C cells perform fast and efficient repair of either strand, which is likely to be caused by the presence of transcription units on both strands. The factor defective in XP-C cells is apparently involved in the processing of DNA damage in inactive parts of the genome, including nontranscribed strands of active genes. These findings have important implications for the understanding of the mechanism of UV-induced excision repair and mutagenesis in mammalian cells.

scholarly journals Ongoing replication forks delay nuclear envelope breakdown upon mitotic entry

2020 ◽  
pp. jbc.RA120.015142
Author(s):  
Yoshitami Hashimoto ◽  
Hirofumi Tanaka
Keyword(s):  
Dna Replication ◽  
Nuclear Envelope ◽  
Dependent Manner ◽  
Repair Synthesis ◽  
Nuclear Envelope Breakdown ◽  
M Phase ◽  
Egg Extracts ◽  
Fork Stalling ◽  
Dna Repair Synthesis ◽  
Xenopus Egg Extracts

DNA replication is a major contributor to genomic instability and protection against DNA replication perturbation is essential for normal cell division. Certain types of replication stress agents, such as aphidicolin and hydroxyurea, have been shown to cause reversible replication fork stalling, wherein replisome complexes are stably maintained with competence to restart in the S-phase of the cell cycle. If these stalled forks persist into the M-phase without a replication restart, replisomes are disassembled in a p97-dependent pathway and under-replicated DNA is subjected to mitotic DNA repair synthesis. Here, using Xenopus egg extracts, we investigated the consequences that arise when stalled forks are released simultaneously with the induction of mitosis. Ara-cytidine-5’-triphosphate (Ara-CTP)-induced stalled forks were able to restart with the addition of excess dCTPduring early mitosis before the nuclear envelope breakdown (NEB). However, stalled forks could no longer restart efficiently after NEB. Although replisome complexes were finally disassembled in a p97-dependent manner during mitotic progression whether or not fork stalling was relieved, the timing of NEB was delayed with the ongoing forks, rather than the stalled forks, and the delay was dependent on Wee1/Myt1 kinase activities. Thus, ongoing DNA replication was found to be directly linked to the regulation of Wee1/Myt1 kinases to modulate cyclin-dependent kinase (CDK) activities, owing to which DNA replication and mitosis occur in a mutually exclusive and sequential manner.

scholarly journals Generation of single-nucleotide repair patches following excision of uracil residues from DNA.

1992 ◽  
Vol 12 (4) ◽  
pp. 1605-1612 ◽  
Author(s):  
G Dianov ◽  
A Price ◽  
T Lindahl
Keyword(s):  
Mammalian Cell ◽  
Excision Repair ◽  
Enzymatic Digestion ◽  
Enzyme Analysis ◽  
Restriction Enzyme Analysis ◽  
Repair Synthesis ◽  
Single Nucleotide ◽  
Cell Extracts ◽  
Dna Repair Synthesis

The extent and location of DNA repair synthesis in a double-stranded oligonucleotide containing a single dUMP residue have been determined. Gently prepared Escherichia coli and mammalian cell extracts were employed for excision repair in vitro. The size of the resynthesized patch was estimated by restriction enzyme analysis of the repaired oligonucleotide. Following enzymatic digestion and denaturing gel electrophoresis, the extent of incorporation of radioactively labeled nucleotides in the vicinity of the lesion was determined by autoradiography. Cell extracts of E. coli and of human cell lines were shown to carry out repair mainly by replacing a single nucleotide. No significant repair replication on the 5' side of the lesion was observed. The data indicate that, after cleavage of the dUMP residue by uracil-DNA glycosylase and incision of the resultant apurinic-apyrimidinic site by an apurinic-apyrimidinic endonuclease activity, the excision step is catalyzed usually by a DNA deoxyribophosphodiesterase rather than by an exonuclease. Gap-filling and ligation complete the repair reaction. Experiments with enzyme inhibitors in mammalian cell extracts suggest that the repair replication step is catalyzed by DNA polymerase beta.

scholarly journals Xeroderma pigmentosum complementation group C cells remove pyrimidine dimers selectively from the transcribed strand of active genes.

1991 ◽  
Vol 11 (8) ◽  
pp. 4128-4134 ◽  
Author(s):  
J Venema ◽  
A van Hoffen ◽  
V Karcagi ◽  
A T Natarajan ◽  
A A van Zeeland ◽  
...  
Keyword(s):  
Xeroderma Pigmentosum ◽  
Mammalian Cells ◽  
Excision Repair ◽  
Complementation Group ◽  
C Cells ◽  
Dhfr Gene ◽  
Normal Cells ◽  
Pyrimidine Dimers ◽  
Normal Human ◽  
Active Genes

We have measured the removal of UV-induced pyrimidine dimers from DNA fragments of the adenosine deaminase (ADA) and dihydrofolate reductase (DHFR) genes in primary normal human and xeroderma pigmentosum complementation group C (XP-C) cells. Using strand-specific probes, we show that in normal cells, preferential repair of the 5' part of the ADA gene is due to the rapid and efficient repair of the transcribed strand. Within 8 h after irradiation with UV at 10 J m-2, 70% of the pyrimidine dimers in this strand are removed. The nontranscribed strand is repaired at a much slower rate, with 30% dimers removed after 8 h. Repair of the transcribed strand in XP-C cells occurs at a rate indistinguishable from that in normal cells, but the nontranscribed strand is not repaired significantly in these cells. Similar results were obtained for the DHFR gene. In the 3' part of the ADA gene, however, both normal and XP-C cells perform fast and efficient repair of either strand, which is likely to be caused by the presence of transcription units on both strands. The factor defective in XP-C cells is apparently involved in the processing of DNA damage in inactive parts of the genome, including nontranscribed strands of active genes. These findings have important implications for the understanding of the mechanism of UV-induced excision repair and mutagenesis in mammalian cells.

