Multiple dispersed spontaneous mutations: a novel pathway of mutation in a malignant human cell line

We analyzed the nature of spontaneous mutations at the autosomal locus coding for adenine phosphoribosyltransferase in the human colorectal carcinoma cell line SW620 to establish whether distinctive mutational pathways exist that might underlie the more complex genome rearrangements arising in tumor cells. Point mutations occur at a low rate in aprt hemizygotes derived from SW620, largely as a result of base substitutions at G.C base pairs to yield transversions and transitions. However, a novel pathway is evident in the form of multiple dispersed mutations in which two errors, separated by as much as 1,800 bp, fall in the same mutant gene. Such mutations could be the result of error-prone DNA synthesis occurring during normal replication or during long-patch excision-repair of spontaneously arising DNA lesions. This process could also contribute to the chromosomal instability evident in these tumor cells.

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Alleviation of C⋅C Mismatches in DNA by the Escherichia coli Fpg Protein

Frontiers in Microbiology ◽

10.3389/fmicb.2021.608839 ◽

2021 ◽

Vol 12 ◽

Author(s):

Almaz Nigatu Tesfahun ◽

Marina Alexeeva ◽

Miglė Tomkuvienė ◽

Aysha Arshad ◽

Prashanna Guragain ◽

...

Keyword(s):

Escherichia Coli ◽

Excision Repair ◽

Dna Lesions ◽

Base Pairs ◽

E Coli ◽

Thymine Glycol ◽

Base Excision ◽

The Cost ◽

Primary Substrate

DNA polymerase III mis-insertion may, where not corrected by its 3′→ 5′ exonuclease or the mismatch repair (MMR) function, result in all possible non-cognate base pairs in DNA generating base substitutions. The most thermodynamically unstable base pair, the cytosine (C)⋅C mismatch, destabilizes adjacent base pairs, is resistant to correction by MMR in Escherichia coli, and its repair mechanism remains elusive. We present here in vitro evidence that C⋅C mismatch can be processed by base excision repair initiated by the E. coli formamidopyrimidine-DNA glycosylase (Fpg) protein. The kcat for C⋅C is, however, 2.5 to 10 times lower than for its primary substrate 8-oxoguanine (oxo8G)⋅C, but approaches those for 5,6-dihydrothymine (dHT)⋅C and thymine glycol (Tg)⋅C. The KM values are all in the same range, which indicates efficient recognition of C⋅C mismatches in DNA. Fpg activity was also exhibited for the thymine (T)⋅T mismatch and for N4- and/or 5-methylated C opposite C or T, Fpg activity being enabled on a broad spectrum of DNA lesions and mismatches by the flexibility of the active site loop. We hypothesize that Fpg plays a role in resolving C⋅C in particular, but also other pyrimidine⋅pyrimidine mismatches, which increases survival at the cost of some mutagenesis.

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Analysis of spontaneous mutations in a chromosomally located hsv-1 thymidine kinase (TK) gene in a human cell line

Somatic Cell and Molecular Genetics ◽

10.1007/bf02422298 ◽

1987 ◽

Vol 13 (1) ◽

pp. 47-56 ◽

Cited By ~ 6

Author(s):

Daphne R. Goring ◽

Kalpana Gupta ◽

Michael S. DuBow

Keyword(s):

Cell Line ◽

Thymidine Kinase ◽

Human Cell ◽

Human Cell Line ◽

Spontaneous Mutations ◽

Hsv 1

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A protein produced by a monocytic human cell line can induce apoptosis on tumor cells

FEBS Letters ◽

10.1016/0014-5793(94)00287-8 ◽

1994 ◽

Vol 344 (1) ◽

pp. 35-40 ◽

Cited By ~ 5

Author(s):

L. Ghibelli ◽

S. Coppola ◽

C. Nosseri ◽

A. Bergamini ◽

S. Beninati

Keyword(s):

Cell Line ◽

Tumor Cells ◽

Human Cell ◽

Human Cell Line

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Single and Coincident Intragenic Mutations Attributable to Gene Conversion in a Human Cell Line

Genetics ◽

10.1093/genetics/146.4.1429 ◽

1997 ◽

Vol 146 (4) ◽

pp. 1429-1439

Author(s):

Cynthia R Giver ◽

Andrew J Grosovsky

Keyword(s):

