scholarly journals DNA clamp function of the mono-ubiquitinated Fanconi Anemia FANCI-FANCD2 complex

2019 ◽  
Author(s):  
Renjing Wang ◽  
Shengliu Wang ◽  
Ankita Dhar ◽  
Christopher Peralta ◽  
Nikola P. Pavletich
Keyword(s):  
Fanconi Anemia ◽  
Translesion Synthesis ◽  
The Other ◽  
Replication Forks ◽  
Dna Structures ◽  
Cancer Predisposition Syndrome ◽  
Interstrand Crosslinks ◽  
Branched Dna ◽  
Closed Ring ◽  
Dna Interstrand Crosslinks

ABSTRACTThe FANCI-FANCD2 (ID) complex, mutated in the Fanconi Anemia (FA) cancer predisposition syndrome, is required for the repair of replication forks stalled at DNA interstrand crosslinks (ICL) and related lesions1. The FA pathway is activated when two replication forks converge onto an ICL2, triggering the mono-ubiquitination of the ID complex. ID mono-ubiquitination is essential for ICL repair by excision, translesion synthesis and homologous recombination, but its function was hitherto unknown1,3. Here, the 3.48 Å cryo-EM structure of mono-ubiquitinated ID (IDUb) bound to DNA reveals that it forms a closed ring that encircles the DNA. Compared to the cryo-EM structure of the non-ubiquitinated ID complex bound to ICL DNA, described here as well, mono-ubiquitination triggers a complete re-arrangement of the open, trough-like ID structure through the ubiquitin of one protomer binding to the other protomer in a reciprocal fashion. The structures, in conjunction with biochemical data, indicate the mono-ubiquitinated ID complex looses its preference for ICL and related branched DNA structures, becoming a sliding DNA clamp that can coordinate the subsequent repair reactions. Our findings also reveal how mono-ubiquitination in general can induce an alternate structure with a new function.

scholarly journals Differential functions of FANCI and FANCD2 ubiquitination stabilize ID2 complex on DNA

2020 ◽  
Author(s):  
Martin L. Rennie ◽  
Kimon Lemonidis ◽  
Connor Arkinson ◽  
Viduth K. Chaugule ◽  
Mairi Clarke ◽  
...  
Keyword(s):  
Fanconi Anemia ◽  
Large Scale ◽  
The Other ◽  
Molecular Function ◽  
Post Translational Modifications ◽  
Hydrophobic Residues ◽  
Interstrand Crosslinks ◽  
Double Stranded Dna ◽  
Dna Interstrand Crosslinks ◽  
Lysine Residues

AbstractThe Fanconi Anemia (FA) pathway is a dedicated pathway for the repair of DNA interstrand crosslinks, and which is additionally activated in response to other forms of replication stress. A key step in the activation of the FA pathway is the monoubiquitination of each of the two subunits (FANCI and FANCD2) of the ID2 complex on specific lysine residues. However, the molecular function of these modifications has been unknown for nearly two decades. Here we find that ubiquitination of FANCD2 acts to increase ID2’s affinity for double stranded DNA via promoting/stabilizing a large-scale conformational change in the complex, resulting in a secondary “Arm” ID2 interphase encircling DNA. Ubiquitination of FANCI, on the other hand, largely protects the ubiquitin on FANCD2 from USP1/UAF deubiquitination, with key hydrophobic residues of FANCI’s ubiquitin being important for this protection. In effect, both of these post-translational modifications function to stabilise a conformation in which the ID2 complex encircles DNA.

scholarly journals Distinct roles of BRCA2 in replication fork protection in response to hydroxyurea and DNA interstrand crosslinks

2019 ◽  
Author(s):  
Kimberly A. Rickman ◽  
Ray Noonan ◽  
Francis P. Lach ◽  
Sunandini Sridhar ◽  
Anderson T. Wang ◽  
...  
Keyword(s):  
Fanconi Anemia ◽  
Essential Gene ◽  
Dna Lesions ◽  
Replication Forks ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Interstrand Crosslinks ◽  
Pathogenic Variants ◽  
Dna Interstrand Crosslinks ◽  
Stalled Replication Forks

