scholarly journals p53 isoforms differentially impact on the POLι dependent DNA damage tolerance pathway

2021 ◽  
Vol 12 (10) ◽  
Author(s):  
Yitian Guo ◽  
Melanie Rall-Scharpf ◽  
Jean-Christophe Bourdon ◽  
Lisa Wiesmüller ◽  
Stephanie Biber
Keyword(s):  
Dna Damage ◽  
Dna Replication ◽  
Damage Tolerance ◽  
Inhibitory Effect ◽  
Dna Damage Tolerance ◽  
Hematopoietic Stem ◽  
Stem And Progenitor Cells ◽  
Fork Stalling ◽  
P53 Isoforms ◽  
Truncated Isoforms

AbstractThe recently discovered p53-dependent DNA damage tolerance (DDT) pathway relies on its biochemical activities in DNA-binding, oligomerization, as well as complex formation with the translesion synthesis (TLS) polymerase iota (POLι). These p53-POLι complexes slow down nascent DNA synthesis for safe, homology-directed bypass of DNA replication barriers. In this study, we demonstrate that the alternative p53-isoforms p53β, p53γ, Δ40p53α, Δ133p53α, and Δ160p53α differentially affect this p53-POLι-dependent DDT pathway originally described for canonical p53α. We show that the C-terminal isoforms p53β and p53γ, comprising a truncated oligomerization domain (OD), bind PCNA. Conversely, N-terminally truncated isoforms have a reduced capacity to engage in this interaction. Regardless of the specific loss of biochemical activities required for this DDT pathway, all alternative isoforms were impaired in promoting POLι recruitment to PCNA in the chromatin and in decelerating DNA replication under conditions of enforced replication stress after Mitomycin C (MMC) treatment. Consistent with this, all alternative p53-isoforms no longer stimulated recombination, i.e., bypass of endogenous replication barriers. Different from the other isoforms, Δ133p53α and Δ160p53α caused a severe DNA replication problem, namely fork stalling even in untreated cells. Co-expression of each alternative p53-isoform together with p53α exacerbated the DDT pathway defects, unveiling impaired POLι recruitment and replication deceleration already under unperturbed conditions. Such an inhibitory effect on p53α was particularly pronounced in cells co-expressing Δ133p53α or Δ160p53α. Notably, this effect became evident after the expression of the isoforms in tumor cells, as well as after the knockdown of endogenous isoforms in human hematopoietic stem and progenitor cells. In summary, mimicking the situation found to be associated with many cancer types and stem cells, i.e., co-expression of alternative p53-isoforms with p53α, carved out interference with p53α functions in the p53-POLι-dependent DDT pathway.

scholarly journals DNA damage tolerance in hematopoietic stem and progenitor cells in mice

2017 ◽  
Vol 114 (33) ◽  
pp. E6875-E6883 ◽  
Author(s):  
Bas Pilzecker ◽  
Olimpia Alessandra Buoninfante ◽  
Paul van den Berk ◽  
Cesare Lancini ◽  
Ji-Ying Song ◽  
...  
Keyword(s):  
Dna Damage ◽  
Progenitor Cells ◽  
Damage Tolerance ◽  
Premature Aging ◽  
Intrinsic Defect ◽  
Dna Damage Tolerance ◽  
Erythroid Progenitor ◽  
Hematopoietic Stem ◽  
Fork Stalling

DNA damage tolerance (DDT) enables bypassing of DNA lesions during replication, thereby preventing fork stalling, replication stress, and secondary DNA damage related to fork stalling. Three modes of DDT have been documented: translesion synthesis (TLS), template switching (TS), and repriming. TLS and TS depend on site-specific PCNA K164 monoubiquitination and polyubiquitination, respectively. To investigate the role of DDT in maintaining hematopoietic stem cells (HSCs) and progenitors, we used PcnaK164R/K164R mice as a unique DDT-defective mouse model. Analysis of the composition of HSCs and HSC-derived multipotent progenitors (MPPs) revealed a significantly reduced number of HSCs, likely owing to increased differentiation of HSCs toward myeloid/erythroid-associated MPP2s. This skewing came at the expense of the number of lymphoid-primed MPP4s, which appeared to be compensated for by increased MPP4 proliferation. Furthermore, defective DDT decreased the numbers of MPP-derived common lymphoid progenitor (CLP), common myeloid progenitor (CMP), megakaryocyte-erythroid progenitor (MEP), and granulocyte-macrophage progenitor (GMP) cells, accompanied by increased cell cycle arrest in CMPs. The HSC and MPP phenotypes are reminiscent of premature aging and stressed hematopoiesis, and indeed progressed with age and were exacerbated on cisplatin exposure. Bone marrow transplantations revealed a strong cell intrinsic defect of DDT-deficient HSCs in reconstituting lethally irradiated mice and a strong competitive disadvantage when cotransplanted with wild-type HSCs. These findings indicate a critical role of DDT in maintaining HSCs and progenitor cells, and in preventing premature aging.

