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Inhibition of host DNA damage response (DDR) is a common mechanism used by viruses to manipulate host cellular machinery and orchestrate viral life cycles. Epstein-Barr virus tegument protein BKRF4 associates with cellular chromatin to suppress host DDR signaling, but the underlying mechanism remains elusive. Here, we identify a BKRF4 histone binding domain (residues 15–102, termed BKRF4-HBD) that can accumulate at the DNA damage sites to disrupt 53BP1 foci formation. The high-resolution structure of the BKRF4-HBD in complex with a human H2A–H2B dimer shows that BKRF4-HBD interacts with the H2A–H2B dimer via the N-terminal region (NTR), the DWP motif (residues 80–86 containing D81, W84, P86), and the C-terminal region (CTR). The “triple-anchor” binding mode confers BKRF4-HBD the ability to associate with the partially unfolded nucleosomes, promoting the nucleosome disassembly. Importantly, disrupting the BKRF4–H2A–H2B interaction impairs the binding between BKRF4-HBD and nucleosome in vitro and inhibits the recruitment of BKRF4-HBD to DNA breaks in vivo. Together, our study reveals the structural basis of BKRF4 bindings to the partially unfolded nucleosome and elucidates an unconventional mechanism of host DDR signal attenuation.

Salmonella is considered one of the leading causes for foodborne diseases in humans. Pork and its products contaminated with Salmonella are increasingly recognized as an important source of human salmonellosis. The aim of this study was to investigate the antimicrobial resistance and prevalence of integrons in Salmonella isolates from pig farms. In total, 92 of 724 (12.7%) samples were Salmonella-positive, including 64 (15.0%) from fecal samples, 27 (12.6%) from floor samples, 1 (4.5%) from water samples, and 0 from feed and air samples. These isolates showed the highest resistance to tetracycline (85.9%), followed by trimethoprim (67.4%), ampicillin (60.9%), and chloramphenicol (51.1%). In addition, 51 isolates carried the complete class 1 integron, most of which (42/51) harbored antibiotic resistance cassettes. A total of six gene cassettes including orfF, est-X, dfrA1+aadA1, aadA1, dfrA12+aadA2, and sat were identified, in which the most prevalent one was orfF (29.4%). Furthermore, all 19 class 1 integron-positive isolates harboring dfr genes showed resistance to trimethoprim (SXT), suggesting that the trimethoprim resistance gene (dfr) may contribute to the emergence of SXT resistance phenotype. Therefore, considering the significance of integrons and related resistance genes for public health, special measures should be taken to control Salmonella spp. on the pig farms and to prevent spread of integrons and associated resistance genes.

Fanconi anemia (FA) is a rare genetic disorder characterized by bone marrow failure, developmental abnormalities and a predisposition to cancer, particularly acute myeloid leukemia. So far, 22 FA genes (FANCA-W) have been identified. Germline inactivation of any one of the 22 currently known FA genes causes FA. Proteins encoded by FA genes are involved in the Fanconi anemia pathway, which repairs DNA interstrand crosslinks (ICLs). The main function of the FA pathway in repairing ICLs has been extensively studied. In addition, several lines of evidence indicate that the FA pathway has crucial roles in genome maintenance upon replication stress, and is involved in common fragile sites, R-loops, and mitotic DNA synthesis. Recently, we found that all the functions of the FA pathway are possibly derived from one unified mechanism, namely that FA proteins play a role in the cleavage-coupled break-induced replication pathway to restart stalled replication forks. In the review, we summarize our current understanding of the functions of the FA pathway, and review our knowledge of the potential endogenous pathogenic factors that contribute to FA.

