The human RAD52 complex undergoes phase separation and facilitates bundling and end-to-end tethering of RAD51 presynaptic filaments.

Human RAD52 is a prime target for synthetical lethality approaches to treat cancers with deficiency in homologous recombination. Among multiple cellular roles of RAD52, its functions in homologous recombination repair and protection of stalled replication forks appear to substitute those of the tumor suppressor protein BRCA2. However, the mechanistic details of how RAD52 can substitute BRCA2 functions are only beginning to emerge.

A double-ring of human RAD52 remodels replication forks restricting fork reversal.

Unlabelled: Human RAD52 is a multifunctional DNA repair protein involved in several cellular events that support genome stability including protection of stalled DNA replication forks from excessive degradation . In its gatekeeper role, RAD52 binds to and stabilizes stalled replication forks during replication stress protecting them from reversal by SMARCAL1 . The structural and molecular mechanism of the RAD52-mediated fork protection remains elusive.

Contrasting roles of different mismatch repair proteins in basal-like breast cancer.

The mismatch repair (MMR) pathway is known as a tumor suppressive pathway and genes involved in MMR are commonly mutated in hereditary colorectal or other cancer types. However, the function of MMR genes/proteins in breast cancer progression and metastasis are largely unknown. We found that MSH2, but not MLH1, is highly enriched in basal-like breast cancer (BLBC) and that its protein expression is inversely correlated with overall survival time (OS).

Single-Molecule Analysis of the Improved Variants of the G-Quadruplex Recognition Protein G4P.

As many as 700,000 unique sequences in the human genome are predicted to fold into G-quadruplexes (G4s), non-canonical structures formed by Hoogsteen guanine-guanine pairing within G-rich nucleic acids. G4s play both physiological and pathological roles in many vital cellular processes including DNA replication, DNA repair and RNA transcription. Several reagents have been developed to visualize G4s in vitro and in cells.

Single-molecule analysis of the improved variants of the G-quadruplex recognition protein G4P.

As many as 700,000 unique sequences in the human genome are predicted to fold into G-quadruplexes (G4s), non-canonical structures formed by Hoogsteen guanine-guanine pairing within G-rich nucleic acids. G4s play both physiological and pathological roles in many vital cellular processes including DNA replication, DNA repair and RNA transcription. Several reagents have been developed to visualize G4s vitro and in cells.

Human hnRNPA1 reorganizes telomere-bound Replication Protein A.

Unlabelled: Human replication protein A (RPA) is a heterotrimeric ssDNA binding protein responsible for many aspects of cellular DNA metabolism. Dynamic interactions of the four RPA DNA binding domains (DBDs) with DNA control replacement of RPA by downstream proteins in various cellular metabolic pathways. RPA plays several important functions at telomeres where it binds to and melts telomeric G-quadruplexes, non-canonical DNA structures formed at the G-rich telomeric ssDNA overhangs.

Therapeutic disruption of RAD52-ssDNA complexation via novel drug-like inhibitors.

RAD52 protein is a coveted target for anticancer drug discovery. Similar to poly-ADP-ribose polymerase (PARP) inhibitors, pharmacological inhibition of RAD52 is synthetically lethal with defects in genome caretakers BRCA1 and BRCA2 (∼25% of breast and ovarian cancers). Emerging structure activity relationships for RAD52 are complex, making it challenging to transform previously identified disruptors of the RAD52-ssDNA interaction into drug-like leads using traditional medicinal chemistry approaches.

RAD52 prevents accumulation of -dependent replication gaps at perturbed replication forks in human cells.

Replication gaps can arise as a consequence of perturbed DNA replication and their accumulation might undermine the stability of the genome. Loss of RAD52, a protein involved in the regulation of fork reversal, promotes accumulation of parental ssDNA gaps during replication perturbation. Here, we demonstrate that this is due to the engagement of downstream of the extensive degradation of perturbed replication forks after their reversal, and is not dependent on PrimPol.

Fork-Remodeling Helicase Rad5 Preferentially Reverses Replication Forks with Gaps in the Leading Strand.

DNA damage bypass pathways promote the replication of damaged DNA when replication forks stall at sites of DNA damage. Template switching is a DNA damage bypass pathway in which fork-reversal helicases convert stalled replication forks into four-way DNA junctions called chicken foot intermediates, which are subsequently extended by replicative DNA polymerases. In yeast, fork-reversal is carried out by the Rad5 helicase using an unknown mechanism.

Gain-of-function mutations in RPA1 cause a syndrome with short telomeres and somatic genetic rescue.

Human telomere biology disorders (TBD)/short telomere syndromes (STS) are heterogeneous disorders caused by inherited loss-of-function mutations in telomere-associated genes. Here, we identify 3 germline heterozygous missense variants in the RPA1 gene in 4 unrelated probands presenting with short telomeres and varying clinical features of TBD/STS, including bone marrow failure, myelodysplastic syndrome, T- and B-cell lymphopenia, pulmonary fibrosis, or skin manifestations. All variants cluster to DNA-binding domain A of RPA1 protein.

Switch-like control of helicase processivity by single-stranded DNA binding protein.