scholarly journals Error-Free DNA Damage Tolerance and Sister Chromatid Proximity during DNA Replication Rely on the Polα/Primase/Ctf4 Complex

2015 ◽  
Vol 57 (5) ◽  
pp. 812-823 ◽  
Author(s):  
Marco Fumasoni ◽  
Katharina Zwicky ◽  
Fabio Vanoli ◽  
Massimo Lopes ◽  
Dana Branzei
Keyword(s):  
Dna Damage ◽  
Dna Replication ◽  
Sister Chromatid ◽  
Damage Tolerance ◽  
Dna Damage Tolerance ◽  
Free Dna

scholarly journals DNA damage tolerance in stem cells, ageing, mutagenesis, disease and cancer therapy

2019 ◽  
Vol 47 (14) ◽  
pp. 7163-7181 ◽  
Author(s):  
Bas Pilzecker ◽  
Olimpia Alessandra Buoninfante ◽  
Heinz Jacobs
Keyword(s):  
Dna Damage ◽  
Dna Damage Response ◽  
Damage Tolerance ◽  
Essential Feature ◽  
Dna Lesions ◽  
Template Switching ◽  
Dna Damage Tolerance ◽  
Response Network ◽  
Damage Response ◽  
Fork Stalling

AbstractThe DNA damage response network guards the stability of the genome from a plethora of exogenous and endogenous insults. An essential feature of the DNA damage response network is its capacity to tolerate DNA damage and structural impediments during DNA synthesis. This capacity, referred to as DNA damage tolerance (DDT), contributes to replication fork progression and stability in the presence of blocking structures or DNA lesions. Defective DDT can lead to a prolonged fork arrest and eventually cumulate in a fork collapse that involves the formation of DNA double strand breaks. Four principal modes of DDT have been distinguished: translesion synthesis, fork reversal, template switching and repriming. All DDT modes warrant continuation of replication through bypassing the fork stalling impediment or repriming downstream of the impediment in combination with filling of the single-stranded DNA gaps. In this way, DDT prevents secondary DNA damage and critically contributes to genome stability and cellular fitness. DDT plays a key role in mutagenesis, stem cell maintenance, ageing and the prevention of cancer. This review provides an overview of the role of DDT in these aspects.

scholarly journals PCNA Ubiquitylation: Instructive or Permissive to DNA Damage Tolerance Pathways?

Biomolecules ◽  
2021 ◽  
Vol 11 (10) ◽  
pp. 1543
Author(s):  
Jun Che ◽  
Xin Hong ◽  
Hai Rao
Keyword(s):  
Dna Damage ◽  
Dna Replication ◽  
Damage Tolerance ◽  
Translesion Synthesis ◽  
Dna Lesions ◽  
Template Switching ◽  
Dna Damage Tolerance ◽  
Future Studies ◽  
Dna Lesion ◽  
Replicative Polymerases

DNA lesions escaping from repair often block the DNA replicative polymerases required for DNA replication and are handled during the S/G2 phases by the DNA damage tolerance (DDT) mechanisms, which include the error-prone translesion synthesis (TLS) and the error-free template switching (TS) pathways. Where the mono-ubiquitylation of PCNA K164 is critical for TLS, the poly-ubiquitylation of the same residue is obligatory for TS. However, it is not known how cells divide the labor between TLS and TS. Due to the fact that the type of DNA lesion significantly influences the TLS and TS choice, we propose that, instead of altering the ratio between the mono- and poly-Ub forms of PCNA, the competition between TLS and TS would automatically determine the selection between the two pathways. Future studies, especially the single integrated lesion “i-Damage” system, would elucidate detailed mechanisms governing the choices of specific DDT pathways.