Fanconi anemia (FA) is a devastating hereditary disease characterized by bone marrow failure (BMF) and acute myeloid leukemia (AML). As FA-deficient cells are hypersensitive to DNA interstrand crosslinks (ICLs), ICLs are widely assumed to be the lesions responsible for FA symptoms. Here, we show that FA-mutated cells are hypersensitive to persistent replication stress and that FA proteins play a role in the break-induced-replication (BIR)-like pathway for fork restart. Both the BIR-like pathway and ICL repair share almost identical molecular mechanisms of 53BP1-BRCA1-controlled signaling response, SLX4- and FAN1-mediated fork cleavage and POLD3-dependent DNA synthesis, suggesting that the FA pathway is intrinsically one of the BIR-like pathways. Replication stress not only triggers BMF in FA-deficient mice, but also specifically induces monosomy 7, which is associated with progression to AML in patients with FA, in FA-deficient cells.

Topoisomerase 3β (TOP3B) and TDRD3 form a dual-activity topoisomerase complex that interacts with FMRP and can change the topology of both DNA and RNA. Here, we investigated the post-transcriptional influence of TOP3B and associated proteins on mRNA translation and turnover. First, we discovered that in human HCT116 colon cancer cells, knock-out (KO) of TOP3B had similar effects on mRNA turnover and translation as did TDRD3-KO, while FMRP-KO resulted in rather distinct effects, indicating that TOP3B had stronger coordination with TDRD3 than FMRP in mRNA regulation. Second, we identified TOP3B-bound mRNAs in HCT116 cells; we found that while TOP3B did not directly influence the stability or translation of most TOP3B target mRNAs, it stabilized a subset of target mRNAs but had a more complex effect on translation–enhancing for some mRNAs whereas reducing for others. Interestingly, a point mutation that specifically disrupted TOP3B catalytic activity only partially recapitulated the effects of TOP3B-KO on mRNA stability and translation, suggesting that the impact of TOP3B on target mRNAs is partly linked to its ability to change topology of mRNAs. Collectively, our data suggest that TOP3B–TDRD3 can regulate mRNA translation and turnover by mechanisms that are dependent and independent of topoisomerase activity.

DNA double strand break (DSB) is a kind of highly toxic damage, causing genomic instability, chromatin aberration and cancer. A number of conserved DSB damage repair pathways have been found in eukaryotes, most notably classical non-homologous end joining (cNHEJ) and homologous recombination (HR). These two repair pathways resolve most DSBs in cells. For different types of DSBs, cells could wittily choose the optimal approach with the lowest risk for genomic instability. Relying on the developing cell imaging techniques and molecular biological methods, sophisticated mechanism of DSB repair pathway choice has been revealed. The regulation mechanism of pathway choice is complex and precise. In this review, the effect factors such as the structures of the broken terminals, the degree of DNA end resection, cell cycle, chromatin environment, histone modification and RNA metabolism on the DSB repair pathway choice are discussed, showing the diversity and flexibility of DSB repair pathway choice.

Introduction Integrons are mobile DNA elements that allow for acquisition and dissemination of antibiotic-resistance genes among pig farm-derived bacteria. Limited information is available on integrons of Staphylococcus aureus from pig farms. The aim of this study was to characterise and investigate the prevalence of class 1 and 2 integrons in multi-drug resistant (MDR) S. aureus isolates from pig farms. Material and Methods A total of 724 swabs were collected from 12 pig farms in Chongqing, China, and examined by conventional microbial and molecular methods. Results In total, 68 isolates were S. aureus, 57 of which were methicillin resistant (MRSA). All 68 isolates were MDR strains and carried integrons, of which 88.2% (60/68) harboured both class 1 and 2. In addition, 85.3% (58/68) of the class 2 integron-positive isolates carried the β-lactam resistance gene (blaTEM-1 ), and 66.7% (40/60) of the class 1 integron–positive isolates carried the aadA1c, aadA1 or dfrA1 gene for respective streptomycin and spectinomycin or trimethoprim resistance. Conclusions Class 1 and 2 integrons are common among the pig farm-derived S. aureus isolates. On account of their significance for public health, the prevalence of the integrons and their associated resistance genes in pig farm-derived S. aureus isolates should be paid special attention.

