Sterile inflammation of peritoneal membrane caused by peritoneal dialysis: focus on the communication between immune cells and peritoneal stroma.

Peritoneal dialysis is a widely used method for treating kidney failure. However, over time, the peritoneal structure and function can deteriorate, leading to the failure of this therapy. This deterioration is primarily caused by infectious and sterile inflammation. Sterile inflammation, which is inflammation without infection, is particularly concerning as it can be subtle and often goes unnoticed. The onset of sterile inflammation involves various pathological processes. Peritoneal cells detect signals that promote inflammation and release substances that attract immune cells from the bloodstream. These immune cells contribute to the initiation and escalation of the inflammatory response. The existing literature extensively covers the involvement of different cell types in the sterile inflammation, including mesothelial cells, fibroblasts, endothelial cells, and adipocytes, as well as immune cells such as macrophages, lymphocytes, and mast cells. These cells work together to promote the occurrence and progression of sterile inflammation, although the exact mechanisms are not fully understood. This review aims to provide a comprehensive overview of the signals from both stromal cells and components of immune system, as well as the reciprocal interactions between cellular components, during the initiation of sterile inflammation. By understanding the cellular and molecular mechanisms underlying sterile inflammation, we may potentially develop therapeutic interventions to counteract peritoneal membrane damage and restore normal function.

Effect of low-calcium and standard-calcium dialysate on serum calcium, phosphorus and full-segment parathyroid hormone in patients on peritoneal dialysis: A retrospective observational study.

OBJECTIVE: To investigate the effects of low-calcium and standard-calcium dialysate in patients with chronic kidney disease on peritoneal dialysis, and find out which dialysate has less vascular calcification effect. METHODS: A total of 141 patients who had undergone peritoneal dialysis (PD) for 2 years in the PD centre from January 2012 to December 2017 were included and divided into two groups according to the calcium concentration of the PD fluid used. There were 79 cases in the low-calcium group, with a dialysate calcium concentration of 1.25 mmol/L and 62 cases in the standard-calcium group, with a dialysate calcium concentration of 1.75 mmol/L. The demographic characteristics and clinical information before initiation of PD were collected and compared between the two groups. Information on the serum calcium, phosphorus and PTH, systolic and diastolic blood pressures and the use of antihypertensive and phosphate-lowering drugs in the second year of dialysis was also collected and compared between the two groups. Vascular calcification was assessed in patients on PD treatment. RESULTS: The mean serum calcium concentrations before initiation of PD in the low- and standard-calcium groups were 1.94 ± 0.27 and 1.89 ± 0.28 mmol/L, respectively. The serum calcium concentrations after PD were 2.30 ± 0.21 and 2.41 ± 0.23 mmol/L, respectively. After PD, the serum calcium concentration in both groups was significantly increased (p < 0.05). The serum calcium concentration in the low-calcium group after PD treatment was lower than that in the standard-calcium group, and the difference was statistically significant (p < 0.05). Compared with the standard-calcium group, patients in the low-calcium group had significantly higher parathyroid hormone concentrations (p < 0.05). More types of phosphate-lowering drugs were used (59.49%) in the low-calcium group than that in the standard-calcium group (35.48%; p < 0.05). The number of antihypertensive drug usage were also higher in the low-calcium group, and the difference was statistically significant (p < 0.05). As for the vascular calcification effect, the two groups have shown no statistical difference in abdominal aortic calcification rate, carotid arteriosclerosis rate and aortic arch calcification rate (p < 0.05). CONCLUSION: We found that low-calcium PD fluid may increase the PTH level and the proportion of CKD patients using antihypertensive drug and phosphorus-lowering drug, but the vascular calcification effect of the low and standard calcium PD fluid needs further exploration. This paper provides new evidence for the choice of dialysate for PD, low-calcium dialysate has no outstanding advantages for long term dialysis.

Association between dialysis effluent leukocyte count after initial antibiotic treatment and outcomes of patients with peritoneal dialysis-associated peritonitis: a retrospective study.