Abstract 3722: Extracellular arginine starvation imposes DNA replication fork stall and permits DNA damage tolerance

2020 ◽  
Author(s):  
Yi-Chang Wang ◽  
Andrew A. Kelso ◽  
Yi-Hsuan Chen ◽  
Chi-An Hsieh ◽  
Wei-Kai Chen ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Replication ◽  
Replication Fork ◽  
Damage Tolerance ◽  
Dna Damage Tolerance

scholarly journals Access to PCNA by Srs2 and Elg1 Controls the Choice between Alternative Repair Pathways in Saccharomyces cerevisiae

mBio ◽  
2020 ◽  
Vol 11 (3) ◽  
Author(s):  
Matan Arbel ◽  
Alex Bronstein ◽  
Soumitra Sau ◽  
Batia Liefshitz ◽  
Martin Kupiec
Keyword(s):  
Dna Damage ◽  
Dna Replication ◽  
Damage Tolerance ◽  
Dna Polymerases ◽  
Strand Transfer ◽  
Dna Damage Tolerance ◽  
Dna Repair Mechanisms ◽  
Dna Damaging Agents ◽  
Recombination Protein ◽  
Repair Mechanisms

ABSTRACT During DNA replication, stalling can occur when the replicative DNA polymerases encounter lesions or hard-to replicate regions. Under these circumstances, the processivity factor PCNA gets ubiquitylated at lysine 164, inducing the DNA damage tolerance (DDT) mechanisms that can bypass lesions encountered during DNA replication. PCNA can also be SUMOylated at the same residue or at lysine 127. Surprisingly, pol30-K164R mutants display a higher degree of sensitivity to DNA-damaging agents than pol30-KK127,164RR strains, unable to modify any of the lysines. Here, we show that in addition to translesion synthesis and strand-transfer DDT mechanisms, an alternative repair mechanism (“salvage recombination”) that copies information from the sister chromatid is repressed by the recruitment of Srs2 to SUMOylated PCNA. Overexpression of Elg1, the PCNA unloader, or of the recombination protein Rad52 allows its activation. We dissect the genetic requirements for this pathway, as well as the interactions between Srs2 and Elg1. IMPORTANCE PCNA, the ring that encircles DNA maintaining the processivity of DNA polymerases, is modified by ubiquitin and SUMO. Whereas ubiquitin is required for bypassing lesions through the DNA damage tolerance (DDT) pathways, we show here that SUMOylation represses another pathway, salvage recombination. The Srs2 helicase is recruited to SUMOylated PCNA and prevents the salvage pathway from acting. The pathway can be induced by overexpressing the PCNA unloader Elg1, or the homologous recombination protein Rad52. Our results underscore the role of PCNA modifications in controlling the various bypass and DNA repair mechanisms.

scholarly journals Multiple biochemical properties of the p53 molecule contribute to activation of polymerase iota-dependent DNA damage tolerance

2020 ◽  
Vol 48 (21) ◽  
pp. 12188-12203
Author(s):  
Stephanie Biber ◽  
Helmut Pospiech ◽  
Vanesa Gottifredi ◽  
Lisa Wiesmüller
Keyword(s):  
Dna Damage ◽  
Dna Replication ◽  
Topoisomerase I ◽  
Mutational Analysis ◽  
Damage Tolerance ◽  
Biochemical Properties ◽  
Replication Proteins ◽  
Dna Damage Tolerance ◽  
C Terminus ◽  
Cancer Tissues

Abstract We have previously reported that p53 decelerates nascent DNA elongation in complex with the translesion synthesis (TLS) polymerase ι (POLι) which triggers a homology-directed DNA damage tolerance (DDT) pathway to bypass obstacles during DNA replication. Here, we demonstrate that this DDT pathway relies on multiple p53 activities, which can be disrupted by TP53 mutations including those frequently found in cancer tissues. We show that the p53-mediated DDT pathway depends on its oligomerization domain (OD), while its regulatory C-terminus is not involved. Mutation of residues S315 and D48/D49, which abrogate p53 interactions with the DNA repair and replication proteins topoisomerase I and RPA, respectively, and residues L22/W23, which disrupt formation of p53-POLι complexes, all prevent this DDT pathway. Our results demonstrate that the p53-mediated DDT requires the formation of a DNA binding-proficient p53 tetramer, recruitment of such tetramer to RPA-coated forks and p53 complex formation with POLι. Importantly, our mutational analysis demonstrates that transcriptional transactivation is dispensable for the POLι-mediated DDT pathway, which we show protects against DNA replication damage from endogenous and exogenous sources.