Deoxyribonucleic acid (DNA) is at a constant risk of damage from endogenous substances, environmental radiation, and chemical stressors. DNA double-strand breaks (DSBs) pose a significant threat to genomic integrity and cell survival. There are two major pathways for DSB repair: nonhomologous end-joining (NHEJ) and homologous recombination (HR). The extent of DNA end resection, which determines the length of the 3′ single-stranded DNA (ssDNA) overhang, is the primary factor that determines whether repair is carried out via NHEJ or HR. NHEJ, which does not require a 3′ ssDNA tail, occurs throughout the cell cycle. 53BP1 and the cofactors PTIP or RIF1-shieldin protect the broken DNA end, inhibit long-range end resection and thus promote NHEJ. In contrast, HR mainly occurs during the S/G2 phase and requires DNA end processing to create a 3′ tail that can invade a homologous region, ensuring faithful gene repair. BRCA1 and the cofactors CtIP, EXO1, BLM/DNA2, and the MRE11–RAD50–NBS1 (MRN) complex promote DNA end resection and thus HR. DNA resection is influenced by the cell cycle, the chromatin environment, and the complexity of the DNA end break. Herein, we summarize the key factors involved in repair pathway selection for DSBs and discuss recent related publications.

Topoisomerase 3b (TOP3B)-TDRD3 is a dual-activity topoisomerase complex in animals that can change topology for both DNA and RNA. A current hypothesis proposes that this complex interacts with Fragile X mental retardation protein, FMRP, to regulate mRNA translation. Here we examined this hypothesis by identifying TOP3B-TDRD3-bound mRNAs in human HCT116 cells; and analyzing the effect of inactivating TOP3B, TOP3B catalytic activity, TDRD3, and FMRP, on translation and abundance of these mRNAs. We found that TOP3B-TDRD3 resembles FMRP in preferentially binding to coding regions of long mRNAs. Complete inactivation of TOP3B protein, but not a point mutation that only disrupts TOP3B catalytic activity, preferentially reduced the abundance of the TOP3B-TDRD3-bound mRNAs. Moreover, ablation of the complex or inactivation of TOP3B catalytic activity alters translation for some but not all TOP3B-TDRD3 bound mRNAs, suggesting that TOP3B-TDRD3 may solve topological problems for specific mRNAs. Finally, several schizophrenia and autism-risk genes are bound and regulated by TOP3B-TDRD3 in mRNA translation and abundance by topoisomerase activity dependent and independent manners. Our data suggest that TOP3B-TDRD3 can regulate mRNA translation and abundance by topoisomerase activity dependent or independent mechanisms; and disruption of this function could contribute to pathogenesis of psychiatric disorders in TOP3B mutation carriers.

Topoisomerases solve topological problems for DNA, but whether they function similarly for RNA remains unclear. Here we show that the Topoisomerase 3b (TOP3B)-TDRD3 complex acts on both DNA and RNA. On DNA, TOP3B-TDRD3 binds transcription start or termination regions; and inactivation of TOP3B topoisomerase activity decreases expression of some though not all genes downregulated in TOP3B-KO and TDRD3-KO cells. On mRNA, TOP3B-TDRD3 preferentially binds coding regions of long mRNAs and stabilizes them independently of TOP3B topoisomerase activity. Moreover, ablation of the complex or inactivation of TOP3B catalytic activity alters translation for some mRNAs, suggesting that TOP3B-TDRD3 may solve RNA topological problems. Finally, several schizophrenia and autism-risk genes are bound and regulated by TOP3B-TDRD3 at either transcription or translation. Our data suggest that TOP3B-TDRD3 can regulate transcription and translation by topoisomerase activity dependent or independent mechanisms; and disruption of either function could contribute to pathogenesis of psychiatric disorders in TOP3B mutation carriers.