SummaryDNA interstrand crosslinks (ICLs) are a form of DNA damage that requires the interplay of a number of repair proteins including those of the Fanconi anemia (FA) and the homologous recombination (HR) pathways. Pathogenic variants in the essential gene BRCA2/FANCD1, when monoallelic, predispose to breast and ovarian cancer, and when biallelic, results in a severe subtype of Fanconi anemia. BRCA2 function in the FA pathway is attributed to its role as a mediator of the RAD51 recombinase in HR repair of the programmed DNA double strand breaks (DSB). BRCA2 and RAD51 functions are also required to protect stalled replication forks from nucleolytic degradation during response to hydroxyurea (HU). While RAD51 has been shown to be necessary in the early steps of ICL repair to prevent aberrant nuclease resection, the role of BRCA2 in this process has not been described. Here, based on the analysis of BRCA2 DNA binding domain (DBD) mutants discovered in FA patients presenting with atypical FA-like phenotypes, we establish that BRCA2 is necessary for protection of DNA at an ICL. Cells carrying DBD BRCA2 mutations are sensitive to ICL inducing agents but resistant to HU treatment consistent with relatively high HR repair in these cells. BRCA2 function at an ICL protects against DNA2-WRN nuclease-helicase complex and not the MRE11 nuclease implicated in the resection of HU-stalled replication forks. Our results also indicate that unlike the processing at HU-stalled forks, function of the SNF2 translocases (SMARCAL1, ZRANB3, or HLTF), implicated in fork reversal, are not an integral component of the ICL repair, pointing to a different mechanism of fork protection at different DNA lesions.

scholarly journals UHRF1 Is a Sensor for DNA Interstrand Crosslinks and Recruits FANCD2 to Initiate the Fanconi Anemia Pathway

Cell Reports ◽  
2015 ◽  
Vol 10 (12) ◽  
pp. 1947-1956 ◽  
Author(s):  
Chih-Chao Liang ◽  
Bao Zhan ◽  
Yasunaga Yoshikawa ◽  
Wilhelm Haas ◽  
Steven P. Gygi ◽  
...  
Keyword(s):  
Fanconi Anemia ◽  
Fanconi Anemia Pathway ◽  
Interstrand Crosslinks ◽  
Dna Interstrand Crosslinks

scholarly journals Bypass of DNA interstrand crosslinks by a Rev1–DNA polymerase ζ complex

2020 ◽  
Vol 48 (15) ◽  
pp. 8461-8473
Author(s):  
Rachel Bezalel-Buch ◽  
Young K Cheun ◽  
Upasana Roy ◽  
Orlando D Schärer ◽  
Peter M Burgers
Keyword(s):  
Catalytic Activity ◽  
Dna Polymerase ◽  
Nitrogen Mustard ◽  
Dna Polymerases ◽  
Translesion Synthesis ◽  
Interstrand Crosslinks ◽  
Interstrand Crosslink ◽  
Dna Interstrand Crosslinks

Abstract DNA polymerase ζ (Pol ζ) and Rev1 are essential for the repair of DNA interstrand crosslink (ICL) damage. We have used yeast DNA polymerases η, ζ and Rev1 to study translesion synthesis (TLS) past a nitrogen mustard-based interstrand crosslink (ICL) with an 8-atom linker between the crosslinked bases. The Rev1–Pol ζ complex was most efficient in complete bypass synthesis, by 2–3 fold, compared to Pol ζ alone or Pol η. Rev1 protein, but not its catalytic activity, was required for efficient TLS. A dCMP residue was faithfully inserted across the ICL-G by Pol η, Pol ζ, and Rev1–Pol ζ. Rev1–Pol ζ, and particularly Pol ζ alone showed a tendency to stall before the ICL, whereas Pol η stalled just after insertion across the ICL. The stalling of Pol η directly past the ICL is attributed to its autoinhibitory activity, caused by elongation of the short ICL-unhooked oligonucleotide (a six-mer in our study) by Pol η providing a barrier to further elongation of the correct primer. No stalling by Rev1–Pol ζ directly past the ICL was observed, suggesting that the proposed function of Pol ζ as an extender DNA polymerase is also required for ICL repair.

scholarly journals Towards a Molecular Understanding of the Fanconi Anemia Core Complex

Anemia ◽  
2012 ◽  
Vol 2012 ◽  
pp. 1-10 ◽  
Author(s):  
Charlotte Hodson ◽  
Helen Walden
Keyword(s):  
Fanconi Anemia ◽  
Molecular Mechanisms ◽  
Genome Instability ◽  
Genetic Disorder ◽  
Single Gene ◽  
Main Function ◽  
Core Complex ◽  
Interstrand Crosslinks ◽  
Dna Interstrand Crosslinks ◽  
Interstrand Crosslinking