scholarly journals The Base Excision Repair Pathway in the Nematode Caenorhabditis elegans

2020 ◽  
Vol 8 ◽  
Author(s):  
Noha Elsakrmy ◽  
Qiu-Mei Zhang-Akiyama ◽  
Dindial Ramotar
Keyword(s):  
Dna Repair ◽  
Caenorhabditis Elegans ◽  
Dna Polymerase ◽  
Base Excision Repair ◽  
Excision Repair ◽  
Dna Glycosylases ◽  
Repair Synthesis ◽  
C Elegans ◽  
Base Excision ◽  
Dna Repair Synthesis

Exogenous and endogenous damage to the DNA is inevitable. Several DNA repair pathways including base excision, nucleotide excision, mismatch, homologous and non-homologous recombinations are conserved across all organisms to faithfully maintain the integrity of the genome. The base excision repair (BER) pathway functions to repair single-base DNA lesions and during the process creates the premutagenic apurinic/apyrimidinic (AP) sites. In this review, we discuss the components of the BER pathway in the nematode Caenorhabditis elegans and delineate the different phenotypes caused by the deletion or the knockdown of the respective DNA repair gene, as well as the implications. To date, two DNA glycosylases have been identified in C. elegans, the monofunctional uracil DNA glycosylase-1 (UNG-1) and the bifunctional endonuclease III-1 (NTH-1) with associated AP lyase activity. In addition, the animal possesses two AP endonucleases belonging to the exonuclease-3 and endonuclease IV families and in C. elegans these enzymes are called EXO-3 and APN-1, respectively. In mammalian cells, the DNA polymerase, Pol beta, that is required to reinsert the correct bases for DNA repair synthesis is not found in the genome of C. elegans and the evidence indicates that this role could be substituted by DNA polymerase theta (POLQ), which is known to perform a function in the microhomology-mediated end-joining pathway in human cells. The phenotypes observed by the C. elegans mutant strains of the BER pathway raised many challenging questions including the possibility that the DNA glycosylases may have broader functional roles, as discuss in this review.

Clustered Sites of DNA Repair Synthesis during Early Nucleotide Excision Repair in Ultraviolet Light-Irradiated Quiescent Human Fibroblasts

2002 ◽  
Vol 276 (2) ◽  
pp. 284-295 ◽  
Author(s):  
Maria Svetlova ◽  
Lioudmila Solovjeva ◽  
Nadezhda Pleskach ◽  
Natalia Yartseva ◽  
Tatyana Yakovleva ◽  
...  
Keyword(s):  
Dna Repair ◽  
Nucleotide Excision Repair ◽  
Ultraviolet Light ◽  
Excision Repair ◽  
Human Fibroblasts ◽  
Repair Synthesis ◽  
Nucleotide Excision ◽  
Dna Repair Synthesis

scholarly journals DNA repair synthesis during base excision repair in vitro is catalyzed by DNA polymerase epsilon and is influenced by DNA polymerases alpha and delta in Saccharomyces cerevisiae.

1993 ◽  
Vol 13 (2) ◽  
pp. 1051-1058 ◽  
Author(s):  
Z Wang ◽  
X Wu ◽  
E C Friedberg
Keyword(s):  
Dna Repair ◽  
Dna Polymerase ◽  
Base Excision Repair ◽  
Excision Repair ◽  
Repair Synthesis ◽  
Dna Polymerase Epsilon ◽  
Nuclear Extracts ◽  
Polymerase Epsilon ◽  
Base Excision ◽  
Dna Repair Synthesis

Base excision repair is an important mechanism for correcting DNA damage produced by many physical and chemical agents. We have examined the effects of the REV3 gene and the DNA polymerase genes POL1, POL2, and POL3 of Saccharomyces cerevisiae on DNA repair synthesis is nuclear extracts. Deletional inactivation of REV3 did not affect repair synthesis in the base excision repair pathway. Repair synthesis in nuclear extracts of pol1, pol2, and pol3 temperature-sensitive mutants was normal at permissive temperatures. However, repair synthesis in pol2 nuclear extracts was defective at the restrictive temperature of 37 degrees C and could be complemented by the addition of purified yeast DNA polymerase epsilon. Repair synthesis in pol1 nuclear extracts was proficient at the restrictive temperature unless DNA polymerase alpha was inactivated prior to the initiation of DNA repair. Thermal inactivation of DNA polymerase delta in pol3 nuclear extracts enhanced DNA repair synthesis approximately 2-fold, an effect which could be specifically reversed by the addition of purified yeast DNA polymerase delta to the extract. These results demonstrate that DNA repair synthesis in the yeast base excision repair pathway is catalyzed by DNA polymerase epsilon but is apparently modulated by the presence of DNA polymerases alpha and delta.

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