BACKGROUND: Among patients with peritoneal dialysis-associated peritonitis (PDAP), It has been regarded as an indicator of deterioration of clinical condition that peritoneal dialysis effluent leukocyte count (PDELC) cannot be restored to normal after initial antibiotic therapy. However, the precise relationship between PDELC on day 5 and the clinical outcomes of PDAP episodes remains uncertain. AIMS: To explore the association between PDELC on day 5 and clinical outcomes of PDAP episodes. METHODS: This retrospective study was based on the medical chart database of the Affiliated Hospital of Guangdong Medical University. Multivariable regressions were used to evaluate the association between PDELC on day 5 and 60-day mortality, half-year mortality, treatment failure, and the length of stay in hospital with adjustment for confounding factors. RESULTS: A total of 549 PDAP episodes in 309 patients were enrolled. The total 60-day mortality, half-year mortality, and rate of treatment failure was 6.0%, 9.8%, and 14.2%, respectively. Compared with patients with normal PDELC, those with PDELC ≥2000 × 106/L on day 5 had significantly higher 60-day mortality (31.1% vs 2.7%), half-year mortality (35.6% vs 5.6%), and treatment failure (46.7% vs 5.7%). In multivariate adjusted regression, the ORs (95%CI) were 6.99 (2.33, 20.92; p = 0.001), 4.97(1.93, 12.77; p = 0.001), and 5.77 (2.07, 16.11; p = 0.001), respectively. Patients with PDELC were 100-2000 × 106/L on day 5 had a higher rate of treatment failure than those with normal PDELC (26.9% vs 5.7%) (OR = 3.03, 95%CI 1.42, 6.46; p = 0.004). After sensitivity analysis, the results remained robust. CONCLUSIONS: Among patients with PDAP, increased PDELC on day 5 was associated with a greater risk of 60-day mortality, half-year mortality, and treatment failure.

Autophagy in peritoneal fibrosis.

Peritoneal dialysis (PD) is a widely accepted renal replacement therapy for patients with end-stage renal disease (ESRD). Morphological and functional changes occur in the peritoneal membranes (PMs) of patients undergoing long-term PD. Peritoneal fibrosis (PF) is a common PD-related complication that ultimately leads to PM injury and peritoneal ultrafiltration failure. Autophagy is a cellular process of "self-eating" wherein damaged organelles, protein aggregates, and pathogenic microbes are degraded to maintain intracellular environment homeostasis and cell survival. Growing evidence shows that autophagy is involved in fibrosis progression, including renal fibrosis and hepatic fibrosis, in various organs. Multiple risk factors, including high-glucose peritoneal dialysis solution (HGPDS), stimulate the activation of autophagy, which participates in PF progression, in human peritoneal mesothelial cells (HPMCs). Nevertheless, the underlying roles and mechanisms of autophagy in PF progression remain unclear. In this review, we discuss the key roles and potential mechanisms of autophagy in PF to offer novel perspectives on future therapy strategies for PF and their limitations.

The Influence of Space Transformation of Land Use on Function Transformation and the Regional Differences in Shaanxi Province.

The development of economy and urbanization promotes the transformation of land use both in space and function. Most existing research perspectives focus only on the transformation of space or function, and analysis of the mutual feedback mechanism between space transformation and function transformation is not deep enough. Therefore, this study constructed a diagnostic method for land use space transformation and function transformation and explored the mutual feedback mechanism between space transformation and functional transformation. The purpose is to deepen the research of land system science, provide a new method for diagnosing the space transformation and function transformation of land use, and clarify the driving mechanism of space transformation on function transformation as well as the mutual feedback mechanism of both. The research results showed that: (1) From 1980 to 2000, the space transformation of land use in the Northern Shaanxi Plateau did not happen, but the degree of trade-off between functions increased, resulting in the occurrence of unsustainable function transformation; the Guanzhong Plain showed a sustainable space transformation during the study period, but the land use functions did not transform; the function transformation and space transformation of the Qinba Mountain area happened in the same direction, and both tended to be sustainable; (2) From 2000 to 2018, the space transformation and function transformation of the Northern Shaanxi Plateau were in opposite directions; the land use space in the Qinba Mountain area was in a state of fluctuation and had not undergone transition, but the land use functions were in an unsustainable transition state; and (3) The function transformation in Shaanxi Province was deeply affected by factors at the policy and cultural levels and the strengthening of its own anti-interference ability, resulting in different performances of space transformation in different regions in different periods. Therefore, Shaanxi Province should rationally plan land resources, coordinate the relationship between space transformation and function transformation, and offer positive feedback to function transformation through sustainable space transformation. Meanwhile, it is necessary to prudently determine the regional land use model according to regional differences.

Insights into homology search from cryo-EM structures of RecA-DNA recombination intermediates.