Abiotic stresses, such as heat, salt, waterlogging, and multiple-stress environments have significantly reduced wheat production in recent decades. There is a need to use effective strategies for overcoming crop losses due to these abiotic stresses. Fertilizer-based approaches are readily available and can be managed in all farming communities. This research revealed the effects of sulfur-coated urea (SCU, 130 kg ha−1, release time of 120 days) on wheat crops under heat, salt, waterlogging, and combined-stress climatic conditions. The research was done using a completely randomized design with three replicates. The results revealed that SCU at a rate of 130 kg of N ha−1 showed a significantly (p ≤ 0.05) high SPAD value (55) in the case of waterlogging stress, while it was the lowest (31) in the case of heat stress, the control had a SPAD value of 58. Stress application significantly (p ≤ 0.05) reduced the leaf area and was the highest in control (1898 cm2), followed by salt stress (1509 cm2), waterlogging (1478 cm2), and heat stress (1298 cm2). A significantly (p ≤ 0.05) lowest crop yield was observed in the case of heat stress (3623.47 kg ha−1) among all stresses, while it was 10,270 kg ha−1 in control and was reduced up to 35% after the application of heat stress. Among all stresses, the salt stress showed the highest crop yield of 5473.16 kg ha−1. A significant correlation was observed among growth rate, spike length, yield, and physiological constraints with N content in the soil. The SCU fertilizer was the least effective against heat stress but could tolerate salt stress in wheat plants. The findings suggested the feasibility of adding SCU as an alternative to normal urea to alleviate salt stresses and improve wheat crop growth and yield traits. For heat stress tolerance, the applicability of SCU with a longer release period of ~180 days is recommended as a future prospect for study.

The 53BP1-RIF1 pathway antagonizes resection of DNA broken ends and confers PARP inhibitor sensitivity on BRCA1-mutated tumors. However, it is unclear how this pathway suppresses initiation of resection. Here, we identify ASF1 as a partner of RIF1 via an interacting manner similar to its interactions with histone chaperones CAF-1 and HIRA. ASF1 is recruited to distal chromatin flanking DNA breaks by 53BP1-RIF1 and promotes non-homologous end joining (NHEJ) using its histone chaperone activity. Epistasis analysis shows that ASF1 acts in the same NHEJ pathway as RIF1, but via a parallel pathway with the shieldin complex, which suppresses resection after initiation. Moreover, defects in end resection and homologous recombination (HR) in BRCA1- deficient cells are largely suppressed by ASF1 deficiency. Mechanistically, ASF1 compacts adjacent chromatin by heterochromatinization to protect broken DNA ends from BRCA1-mediated resection. Taken together, our findings identified a RIF1-ASF1 histone chaperone complex that promotes changes in high-order chromatin structure to stimulate the NHEJ pathway for DSB repair.

Pasteurella multocida (P. multocida) can cause severe respiratory disease in cattle, resulting in high mortality and morbidity. Inflammasomes are multiprotein complexes in the cytoplasm that recognize pathogens and play an important role in the host defense against microbial infection. In this study, the mechanism of P. multocida-induced NLRP6 inflammasome activation was investigated in vitro and in vivo. Firstly, P. multocida induced severe inflammation with a large number of inflammatory cells infiltrating the lungs of WT and Nlrp6−/− mice. Nlrp6−/− mice were more susceptible to P. multocida infection and they had more bacterial burden in the lungs. Then, the recruitment of macrophages and neutrophils in the lungs was investigated and the results show that the number of immune cells was significantly decreased in Nlrp6−/− mice. Subsequently, NLRP6 was shown to regulate P. multocida-induced inflammatory cytokine secretion including IL-1β and IL-6 both in vivo and in vitro while TNF-α secretion was not altered. Moreover, NLRP6 was found to mediate caspase-1 activation and ASC oligomerization, resulting in IL-1β secretion. Furthermore, NLRP6 inflammasome mediated the gene expression of chemokines including CXCL1, CXCL2 and CXCR2 which drive the activation of NLRP3 inflammasomes. Finally, NLRP3 protein expression was detected to be abrogated in P. multocida-infected Nlrp6−/− macrophages, indicating the synergic effect of NLRP6 and NLRP3. Our study demonstrates that NLRP6 inflammasome plays an important role in the host against P. multocida infection and contributes to the development of immune therapeutics against P. multocida.