Helicases utilize nucleotide triphosphate (NTP) hydrolysis to translocate along single-stranded nucleic acids (NA) and unwind the duplex. In the cell, helicases function in the context of other NA-associated proteins such as single-stranded DNA binding proteins. Such encounters regulate helicase function, although the underlying mechanisms remain largely unknown.

RPA complexes in Caenorhabditis elegans meiosis; unique roles in replication, meiotic recombination and apoptosis.

Replication Protein A (RPA) is a critical complex that acts in replication and promotes homologous recombination by allowing recombinase recruitment to processed DSB ends. Most organisms possess three RPA subunits (RPA1, RPA2, RPA3) that form a trimeric complex critical for viability. The Caenorhabditis elegans genome encodes RPA-1, RPA-2 and an RPA-2 paralog RPA-4.

PCNA activates the MutLγ endonuclease to promote meiotic crossing over.

During meiosis, crossover recombination connects homologous chromosomes to direct their accurate segregation. Defective crossing over causes infertility, miscarriage and congenital disease. Each pair of chromosomes attains at least one crossover via the formation and biased resolution of recombination intermediates known as double Holliday junctions.

Physiological and Pathological Roles of RAD52 at DNA Replication Forks.

Understanding basic molecular mechanisms underlying the biology of cancer cells is of outmost importance for identification of novel therapeutic targets and biomarkers for patient stratification and better therapy selection. One of these mechanisms, the response to replication stress, fuels cancer genomic instability. It is also an Achille's heel of cancer.

Author Correction: Rad52 prevents excessive replication fork reversal and protects from nascent strand degradation.

The original version of this Article contained an error in Fig. 2. The immunofluorescence images in panel d were inadvertently replaced with duplicates of those in panel c during final assembly of the figure.

Rad52 prevents excessive replication fork reversal and protects from nascent strand degradation.

Stabilisation of stalled replication forks prevents excessive fork reversal and their pathological degradation, which can undermine genome integrity. Here we investigate a physiological role of RAD52 at stalled replication forks by using human cell models depleted of RAD52, a specific small-molecule inhibitor of the RAD52-ssDNA interaction, in vitro and single-molecule analyses. We demonstrate that RAD52 prevents excessive degradation of reversed replication forks by MRE11.

Direct Copolymerization of CO2 and Diols.

Direct polymerization of CO2 and diols is promising as a simple and environmental-benign method in place of conventional processes using high-cost and/or hazardous reagents such as phosgene, carbon monoxide and epoxides, however, there are no reports on the direct method due to the inertness of CO2 and severe equilibrium limitation of the reaction. Herein, we firstly substantiate the direct copolymerization of CO2 and diols using CeO2 catalyst and 2-cyanopyridine promotor, providing the alternating cooligomers in high diol-based yield (up to 99%) and selectivity (up to >99%). This catalyst system is applicable to various diols including linear C4-C10 α,ω-diols to provide high yields of the corresponding cooligomers, which cannot be obtained by well-known methods such as copolymerization of CO2 and cyclic ethers and ring-opening polymerization of cyclic carbonates.

Mismatch repair protein hMSH2-hMSH6 recognizes mismatches and forms sliding clamps within a D-loop recombination intermediate.

High fidelity homologous DNA recombination depends on mismatch repair (MMR), which antagonizes recombination between divergent sequences by rejecting heteroduplex DNA containing excessive nucleotide mismatches. The hMSH2-hMSH6 heterodimer is the first responder in postreplicative MMR and also plays a prominent role in heteroduplex rejection. Whether a similar molecular mechanism underlies its function in these two processes remains enigmatic.

Single-molecule study of the CUG repeat-MBNL1 interaction and its inhibition by small molecules.

Effective drug discovery and optimization can be accelerated by techniques capable of deconvoluting the complexities often present in targeted biological systems. We report a single-molecule approach to study the binding of an alternative splicing regulator, muscleblind-like 1 protein (MBNL1), to (CUG)n = 4,6 and the effect of small molecules on this interaction. Expanded CUG repeats (CUG(exp)) are the causative agent of myotonic dystrophy type 1 by sequestering MBNL1.

Tyrosine phosphorylation enhances RAD52-mediated annealing by modulating its DNA binding.

RAD52 protein has an important role in homology-directed DNA repair by mediating RAD51 nucleoprotein filament formation on single-stranded DNA (ssDNA) protected by replication protein-A (RPA) and annealing of RPA-coated ssDNA. In human, cellular response to DNA damage includes phosphorylation of RAD52 by c-ABL kinase at tyrosine 104. To address how this phosphorylation modulates RAD52 function, we used an amber suppressor technology to substitute tyrosine 104 with chemically stable phosphotyrosine analogue (p-Carboxymethyl-L-phenylalanine, pCMF).

Ensemble and single-molecule fluorescence-based assays to monitor DNA binding, translocation, and unwinding by iron-sulfur cluster containing helicases.

Many quantitative approaches for analysis of helicase-nucleic acid interactions require a robust and specific signal, which reports on the presence of the helicase and its position on a nucleic acid lattice. Since 2006, iron-sulfur (FeS) clusters have been found in a number of helicases. They serve as endogenous quenchers of Cy3 and Cy5 fluorescence which can be exploited to characterize FeS cluster containing helicases both in ensemble-based assays and at the single-molecule level.