The fundamental reaction in homologous recombination is the exchange of strands between two homologous DNA molecules. This reaction is carried out by the RecA family of ATPases that polymerize on ssDNA to form a presynaptic filament. This filament then binds to dsDNA to form a synaptic filament, a key intermediate that mediates the search for homology and subsequent strand exchange to produce a new heteroduplex. A recent cryo-EM analysis of synaptic filaments has now shed light on this process. The dsDNA strands are separated on binding to the filament. One strand is sequestrated while the other is freed to sample pairing with the ssDNA. Homology, through heteroduplex formation, promotes dsDNA opening. Lack of homology suppresses it, keeping local synapses short so that multiple synapses can form and increasing the probability of encountering homology.

Mechanism of strand exchange from RecA-DNA synaptic and D-loop structures.

The strand-exchange reaction is central to homologous recombination. It is catalysed by the RecA family of ATPases, which form a helical filament with single-stranded DNA (ssDNA) and ATP. This filament binds to a donor double-stranded DNA (dsDNA) to form synaptic filaments, which search for homology and then catalyse the exchange of the complementary strand, forming either a new heteroduplex or-if homology is limited-a D-loop1,2. How synaptic filaments form, search for homology and catalyse strand exchange is poorly understood. Here we report the cryo-electron microscopy analysis of synaptic mini-filaments with both non-complementary and partially complementary dsDNA, and structures of RecA-D-loop complexes containing a 10- or a 12-base-pair heteroduplex. The C-terminal domain of RecA binds to dsDNA and directs it to the RecA L2 loop, which inserts into and opens up the duplex. The opening propagates through RecA sequestering the homologous strand at a secondary DNA-binding site, which frees the complementary strand to sample pairing with the ssDNA. At each RecA step, there is a roughly 20% probability that duplex opening will terminate and the as-yet-unopened dsDNA portion will bind to another C-terminal domain. Homology suppresses this process, through the cooperation of heteroduplex pairing with the binding of ssDNA to the secondary site, to extend dsDNA opening. This mechanism locally limits the length of ssDNA sampled for pairing if homology is not encountered, and could allow for the formation of multiple, widely separated synapses on the donor dsDNA, which would increase the likelihood of encountering homology. These findings provide key mechanistic insights into homologous recombination.

Preparation and Properties of Self-Healing and Self-Lubricating Epoxy Coatings with Polyurethane Microcapsules Containing Bifunctional Linseed Oil.

An organic coating is commonly used to protect metal from corrosion, but it is prone to failure due to microcracks generated by internal stress and external mechanical action. The self-healing and self-lubricating achieved in the coating is novel, which allows an extension of life by providing resistance to damage and repair after damage. In this study, a new approach to microencapsulating bifunctional linseed oil with polyurethane shell by interfacial polymerization. Moreover, the self-healing and self-lubricating coatings with different concentrations of microcapsules were developed. The well-dispersed microcapsules showed a regular spherical morphology with an average diameter of ~64.9 µm and a core content of 74.0 wt.%. The results of the salt spray test demonstrated that coatings containing microcapsules still possess anticorrosion, which is improved with the increase of microcapsules content, after being scratched. The results of electrochemical impedance spectroscopy showed a |Z|f=0.01Hz value of 104 Ω·cm2 for pure epoxy coating after being immersed for 3 days, whereas the coating with 20 wt.% microcapsules was the highest, 1010 Ω·cm2. The results of friction wear showed that the tribological performance of the coating was enhanced greatly as microcapsule concentration reached 10 wt.% or more, which showed a 86.8% or more reduction in the friction coefficient compared to the pure epoxy coating. These results indicated that the coatings containing microcapsules exhibited excellent self-healing and self-lubricating properties, which are positively correlated with microcapsules content.

The Influence of Land Use Change on Ecosystem Service Value in Shangzhou District.

Land use change has an impact on the ecosystem service value because it changes the structure and function of ecosystems. This paper analyzed the changes in land use during the period from 2000 to 2015 in Shangzhou district, and used the equivalent value of ecological services per unit area of land ecosystem combining the natural and economic conditions of Shangzhou district. Based on this method, the ecological service value of Shangzhou district was estimated, and the impact of land use change on the ecological service value was analyzed. The results showed that: (1) the main types of land use in Shangzhou district were grassland, woodland and farmland, among which the contribution rate of woodland to the value of local ecosystem services was the highest; (2) the overall trend in the ecosystem service value in Shangzhou district increased between 2000 and 2015, from 10.74 × 108 yuan in 2000 to 20.32 × 108 yuan in 2015, which is the result of the combined effects of regional economic development and changes in the natural environment and land use patterns; and (3) the main reason for the value increase of ecosystem services in Shangzhou district between 2000 and 2015 was that the grain-for-green policy transformed a considerable amount of farmland into woodland, while the main reasons for a decline in value was the expansion of built-up land that occupied other types of land.