Background Salmonella is one of the most important foodborne pathogens, causing outbreaks of human salmonellosis worldwide. Owing to large scales of consumption markets, pork and poultry that contaminated by Salmonella could pose a tremendous threat to public health. The aim of this study was to investigate the contamination of Salmonella from chicken, pork and the environment in slaughtering and retail processes in Chongqing, China. Results A total of 115 Salmonella isolates were recovered from 1112 samples collected from pork, chicken and the environment. Compared with the isolation rate of samples from chicken (9.50%) and the environment (6.23%), samples from pork had a significant higher isolation rate (44.00%). The isolation rates in slaughterhouses (10.76%) and in supermarkets (10.07%) showed no statistical difference. Thirty different serotypes were identified among all the isolates. S. Derby (n = 26), S. London (n = 16) and S. Rissen (n = 12) were the dominant serotypes. Antimicrobial susceptibility testing revealed that 73.04% isolates were resistant to tetracycline, followed by 66.96% to ampicillin and 59.13% to doxycycline. More than half (50.43%) of the isolates were multidrug resistant (MDR), and most of the MDR isolates were from supermarkets. Multilocus sequence typing results showed 24 out of 115 isolates were ST40, which was the most prevalent. Furthermore, isolates from supermarkets had 20 different sequence types while isolates from slaughterhouses only had 8 different sequence types. Conclusion Our study highlighted that Salmonella was more frequently isolated in pork production chain than that in chicken. Compared with isolates from slaughterhouses, isolates from supermarkets had more MDR profiles and represented a wider range of serotypes and sequence types, indicating that the retail process had more diverse sources of Salmonella contamination than that of slaughtering process.

Chromosome translocation is a major cause of the onset and progression of diverse types of cancers. However, the mechanisms underlying this process remain poorly understood. Here, we identified a non-homologous end-joining protein, IFFO1, which structurally forms a heterotetramer with XRCC4. IFFO1 is recruited to the sites of DNA damage by XRCC4 and promotes the repair of DNA double-strand breaks in a parallel pathway with XLF. Interestingly, IFFO1 interacts with lamin A/C, forming an interior nucleoskeleton. Inactivating IFFO1 or its interaction with XRCC4 or lamin A/C leads to increases in both the mobility of broken ends and the frequency of chromosome translocation. Importantly, the destruction of this nucleoskeleton accounts for the elevated frequency of chromosome translocation in many types of cancer cells. Our results reveal that the lamin A/C-IFFO1-constituted nucleoskeleton prevents chromosome translocation by immobilizing broken DNA ends during tumorigenesis.

Enzootic nasal adenocarcinoma (ENA) of goats, characterized by transformation of epithelial cells of the ethmoid turbinates, is caused by enzootic nasal tumor virus 2 (ENTV-2). ENTV-2 belongs to the genus Betaretrovirus and has extended its distribution globally with a high prevalence; however, the genetic diversity and genotypic distribution for ENTV-2 have not been analyzed systematically due to the limited availability of sequence data. In this study, an infection by ENTV-2 was detected by RT-PCR in Chongqing in July 2018, and the complete sequence of one strain (CQ1) was determined. Phylogenetic analysis indicated a high degree of genetic heterogeneity among ENTV-2 sequences, with the existence of two main lineages. Lineage 1 and 2 were composed of ENTV-2 from China and the UK, respectively. Although CQ1 was closely related to recent ENTV-2 strains collected in the neighboring provinces of Chongqing (Shaanxi and Sichuan), it formed a separate sublineage of lineage 1 (sublineage 1.3). This report will enhance our understanding of the epidemiology of ENTV-2 in China.

