Carcinogen-induced DNA structural distortion differences in the RAS gene isoforms; the importance of local sequence

Abstract Background Local sequence context is known to have an impact on the mutational pattern seen in cancer. The RAS genes and a smoking carcinogen, Benzo[a]pyrene diol epoxide (BPDE), have been utilised to explore these context effects. BPDE is known to form an adduct at the guanines in a number of RAS gene sites, KRAS codons 12, 13 and 14, NRAS codon 12, and HRAS codons 12 and 14. Results Molecular modelling techniques, along with multivariate analysis, have been utilised to determine the sequence influenced differences between BPDE-adducted RAS gene sequences as well as the local distortion caused by the adducts. Conclusions We conclude that G:C > T:A mutations at KRAS codon 12 in the tumours of lung cancer patients (who smoke), proposed to be predominantly caused by BPDE, are due to the effect of the interaction methyl group at the C5 position of the thymine base in the KRAS sequence with the BPDE carcinogen investigated causing increased distortion. We further suggest methylated cytosine would have a similar effect, showing the importance of methylation in cancer development.

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Sequence Context Effects on Mutational Properties of cis-Opened Benzo[c]phenanthrene Diol Epoxide−Deoxyadenosine Adducts in Site-Specific Mutation Studies†

Biochemistry ◽

10.1021/bi982436l ◽

1999 ◽

Vol 38 (3) ◽

pp. 1144-1152 ◽

Cited By ~ 12

Author(s):

Ingrid Pontén ◽

Jane M. Sayer ◽

Anthony S. Pilcher ◽

Haruhiko Yagi ◽

Subodh Kumar ◽

...

Keyword(s):

Context Effects ◽

Sequence Context ◽

Site Specific ◽

Specific Mutation ◽

Diol Epoxide ◽

Deoxyadenosine Adducts

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Base damage, local sequence context andTP53mutation hotspots: a molecular dynamics study of benzo[a]pyrene induced DNA distortion and mutability

Nucleic Acids Research ◽

10.1093/nar/gkv910 ◽

2015 ◽

Vol 43 (19) ◽

pp. 9133-9146 ◽

Cited By ~ 9

Author(s):

Georgina E. Menzies ◽

Simon H. Reed ◽

Andrea Brancale ◽

Paul D. Lewis

Keyword(s):

Molecular Dynamics ◽

Sequence Context ◽

Base Damage ◽

Local Sequence

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Disruption of origin chromatin structure by helicase activation in the absence of DNA replication

10.1101/2021.03.17.435814 ◽

2021 ◽

Author(s):

Rachel A. Hoffman ◽

David M. MacAlpine

Keyword(s):

Dna Replication ◽

S Phase ◽

Replication Initiation ◽

Micrococcal Nuclease ◽

Sequence Context ◽

Α Subunit ◽

Chromatin Dynamics ◽

Conditional Mutant ◽

Local Sequence ◽

Efficient Replication

Prior to initiation of DNA replication, the eukaryotic helicase, Mcm2-7, must be activated to unwind DNA at replication start sites in early S-phase. To study helicase activation within origin chromatin, we constructed a conditional mutant of the polymerase α subunit Cdc17 (or Pol1) to prevent priming and block replication. Recovery of these cells at permissive conditions resulted in the generation of unreplicated gaps at origins, likely due to helicase activation prior to replication initiation. We used micrococcal nuclease (MNase)-based chromatin occupancy profiling under restrictive conditions to study chromatin dynamics associated with helicase activation. Helicase activation in the absence of DNA replication resulted in the disruption and disorganization of chromatin which extends up to one kilobase from early, efficient replication origins. The CMG holo-helicase complex also moves the same distance out from the origin, producing single-stranded DNA that activates the intra-S-phase checkpoint. Loss of the checkpoint did not regulate the progression and stalling of the CMG complex, but rather resulted in the disruption of chromatin at both early and late origins. Finally, we found that the local sequence context regulates helicase progression in the absence of DNA replication, suggesting that the helicase is intrinsically less processive when uncoupled from replication.