Voting Simulation based Agglomerative Hierarchical Method for Network Community Detection.

Community detection has been paid much attention in many fields in recent years, and a great deal of community-detection methods have been proposed. But the time consumption of some of them is heavy, limiting them from being applied to large-scale networks. On the contrary, there exist some lower-time-complexity methods. But most of them are non-deterministic, meaning that running the same method many times may yield different results from the same network, which reduces their practical utility greatly in real-world applications. To solve these problems, we propose a community-detection method in this paper, which takes both the quality of the results and the efficiency of the detecting procedure into account. Moreover, it is a deterministic method which can extract definite community structures from networks. The proposed method is inspired by the voting behaviours in election activities in the social society, in which we first simulate the voting procedure on the network. Every vertex votes for the nominated candidates following the proposed voting principles, densely connected groups of vertices can quickly reach a consensus on their candidates. At the end of this procedure, candidates and their own voters form a group of clusters. Then, we take the clusters as initial communities, and agglomerate some of them into larger ones with high efficiency to obtain the resulting community structures. We conducted extensive experiments on some artificial networks and real-world networks, the experimental results show that our proposed method can efficiently extract high-quality community structures from networks, and outperform the comparison algorithms significantly.

Mechanisms of mTORC1 activation by RHEB and inhibition by PRAS40.

The mechanistic target of rapamycin complex 1 (mTORC1) controls cell growth and metabolism in response to nutrients, energy levels, and growth factors. It contains the atypical kinase mTOR and the RAPTOR subunit that binds to the Tor signalling sequence (TOS) motif of substrates and regulators. mTORC1 is activated by the small GTPase RHEB (Ras homologue enriched in brain) and inhibited by PRAS40. Here we present the 3.0 ångström cryo-electron microscopy structure of mTORC1 and the 3.4 ångström structure of activated RHEB-mTORC1. RHEB binds to mTOR distally from the kinase active site, yet causes a global conformational change that allosterically realigns active-site residues, accelerating catalysis. Cancer-associated hyperactivating mutations map to structural elements that maintain the inactive state, and we provide biochemical evidence that they mimic RHEB relieving auto-inhibition. We also present crystal structures of RAPTOR-TOS motif complexes that define the determinants of TOS recognition, of an mTOR FKBP12-rapamycin-binding (FRB) domain-substrate complex that establishes a second substrate-recruitment mechanism, and of a truncated mTOR-PRAS40 complex that reveals PRAS40 inhibits both substrate-recruitment sites. These findings help explain how mTORC1 selects its substrates, how its kinase activity is controlled, and how it is activated by cancer-associated mutations.

Role of basophils in rheumatoid arthritis (Review).

The T helper (Th)1/Th2 imbalance plays a crucial role in the development of rheumatoid arthritis (RA). It is well known that basophils can affect the Th1/Th2 balance by enhancing the Th2 response, while impairing the Th1 response, which is known to be involved in the development of a number of diseases. However, limited information is available with regard to the role of basophils in RA. Decreased levels of circulating basophils and a dominant Th1 response have been reported in adult patients with RA, while children with juvenile RA have been largely found to have increased levels of circulating basophils and a dominant Th2 response. Furthermore, the circulating basophils in the two conditions have an activated phenotype and are associated with disease activity. In addition, a longitudinal study found the Th2 response was dominant in the early stages of RA, while the Th1 response was dominant in long-term chronic RA. These observations indicate that basophils may be involved in the development of RA by affecting the Th1/Th2 balance, particularly in the early stages of RA. Therefore, targeting basophils may be a novel therapeutic strategy for the treatment of RA; however, further investigation is required.

Optical microscopy with flexible axial capabilities using a vari-focus liquid lens.