Cell Line ◽

Gene Conversion ◽

Point Mutations ◽

Double Exchange ◽

Detailed Examination ◽

Human Cell Line ◽

Human Lymphoblastoid Cell ◽

Dna Base ◽

Dna Strand ◽

Conversion Tract

Two polymorphic sites are located within the heterozygous TK1 locus in the human lymphoblastoid cell line TK6: an inactivating frameshift in exon 4 of the nonfunctional allele and a phenotypically silent frameshift in exon 7 of the functional allele. Through the use of these intragenic polymorphisms and microsatellite markers that flank TK1, we demonstrate that partial gene conversion accounts for 3/75 (0.04) spontaneous and 9/163 (0.06) X-ray-induced TK1– mutants, thus comprising a significant component of forward mutations at this locus. In all cases, the conversion tract is <1 cM, rendering double exchange a remote alternate explanation for these results. Sequence analysis of full length TK1 cDNA provides rigorous exclusion of deletion events as a mechanism for generation of these allelotypes. Detailed examination of allelotypes in TK1– mutants identified two mechanisms for the generation of coincident sequence alterations that sometimes accompanied gene conversions. Mutations within the conversion tract were attributed to either error-prone gap filling synthesis during recombinational repair or mismatch repair within a heteroduplex region following branch migration. These findings suggest that a proportion of point mutations may not be targeted to sites of DNA base damage, but rather may arise as secondary consequences from the repair of DNA strand breaks.

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Circulating human B lymphocytes are deficient in nucleotide excision repair and accumulate mutations upon proliferation

Blood ◽

10.1182/blood-2010-12-326637 ◽

2011 ◽

Vol 117 (23) ◽

pp. 6277-6286 ◽

Cited By ~ 14

Author(s):

Nevila Hyka-Nouspikel ◽

Kimon Lemonidis ◽

Wei-Ting Lu ◽

Thierry Nouspikel

Keyword(s):

Cell Cycle ◽

B Lymphocytes ◽

Nucleotide Excision Repair ◽

Excision Repair ◽

Point Mutations ◽

Dna Lesions ◽

Repair System ◽

Differentiated Cells ◽

Nucleotide Excision ◽

Dividing Cells

Abstract Faithful repair of DNA lesions is a crucial task that dividing cells must actively perform to maintain genome integrity. Strikingly, nucleotide excision repair (NER), the most versatile DNA repair system, is specifically down-regulated in terminally differentiated cells. This prompted us to examine whether NER attenuation might be a common feature of all G0-arrested cells, and in particular of those that retain the capacity to reenter cell cycle and might thus convert unrepaired DNA lesions into mutations, a prerequisite for malignant transformation. Here we report that quiescent primary human B lymphocytes down-regulate NER at the global genome level while maintaining proficient repair of constitutively expressed genes. Quiescent B cells exposed to an environment that causes both DNA damage and proliferation accumulate point mutations in silent and inducible genes crucial for cell replication and differentiation, such as BCL6 and Cyclin D2. Similar to differentiated cells, NER attenuation in quiescent cells is associated with incomplete phosphorylation of the ubiquitin activating enzyme Ube1, which is required for proficient NER. Our data establish a mechanistic link between NER attenuation during quiescence and cell mutagenesis and also support the concept that oncogenic events targeting cell cycle- or activation-induced genes might initiate genomic instability and lymphomagenesis.

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Combined mismatch and nucleotide excision repair defects in a human cell line: mismatch repair processes methylation but not UV- or ionizing radiation-induced DNA damage

Carcinogenesis ◽

10.1093/carcin/20.5.799 ◽

1999 ◽

Vol 20 (5) ◽

pp. 799-804 ◽

Cited By ~ 12

Author(s):

M. O'Driscoll ◽

S. Martinelli ◽

C. Ciotta ◽

P. Karran

Keyword(s):

Dna Damage ◽

Ionizing Radiation ◽

Cell Line ◽

Mismatch Repair ◽

Nucleotide Excision Repair ◽

Human Cell ◽

Excision Repair ◽

Human Cell Line ◽

Nucleotide Excision ◽

Radiation Induced

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The human myeloma cell line LP-1: a versatile model in which to study early plasma-cell differentiation and c-myc activation

Blood ◽

10.1182/blood.v73.4.1020.bloodjournal7341020 ◽

1989 ◽

Vol 73 (4) ◽

pp. 1020-1027

Author(s):

L Pegoraro ◽

F Malavasi ◽

G Bellone ◽

M Massaia ◽

M Boccadoro ◽

...