Streptococcus pneumoniae (S. pneumoniae) causes severe pulmonary diseases, leading to high morbidity and mortality. It has been reported that inflammasomes such as NLR family pyrin domain containing 3 (NLRP3) and absent in melanoma 2 (AIM2) play an important role in the host defense against S. pneumoniae infection. However, the role of NLRP6 in vivo and in vitro against S. pneumoniae remains unclear. Therefore, we investigated the role of NLRP6 in regulating the S. pneumoniae-induced inflammatory signaling pathway in vitro and the role of NLRP6 in the host defense against S. pneumoniae in vivo by using NLRP6−/− mice. The results showed that the NLRP6 inflammasome regulated the maturation and secretion of IL-1β, but it did not affect the induction of IL-1β transcription in S. pneumoniae-infected macrophages. Furthermore, the activation of caspase-1, caspase-11, and gasdermin D (GSDMD) as well as the oligomerization of apoptosis-associated speck-like protein (ASC) were also mediated by NLRP6 in S. pneumoniae-infected macrophages. However, the activation of NLRP6 reduced the expression of NF-κB and ERK signaling pathways in S. pneumoniae-infected macrophages. In vivo study showed that NLRP6−/− mice had a higher survival rate, lower number of bacteria, and milder inflammatory response in the lung compared with wild-type (WT) mice during S. pneumoniae infection, indicating that NLRP6 plays a negative role in the host defense against S. pneumoniae. Furthermore, increased bacterial clearance in NLRP6 deficient mice was modulated by the recruitment of macrophages and neutrophils. Our study provides a new insight on S. pneumoniae-induced activation of NLRP6 and suggests that blocking NLRP6 could be considered as a potential therapeutic strategy to treat S. pneumoniae infection.

Pseudorabies virus (PRV), an alphaherpesvirus, causes respiratory and reproductive diseases in pigs and severe nervous symptom in other susceptible hosts. Previous studies showed that PRV infection induced a systemic inflammatory response in mice, indicating that pro-inflammatory cytokines participated in viral neuropathy in mice. The pro-inflammatory cytokine IL-1β is a key mediator of the inflammatory response and plays an important role in host-response to pathogens. However, the secretion of IL-1β and its relationship with inflammasome activation during PRV infection remains poorly understood. In this study, we found that PRV infection caused significant secretion of several pro-inflammatory cytokines in macrophages and promoted IL-1β secretion in an ATP-dependent manner. Furthermore, the expression of IL-1β can be induced by only PRV infection and depended on NF-κB pathway activation, while the subsequent secretion of IL-1β was mediated by ATP-induced P2 × 7R activation, loss of intracellular K+, and the subsequent NLRP3 inflammasome activation. By using a mouse infection model, we also found that ATP exacerbated clinical signs and death of mice infected by PRV in a NLRP3-dependent manner. These results indicate that ATP facilitates activation of NLRP3 inflammasome and enhances the pathogenicity of PRV in mice during its acute infection.

Staphylococcus aureus is a Gram-positive opportunistic pathogen which causes infections in a variety of vertebrates. Virulence factors are the main pathogenesis of S. aureus as a pathogen, which induce the host&rsquo, s innate and adaptive immune responses. Toxic shock syndrome toxin 1 (TSST-1) is one of the most important virulence factors of S. aureus. However, the role of nucleotide-binding oligomerization domain-like receptor family pyrin domain containing 3 (NLRP3) in TSST-1-induced innate immune response is still unclear. Here, purified recombinant TSST-1 (rTSST-1) was prepared and used to stimulate mouse peritoneal macrophages. The results showed that under the action of adenosine-triphosphate (ATP), rTSST-1 significantly induced interleukin-1&beta, (IL-1&beta, ) and tumor necrosis factor-&alpha, (TNF-&alpha, ) production in mouse macrophages and the production was dose-dependent. In addition, rTSST-1+ATP-stimulated cytokine production in macrophage depends on the activation of toll like receptor 4 (TLR4), but not TLR2 on the cells. Furthermore, the macrophages of NLRP3&minus, /&minus, mice stimulated with rTSST-1+ATP showed significantly low levels of IL-1&beta, production compared to that of wild-type mice. These results demonstrated that TSST-1 can induce the expression of inflammatory cytokines in macrophages via the activation of the TLR4 and NLRP3 signaling pathways. Our study provides new information about the mechanism of the TSST-1-inducing host&rsquo, s innate immune responses.