The axial imaging range of optical microscopy is restricted by its fixed working plane and limited depth of field. In this paper, the axial capabilities of an off-the-shelf microscope is improved by inserting a liquid lens, which can be controlled by a driving electrical voltage, into the optical path of the microscope. First, the numerical formulas of the working distance and the magnification with the variation of the focus of the liquid lens are inferred using a ray tracing method and conclusion is obtained that the best position for inserting a liquid lens with consistent magnification is the aperture plane and the rear focal plane of the objective lens. Second, with the liquid lens embedded in the microscope, the numerical relationship between the magnification and the working distance of the proposed flexible-axial-capability microscope and the liquid lens driving voltage is calibrated and fitted using the inferred numerical formulas. Third, techniques including autofocus, extending depth of field and three-dimensional imaging are researched and applied, improving the designed microscope to not only flexibly control its working distance, but also to extend the depth of field near the variable working plane. Experiments show that the presented flexible-axial-capability microscope has a long working distance range of 8 mm, and by calibrating the magnification curve within the working distance range, samples can be observed and measured precisely. The depth of field can be extended to 400 µm from the variable working plane and is 20 times that of the off-the-shelf microscope.

mTOR kinase structure, mechanism and regulation.

The mammalian target of rapamycin (mTOR), a phosphoinositide 3-kinase-related protein kinase, controls cell growth in response to nutrients and growth factors and is frequently deregulated in cancer. Here we report co-crystal structures of a complex of truncated mTOR and mammalian lethal with SEC13 protein 8 (mLST8) with an ATP transition state mimic and with ATP-site inhibitors. The structures reveal an intrinsically active kinase conformation, with catalytic residues and a catalytic mechanism remarkably similar to canonical protein kinases. The active site is highly recessed owing to the FKBP12-rapamycin-binding (FRB) domain and an inhibitory helix protruding from the catalytic cleft. mTOR-activating mutations map to the structural framework that holds these elements in place, indicating that the kinase is controlled by restricted access. In vitro biochemistry shows that the FRB domain acts as a gatekeeper, with its rapamycin-binding site interacting with substrates to grant them access to the restricted active site. Rapamycin-FKBP12 inhibits the kinase by directly blocking substrate recruitment and by further restricting active-site access. The structures also reveal active-site residues and conformational changes that underlie inhibitor potency and specificity.

Mechanism of homologous recombination from the RecA-ssDNA/dsDNA structures.

The RecA family of ATPases mediates homologous recombination, a reaction essential for maintaining genomic integrity and for generating genetic diversity. RecA, ATP and single-stranded DNA (ssDNA) form a helical filament that binds to double-stranded DNA (dsDNA), searches for homology, and then catalyses the exchange of the complementary strand, producing a new heteroduplex. Here we have solved the crystal structures of the Escherichia coli RecA-ssDNA and RecA-heteroduplex filaments. They show that ssDNA and ATP bind to RecA-RecA interfaces cooperatively, explaining the ATP dependency of DNA binding. The ATP gamma-phosphate is sensed across the RecA-RecA interface by two lysine residues that also stimulate ATP hydrolysis, providing a mechanism for DNA release. The DNA is underwound and stretched globally, but locally it adopts a B-DNA-like conformation that restricts the homology search to Watson-Crick-type base pairing. The complementary strand interacts primarily through base pairing, making heteroduplex formation strictly dependent on complementarity. The underwound, stretched filament conformation probably evolved to destabilize the donor duplex, freeing the complementary strand for homology sampling.

Tel2 regulates the stability of PI3K-related protein kinases.

We report an unexpected role for Tel2 in the expression of all mammalian phosphatidylinositol 3-kinase-related protein kinases (PIKKs). Although Tel2 was identified as a budding yeast gene required for the telomere length maintenance, we found no obvious telomeric function for mammalian Tel2. Targeted gene deletion showed that mouse Tel2 is essential in embryonic development, embryonic stem (ES) cells, and embryonic fibroblasts. Conditional deletion of Tel2 from embryonic fibroblasts compromised their response to IR and UV, diminishing the activation of checkpoint kinases and their downstream effectors. The effects of Tel2 deletion correlated with significantly reduced protein levels for the PI3K-related kinases ataxia telangiectasia mutated (ATM), ATM and Rad3 related (ATR), DNA-dependent protein kinase catalytic subunit ataxia (DNA-PKcs). Tel2 deletion also elicited specific depletion of the mammalian target of rapamycin (mTOR), suppressor with morphological effect on genitalia 1 (SMG1), and transformation/transcription domain-associated protein (TRRAP), and curbed mTOR signaling, indicating that Tel2 affects all six mammalian PIKKs. While Tel2 deletion did not alter PIKK mRNA levels, in vivo pulse labeling experiments showed that Tel2 controls the stability of ATM and mTOR. Each of the PIKK family members associated with Tel2 in vivo and in vitro experiments indicated that Tel2 binds to part of the HEAT repeat segments of ATM and mTOR. These data identify Tel2 as a highly conserved regulator of PIKK stability.