Keyword(s):

Cell Line ◽

Plasma Cells ◽

Point Mutations ◽

Myeloma Cell ◽

Human Cell Line ◽

B Cell Maturation ◽

Potential Partner ◽

Human Myeloma Cell Line ◽

Myc Gene ◽

Conventional Cytogenetics

The characteristics of a human cell line (LP-1) derived from the peripheral blood of a patient with IgG-lambda myeloma in leukemic transformation are described. The cells resemble immature plasma cells in that they exhibit a membrane phenotype that is intermediate between late B lymphocytes and plasma cells, even though they secrete IgG- lambda chains. Treatment of LP-1 cells with 12–0 tetradecanoylphorbol- 13-acetate (TPA) or pokeweek mitogen (PWM) induces the appearance of surface markers and ultrastructural features typical of mature plasma cells but does not affect their proliferative activity. Molecular analysis of the cell line showed an increased expression of the c-myc protooncogene and the presence of abnormally sized transcripts. Conventional cytogenetics and pulsed-field gel electrophoresis showed no structural rearrangements of the c-myc gene, suggesting that the abnormal c-myc expression may be due to point mutations or small deletions within the gene. The LP-1 cell line is a useful model in which to study the process of B-cell maturation; such study may lead to the uncovering of unusual mechanisms of c-myc activation. Furthermore, the LP-1 cell is a potential partner in the generation of human hybridomas.

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The Influence of 5′,8-Cyclo-2′-deoxypurines on the Mitochondrial Repair of Clustered DNA Damage in Xrs5 Cells: The Preliminary Study

Molecules ◽

10.3390/molecules26227042 ◽

2021 ◽

Vol 26 (22) ◽

pp. 7042

Author(s):

Karolina Boguszewska ◽

Julia Kaźmierczak-Barańska ◽

Bolesław T. Karwowski

Keyword(s):

Excision Repair ◽

Dna Lesions ◽

Base Pairs ◽

Apurinic Site ◽

Ap Sites ◽

Base Excision ◽

First Time ◽

Ap Site ◽

Clustered Dna Damage ◽

Single Lesion

The 5′,8-cyclo-2′-deoxypurines (cdPus) affect the DNA structure. When these bulky structures are a part of clustered DNA lesions (CDL), they affect the repair of the other lesions within the cluster. Mitochondria are crucial for cell survival and have their own genome, hence, are highly interesting in the context of CDL repair. However, no studies are exploring this topic. Here, the initial stages of mitochondrial base excision repair (mtBER) were considered—the strand incision and elongation. The repair of a single lesion (apurinic site (AP site)) accompanying the cdPu within the double-stranded CDL has been investigated for the first time. The type of cdPu, its diastereomeric form, and the interlesion distance were taken into consideration. For these studies, the established experimental model of short oligonucleotides (containing AP sites located ≤7 base pairs to the cdPu in both directions) and mitochondrial extracts of the xrs5 cells were used. The obtained results have shown that the presence of cdPus influenced the processing of an AP site within the CDL. Levels of strand incision and elongation were higher for oligos containing RcdA and ScdG than for those with ScdA and RcdG. Investigated stages of mtBER were more efficient for DNA containing AP sites located on 5′-end side of cdPu than on its 3′-end side. In conclusion, the presence of cdPus in mtDNA structure may affect mtBER (processing the second mutagenic lesion within the CDL). As impaired repair processes may lead to serious biological consequences, further studies concerning the mitochondrial repair of CDL are highly demanded.

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Structural basis of the XPB helicase–Bax1 nuclease complex interacting with the repair bubble DNA

Nucleic Acids Research ◽

10.1093/nar/gkaa801 ◽

2020 ◽

Vol 48 (20) ◽

pp. 11695-11705

Author(s):

Feng He ◽

Kevin DuPrez ◽

Eduardo Hilario ◽

Zhenhang Chen ◽

Li Fan

Keyword(s):

Excision Repair ◽

Uv Light ◽

Dna Helicase ◽

Dna Lesions ◽

Structural Basis ◽

Base Pairs ◽

Dna Strands ◽

Underlying Mechanisms ◽

Dna Strand ◽

Atp Binding And Hydrolysis

Abstract Nucleotide excision repair (NER) removes various DNA lesions caused by UV light and chemical carcinogens. The DNA helicase XPB plays a key role in DNA opening and coordinating damage incision by nucleases during NER, but the underlying mechanisms remain unclear. Here, we report crystal structures of XPB from Sulfurisphaera tokodaii (St) bound to the nuclease Bax1 and their complex with a bubble DNA having one arm unwound in the crystal. StXPB and Bax1 together spirally encircle 10 base pairs of duplex DNA at the double-/single-stranded (ds–ss) junction. Furthermore, StXPB has its ThM motif intruding between the two DNA strands and gripping the 3′-overhang while Bax1 interacts with the 5′-overhang. This ternary complex likely reflects the state of repair bubble extension by the XPB and nuclease machine. ATP binding and hydrolysis by StXPB could lead to a spiral translocation along dsDNA and DNA strand separation by the ThM motif, revealing an unconventional DNA unwinding mechanism. Interestingly, the DNA is kept away from the nuclease domain of Bax1, potentially preventing DNA incision by Bax1 during repair bubble extension.

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