Fanconi Anemia (FA) is a genetic disorder characterized by the inability of patient cells to repair DNA damage caused by interstrand crosslinking agents. There are currently 14 verified FA genes, where mutation of any single gene prevents repair of DNA interstrand crosslinks (ICLs). The accumulation of ICL damage results in genome instability and patients having a high predisposition to cancers. The key event of the FA pathway is dependent on an eight-protein core complex (CC), required for the monoubiquitination of each member of the FANCD2-FANCI complex. Interestingly, the majority of patient mutations reside in the CC. The molecular mechanisms underlying the requirement for such a large complex to carry out a monoubiquitination event remain a mystery. This paper documents the extensive efforts of researchers so far to understand the molecular roles of the CC proteins with regard to its main function in the FA pathway, the monoubiquitination of FANCD2 and FANCI.

scholarly journals Bypass of DNA Interstrand crosslinks by a Rev1-DNA polymerase ζ complex

2020 ◽  
Author(s):  
Rachel Bezalel-Buch ◽  
Young K. Cheun ◽  
Upasana Roy ◽  
Orlando D. Schärer ◽  
Peter M. Burgers
Keyword(s):  
Catalytic Activity ◽  
Dna Polymerase ◽  
Nitrogen Mustard ◽  
Dna Polymerases ◽  
Translesion Synthesis ◽  
Interstrand Crosslinks ◽  
Interstrand Crosslink ◽  
Dna Interstrand Crosslinks

AbstractDNA polymerase ζ (Pol ζ) and Rev1 are essential for the repair of DNA interstrand crosslink (ICL) damage. We have used yeast DNA polymerases η, ζ, and Rev1 to study translesion synthesis (TLS) past a nitrogen mustard-based ICL with an 8-atom linker between the crosslinked bases. The Rev1-Pol ζ complex was most efficient in complete bypass synthesis, by 2-3 fold, compared to Pol ζ alone or Pol η. Rev1 protein, but not its catalytic activity, was required for efficient TLS. A dCMP residue was faithfully inserted across the ICL-G by Pol η, Pol ζ, and Rev1-Pol ζ. Rev1-Pol ζ, and particularly Pol ζ alone showed a tendency to stall before the ICL, whereas Pol η stalled just after insertion across the ICL. The stalling of Pol η directly past the ICL is attributed to its autoinhibitory activity, caused by elongation of the short ICL-unhooked oligonucleotide (a six-mer in our study) by Pol η providing a barrier to further elongation of the correct primer. No stalling by Rev1-Pol ζ directly past the ICL was observed, suggesting that the proposed function of Pol ζ as an extender DNA polymerase is also required for ICL repair.

AN ONIOM STUDY ON THE CROSSLINKED BASE PAIRS IN DNA REACTED WITH CHLOROETHYLNITROSOUREAS

2007 ◽  
Vol 06 (03) ◽  
pp. 631-639 ◽  
Author(s):  
LIJIAO ZHAO ◽  
RUGANG ZHONG ◽  
YAN ZHEN
Keyword(s):  
Anticancer Agents ◽  
Hybrid Method ◽  
The Other ◽  
Base Pairs ◽  
Interstrand Crosslinks ◽  
Dna Crosslinks ◽  
Dna Interstrand Crosslinks

Chloroethylnitrosoureas (CENUs) are clinically useful anticancer agents. Their cytotoxicity is associated with the generation of DNA interstrand crosslinks. QM/MM computations are carried out to investigate DNA crosslinks by CENUs with ONIOM hybrid method. The crosslinked DNA are subdivided into three layers, each of which are described at B3LYP/6-311+G(d,p), AM1, and UFF level of theory, respectively. The result shows that the deformation of DNA with dG ( N 1)– dC ( N 3) crosslink is much less than the other crosslinks, which indicate that the most favorable crosslink is between the N1 atom of guanine and the N3 atom of the complementary cytosine.

scholarly journals Structure-dependent bypass of DNA interstrand crosslinks by translesion synthesis polymerases

2011 ◽  
Vol 39 (17) ◽  
pp. 7455-7464 ◽  
Author(s):  
The Vinh Ho ◽  
Angelo Guainazzi ◽  
Semsi Burak Derkunt ◽  
Milica Enoiu ◽  
Orlando D. Schärer
Keyword(s):  
Translesion Synthesis ◽  
Interstrand Crosslinks ◽  
Dna Interstrand Crosslinks
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