Ubiquitination regulates PTEN nuclear import and tumor suppression.

The PTEN tumor suppressor is frequently affected in cancer cells, and inherited PTEN mutation causes cancer-susceptibility conditions such as Cowden syndrome. PTEN acts as a plasma-membrane lipid-phosphatase antagonizing the phosphoinositide 3-kinase/AKT cell survival pathway. However, PTEN is also found in cell nuclei, but mechanism, function, and relevance of nuclear localization remain unclear. We show that nuclear PTEN is essential for tumor suppression and that PTEN nuclear import is mediated by its monoubiquitination. A lysine mutant of PTEN, K289E associated with Cowden syndrome, retains catalytic activity but fails to accumulate in nuclei of patient tissue due to an import defect. We identify this and another lysine residue as major monoubiquitination sites essential for PTEN import. While nuclear PTEN is stable, polyubiquitination leads to its degradation in the cytoplasm. Thus, we identify cancer-associated mutations of PTEN that target its posttranslational modification and demonstrate how a discrete molecular mechanism dictates tumor progression by differentiating between degradation and protection of PTEN.

The BRCA2 homologue Brh2 nucleates RAD51 filament formation at a dsDNA-ssDNA junction.

The BRCA2 tumour suppressor is essential for the error-free repair of double-strand breaks (DSBs) in DNA by homologous recombination. This is mediated by RAD51, which forms a nucleoprotein filament with the 3' overhanging single-stranded DNA (ssDNA) of the resected DSB, searches for a homologous donor sequence, and catalyses strand exchange with the donor DNA. The 3,418-amino-acid BRCA2 contains eight approximately 30-amino-acid BRC repeats that bind RAD51 (refs 5, 6) and a approximately 700-amino-acid DBD domain that binds ssDNA. The isolated BRC and DBD domains have the opposing effects of inhibiting and stimulating recombination, respectively, and the role of BRCA2 in repair has been unclear. Here we show that a full-length BRCA2 homologue (Brh2) stimulates Rad51-mediated recombination at substoichiometric concentrations relative to Rad51. Brh2 recruits Rad51 to DNA and facilitates the nucleation of the filament, which is then elongated by the pool of free Rad51. Brh2 acts preferentially at a junction between double-stranded DNA (dsDNA) and ssDNA, with strict specificity for the 3' overhang polarity of a resected DSB. These results establish a BRCA2 function in RAD51-mediated DSB repair and explain the loss of this repair capacity in BRCA2-associated cancers.

The BRCA2-interacting protein DSS1 is vital for DNA repair, recombination, and genome stability in Ustilago maydis.

DSS1 encodes a small acidic protein shown in recent structural studies to interact with the DNA binding domain of BRCA2. Here we report that an ortholog of DSS1 is present in Ustilago maydis and associates with Brh2, the BRCA2-related protein, thus recapitulating the protein partnership in this genetically amenable fungus. Mutants of U. maydis deleted of DSS1 are extremely radiation sensitive, deficient in recombination, defective in meiosis, and disturbed in genome stability; these phenotypes mirror previous observations of U. maydis mutants deficient in Brh2 or Rad51. These findings conclusively show that Dss1 constitutes a protein with a significant role in the recombinational repair pathway in U. maydis, and imply that it plays a similar key role in the recombination systems of organisms in which recombinational repair is BRCA2 dependent.

BRCA2 function in DNA binding and recombination from a BRCA2-DSS1-ssDNA structure.

Mutations in the BRCA2 (breast cancer susceptibility gene 2) tumor suppressor lead to chromosomal instability due to defects in the repair of double-strand DNA breaks (DSBs) by homologous recombination, but BRCA2's role in this process has been unclear. Here, we present the 3.1 angstrom crystal structure of a approximately 90-kilodalton BRCA2 domain bound to DSS1, which reveals three oligonucleotide-binding (OB) folds and a helix-turn-helix (HTH) motif. We also (i) demonstrate that this BRCA2 domain binds single-stranded DNA, (ii) present its 3.5 angstrom structure bound to oligo(dT)9, (iii) provide data that implicate the HTH motif in dsDNA binding, and (iv) show that BRCA2 stimulates RAD51-mediated recombination in vitro. These findings establish that BRCA2 functions directly in homologous recombination and provide a structural and biochemical basis for understanding the loss of recombination-mediated DSB repair in BRCA2-associated cancers.