Fanconi anemia (FA) is a devastating hereditary disease characterized by bone marrow failure (BMF) and acute myeloid leukemia (AML). As FA-deficient cells are hypersensitive to DNA interstrand crosslinks (ICLs), ICLs are widely assumed to be the lesions responsible for FA symptoms. Here, we show that FA-mutated cells are hypersensitive to persistent replication stress and that FA proteins play a role in the break-induced-replication (BIR)-like pathway for fork restart. Both the BIR-like pathway and ICL repair share almost identical molecular mechanisms of 53BP1-BRCA1-controlled signaling response, SLX4- and FAN1-mediated fork cleavage and POLD3-dependent DNA synthesis, suggesting that the FA pathway is intrinsically one of the BIR-like pathways. Replication stress not only triggers BMF in FA-deficient mice, but also specifically induces monosomy 7, which is associated with progression to AML in patients with FA, in FA-deficient cells.

Background: Salmonella is one of the most important foodborne pathogens, causing outbreaks of human salmonellosis worldwide. Owing to large scales of consumption markets, pork and poultry that contaminated by Salmonella could pose a tremendous threat to public health. The aim of this study was to investigate the contamination of Salmonella from chicken, pork and the environment in slaughtering and retail processes in Chongqing, China. Results: A total of 115 Salmonella isolates were recovered from 1112 samples collected from pork, chicken and the environment. Compared with the isolation rate of samples from chicken (9.50%) and the environment (6.23%), samples from pork had a significant higher isolation rate (44.00%). The isolation rates in slaughterhouses (10.76%) and in supermarkets (10.07%) showed no statistical different. 30 different serotypes were identified among all the isolates. S. Derby (n=26), S. London (n=16) and S. Rissen (n=12) were the dominant serotypes. Antimicrobial susceptibility testing revealed that 73.04% isolates were resistant to tetracycline, followed by 66.96% to ampicillin and 59.13% to doxycycline. More than half (50.43%) of the isolates were multidrug resistant (MDR), and most of the MDR isolates were from supermarkets. Multilocus sequence typing results showed 24 out of 115 isolates were ST40, which was the most prevalent. Furthermore, isolates from supermarkets had 20 different sequence types while isolates from slaughterhouses only had 8 different sequence types. Conclusion: Our study highlighted that Salmonella was more frequently isolated in pork production chain than that of chicken. Compared with isolates from slaughterhouses, isolates from supermarkets had more MDR profiles and represented a wider range of serotypes and sequence types, indicating that the retail process had more diverse sources of Salmonella contamination than that of slaughtering process.

Background: Salmonella is one of the most important foodborne pathogens, causing outbreaks of human salmonellosis worldwide. Owing to large scales of consumption markets, pork and poultry that contaminated by Salmonella could pose a tremendous threat to public health. The aim of this study was to investigate the contamination of Salmonella from chicken, pork and the environment in slaughtering and retailing segments in Chongqing, China. Results: A total of 115 Salmonella isolates were recovered from 1122 samples, with 10.76% (47/437) in slaughterhouses and 10.07% (68/675) in supermarkets. 30 different serotypes were identified, in which S. Derby, S. London and S. Rissen were mostly detected. Antimicrobial susceptibility testing and resistance genes verification were carried out to investigate the relationship between phenotypes and genotypes. Altogether, 75.65% isolates showed resistance to tetracycline, followed by 60.87% to doxycycline and 69.5% to ampicillin. More than half (50.43%) of the isolates were multidrug resistant, which were mostly from supermarkets (P

SMC5/6 function in genome integrity remains elusive. Here, we show that SMC5 dysfunction in avian DT40 B cells causes mitotic delay and hypersensitivity toward DNA intra‐ and inter‐strand crosslinkers (ICLs), with smc5 mutants being epistatic to FANCC and FANCM mutations affecting the Fanconi anemia (FA) pathway. Mutations in the checkpoint clamp loader RAD17 and the DNA helicase DDX11, acting in an FA‐like pathway, do not aggravate the damage sensitivity caused by SMC5 dysfunction in DT40 cells. SMC5/6 knockdown in HeLa cells causes MMC sensitivity, increases nuclear bridges, micronuclei, and mitotic catastrophes in a manner similar and non‐additive to FANCD2 knockdown. In both DT40 and HeLa systems, SMC5/6 deficiency does not affect FANCD2 ubiquitylation and, unlike FANCD2 depletion, RAD51 focus formation. SMC5/6 components further physically interact with FANCD2‐I in human cells. Altogether, our data suggest that SMC5/6 functions jointly with the FA pathway to support genome integrity and DNA repair and may be implicated in FA or FA‐related human disorders., The structural maintenance complex SMC5/6 interacts with and functions jointly with key components of the Fanconi anemia pathway to facilitate DNA repair and preserve genome stability.