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Distant Neighbor Base Sequence Context Effects in Human Nucleotide Excision Repair of a Benzo[a]pyrene-derived DNA Lesion

Journal of Molecular Biology ◽

10.1016/j.jmb.2010.04.004 ◽

2010 ◽

Vol 399 (3) ◽

pp. 397-409 ◽

Cited By ~ 24

Author(s):

Yuqin Cai ◽

Konstantin Kropachev ◽

Rong Xu ◽

Yijin Tang ◽

Marina Kolbanovskii ◽

...

Keyword(s):

Nucleotide Excision Repair ◽

Context Effects ◽

Excision Repair ◽

Base Sequence ◽

Sequence Context ◽

Dna Lesion ◽

Nucleotide Excision ◽

Distant Neighbor

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Sequence context effects on 8-methoxypsoralen photobinding to defined DNA fragments

Biochemistry ◽

10.1021/bi00386a010 ◽

1987 ◽

Vol 26 (12) ◽

pp. 3307-3314 ◽

Cited By ~ 84

Author(s):

Evelyne Sage ◽

Ethel Moustacchi

Keyword(s):

Context Effects ◽

Dna Fragments ◽

Sequence Context

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Thermodynamic, Spectroscopic, and Equilibrium Binding Studies of DNA Sequence Context Effects in Six 22-Base Pair Deoxyoligonucleotides

Biochemistry ◽

10.1021/bi9904407 ◽

1999 ◽

Vol 38 (34) ◽

pp. 11197-11208 ◽

Cited By ~ 10

Author(s):

Peter V. Riccelli ◽

Peter M. Vallone ◽

Irina Kashin ◽

Brian D. Faldasz ◽

Michael J. Lane ◽

...

Keyword(s):

Dna Sequence ◽

Base Pair ◽

Context Effects ◽

Binding Studies ◽

Sequence Context ◽

Equilibrium Binding

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Locally acting transcription factors are required for p53-dependent cis-regulatory element activity

10.1101/761387 ◽

2019 ◽

Author(s):

Allison N. Catizone ◽

Gizem Karsli Uzunbas ◽

Petra Celadova ◽

Sylvia Kuang ◽

Daniel Bose ◽

...

Keyword(s):

Transcription Factors ◽

Regulatory Element ◽

Regulatory Elements ◽

Mrna Levels ◽

Sequence Context ◽

Endogenous Expression ◽

Damage Repair ◽

Local Sequence ◽

Element Activity ◽

Massively Parallel Reporter Assay

AbstractThe master tumor suppressor p53 controls transcription of a wide-ranging gene network involved in apoptosis, cell cycle arrest, DNA damage repair, and senescence. Recent studies revealed pervasive binding of p53 to cis-regulatory elements (CRE), which are non-coding segments of DNA that spatially and temporally control transcription through the combinatorial binding of local transcription factors (TFs). Although the role of p53 as a strong trans-activator of gene expression is well known, the co-regulatory factors and local sequences acting at p53-bound CREs are comparatively understudied. We designed and executed a massively parallel reporter assay (MPRA) to investigate the effect of transcription factor binding motifs and local sequence context on p53-bound CRE activity. Our data indicate that p53-bound CREs are both positively and negatively affected by alterations in local sequence context and changes to co-regulatory TF motifs. We identified a SP1/KLF family motif located in an intronic p53 CRE that is required for the endogenous expression of the p53-dependent gene CCNG1. We also identified ATF3 as a factor that co-regulates the expression of the p53-dependent gene GDF15 through binding with p53 in an upstream CRE. Loss of either p53 or ATF3 severely reduces CRE activity and alters endogenous GDF15 mRNA levels in the cell. Our data suggests that p53 has the flexibility to cooperate with a variety of transcription factors in order to regulate CRE activity. By utilizing different sets of co-factors across CREs, we hypothesize that p53 activity is guarded against loss of any one regulatory partner allowing for dynamic and redundant control of p53-mediated transcription.