scholarly journals DNA Damage Tolerance Mechanisms Revealed from the Analysis of Immunoglobulin V Gene Diversification in Avian DT40 Cells

Genes ◽  
2018 ◽  
Vol 9 (12) ◽  
pp. 614 ◽  
Author(s):  
Takuya Abe ◽  
Dana Branzei ◽  
Kouji Hirota
Keyword(s):  
Dna Damage ◽  
Dna Replication ◽  
B Lymphocyte ◽  
Damage Tolerance ◽  
Base Pairs ◽  
Dna Damage Tolerance ◽  
Abasic Sites ◽  
Activation Induced Deaminase ◽  
Template Switch ◽  
Dt40 Cells

DNA replication is an essential biochemical reaction in dividing cells that frequently stalls at damaged sites. Homologous/homeologous recombination (HR)-mediated template switch and translesion DNA synthesis (TLS)-mediated bypass processes release arrested DNA replication forks. These mechanisms are pivotal for replication fork maintenance and play critical roles in DNA damage tolerance (DDT) and gap-filling. The avian DT40 B lymphocyte cell line provides an opportunity to examine HR-mediated template switch and TLS triggered by abasic sites by sequencing the constitutively diversifying immunoglobulin light-chain variable gene (IgV). During IgV diversification, activation-induced deaminase (AID) converts dC to dU, which in turn is excised by uracil DNA glycosylase and yields abasic sites within a defined window of around 500 base pairs. These abasic sites can induce gene conversion with a set of homeologous upstream pseudogenes via the HR-mediated template switch, resulting in templated mutagenesis, or can be bypassed directly by TLS, resulting in non-templated somatic hypermutation at dC/dG base pairs. In this review, we discuss recent works unveiling IgV diversification mechanisms in avian DT40 cells, which shed light on DDT mode usage in vertebrate cells and tolerance of abasic sites.

scholarly journals DNA damage tolerance pathway involving DNA polymerase ι and the tumor suppressor p53 regulates DNA replication fork progression

2016 ◽  
Vol 113 (30) ◽  
pp. E4311-E4319 ◽  
Author(s):  
Stephanie Hampp ◽  
Tina Kiessling ◽  
Kerstin Buechle ◽  
Sabrina F. Mansilla ◽  
Jürgen Thomale ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Replication ◽  
Tumor Suppressor ◽  
Damage Tolerance ◽  
Replication Stress ◽  
Nuclear Antigen ◽  
Replication Forks ◽  
Tumor Suppressor P53 ◽  
Dna Damage Tolerance ◽  
Direct Role

DNA damage tolerance facilitates the progression of replication forks that have encountered obstacles on the template strands. It involves either translesion DNA synthesis initiated by proliferating cell nuclear antigen monoubiquitination or less well-characterized fork reversal and template switch mechanisms. Herein, we characterize a novel tolerance pathway requiring the tumor suppressor p53, the translesion polymerase ι (POLι), the ubiquitin ligase Rad5-related helicase-like transcription factor (HLTF), and the SWI/SNF catalytic subunit (SNF2) translocase zinc finger ran-binding domain containing 3 (ZRANB3). This novel p53 activity is lost in the exonuclease-deficient but transcriptionally active p53(H115N) mutant. Wild-type p53, but not p53(H115N), associates with POLι in vivo. Strikingly, the concerted action of p53 and POLι decelerates nascent DNA elongation and promotes HLTF/ZRANB3-dependent recombination during unperturbed DNA replication. Particularly after cross-linker–induced replication stress, p53 and POLι also act together to promote meiotic recombination enzyme 11 (MRE11)-dependent accumulation of (phospho-)replication protein A (RPA)-coated ssDNA. These results implicate a direct role of p53 in the processing of replication forks encountering obstacles on the template strand. Our findings define an unprecedented function of p53 and POLι in the DNA damage response to endogenous or exogenous replication stress.

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