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Primary orthologs from local sequence context

BMC Bioinformatics ◽

10.1186/s12859-020-3384-2 ◽

2020 ◽

Vol 21 (1) ◽

Cited By ~ 2

Author(s):

Kun Gao ◽

Jonathan Miller

Keyword(s):

Sequence Context ◽

Local Sequence

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Locally acting transcription factors regulate p53-dependent cis-regulatory element activity

Nucleic Acids Research ◽

10.1093/nar/gkaa147 ◽

2020 ◽

Vol 48 (8) ◽

pp. 4195-4213 ◽

Cited By ~ 2

Author(s):

Allison N Catizone ◽

Gizem Karsli Uzunbas ◽

Petra Celadova ◽

Sylvia Kuang ◽

Daniel Bose ◽

...

Keyword(s):

Transcription Factors ◽

Regulatory Element ◽

Regulatory Elements ◽

Sequence Context ◽

Binding Motifs ◽

Cycle Arrest ◽

Damage Repair ◽

Local Sequence ◽

Element Activity ◽

Massively Parallel Reporter Assay

Abstract The master tumor suppressor p53 controls transcription of a wide-ranging gene network involved in apoptosis, cell cycle arrest, DNA damage repair, and senescence. Recent studies revealed pervasive binding of p53 to cis-regulatory elements (CREs), which are non-coding segments of DNA that spatially and temporally control transcription through the combinatorial binding of local transcription factors. Although the role of p53 as a strong trans-activator of gene expression is well known, the co-regulatory factors and local sequences acting at p53-bound CREs are comparatively understudied. We designed and executed a massively parallel reporter assay (MPRA) to investigate the effect of transcription factor binding motifs and local sequence context on p53-bound CRE activity. Our data indicate that p53-bound CREs are both positively and negatively affected by alterations in local sequence context and changes to co-regulatory TF motifs. Our data suggest p53 has the flexibility to cooperate with a variety of transcription factors in order to regulate CRE activity. By utilizing different sets of co-factors across CREs, we hypothesize that global p53 activity is guarded against loss of any one regulatory partner, allowing for dynamic and redundant control of p53-mediated transcription.

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Massively parallel profiling and predictive modeling of the outcomes of CRISPR/Cas9-mediated double-strand break repair

Nucleic Acids Research ◽

10.1093/nar/gkz487 ◽

2019 ◽

Vol 47 (15) ◽

pp. 7989-8003 ◽

Cited By ~ 35

Author(s):

Wei Chen ◽

Aaron McKenna ◽

Jacob Schreiber ◽

Maximilian Haeussler ◽

Yi Yin ◽

...

Keyword(s):

Human Cell Line ◽

Strand Break ◽

Double Strand Break ◽

Lineage Tracing ◽

Dsb Repair ◽

Sequence Context ◽

Local Sequence ◽

Lentiviral Infection ◽

Repair Mechanisms ◽

Non Homologous End Joining

Abstract Non-homologous end-joining (NHEJ) plays an important role in double-strand break (DSB) repair of DNA. Recent studies have shown that the error patterns of NHEJ are strongly biased by sequence context, but these studies were based on relatively few templates. To investigate this more thoroughly, we systematically profiled ∼1.16 million independent mutational events resulting from CRISPR/Cas9-mediated cleavage and NHEJ-mediated DSB repair of 6872 synthetic target sequences, introduced into a human cell line via lentiviral infection. We find that: (i) insertions are dominated by 1 bp events templated by sequence immediately upstream of the cleavage site, (ii) deletions are predominantly associated with microhomology and (iii) targets exhibit variable but reproducible diversity with respect to the number and relative frequency of the mutational outcomes to which they give rise. From these data, we trained a model that uses local sequence context to predict the distribution of mutational outcomes. Exploiting the bias of NHEJ outcomes towards microhomology mediated events, we demonstrate the programming of deletion patterns by introducing microhomology to specific locations in the vicinity of the DSB site. We anticipate that our results will inform investigations of DSB repair mechanisms as well as the design of CRISPR/Cas9 experiments for diverse applications including genome-wide screens, gene therapy, lineage tracing and molecular recording.

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