Human cells contain five topoisomerases in the nucleus and cytoplasm, but which one is the major topoisomerase for mRNAs is unclear. To date, Top3β is the only known topoisomerase that possesses RNA topoisomerase activity, binds mRNA translation machinery and interacts with an RNA-binding protein, FMRP, to promote synapse formation; and Top3β gene deletion has been linked to schizophrenia. Here, we show that Top3β is also the most abundant mRNA-binding topoisomerase in cells. Top3β, but not other topoisomerases, contains a distinctive RNA-binding domain; and deletion of this domain diminishes the amount of Top3β that associates with mRNAs, indicating that Top3β is specifically targeted to mRNAs by its RNA binding domain. Moreover, Top3β mutants lacking either its RNA-binding domain or catalytic residue fail to promote synapse formation, suggesting that Top3β requires both its mRNA-binding and catalytic activity to facilitate neurodevelopment. Notably, Top3β proteins bearing point mutations from schizophrenia and autism individuals are defective in association with FMRP; whereas one of the mutants is also deficient in binding mRNAs, catalyzing RNA topoisomerase reaction, and promoting synapse formation. Our data suggest that Top3β is the major topoisomerase for mRNAs, and requires both RNA binding and catalytic activity to promote neurodevelopment and prevent mental dysfunction.

The MRE11–RAD50–NBS1 (MRN) complex is well known for participating in DNA damage response pathways in all phases of cell cycle. Here, we show that MRN constitutes a mitosis-specific complex, named mMRN, with a protein, MMAP. MMAP directly interacts with MRE11 and is required for optimal stability of the MRN complex during mitosis. MMAP colocalizes with MRN in mitotic spindles, and MMAP-deficient cells display abnormal spindle dynamics and chromosome segregation similar to MRN-deficient cells. Mechanistically, both MMAP and MRE11 are hyperphosphorylated by the mitotic kinase, PLK1; and the phosphorylation is required for assembly of the mMRN complex. The assembled mMRN complex enables PLK1 to interact with and activate the microtubule depolymerase, KIF2A, leading to spindle turnover and chromosome segregation. Our study identifies a mitosis-specific version of the MRN complex that acts in the PLK1–KIF2A signaling cascade to regulate spindle dynamics and chromosome distribution.

Generation of single‐stranded DNA (ssDNA) is required for the template strand formation during DNA replication. Replication Protein A (RPA) is an ssDNA‐binding protein essential for protecting ssDNA at replication forks in eukaryotic cells. While significant progress has been made in characterizing the role of the RPA–ssDNA complex, how RPA is loaded at replication forks remains poorly explored. Here, we show that the Saccharomyces cerevisiae protein regulator of Ty1 transposition 105 (Rtt105) binds RPA and helps load it at replication forks. Cells lacking Rtt105 exhibit a dramatic reduction in RPA loading at replication forks, compromised DNA synthesis under replication stress, and increased genome instability. Mechanistically, we show that Rtt105 mediates the RPA–importin interaction and also promotes RPA binding to ssDNA directly in vitro, but is not present in the final RPA–ssDNA complex. Single‐molecule studies reveal that Rtt105 affects the binding mode of RPA to ssDNA. These results support a model in which Rtt105 functions as an RPA chaperone that escorts RPA to the nucleus and facilitates its loading onto ssDNA at replication forks.

53BP1 with its downstream proteins, RIF1, PTIP and REV7, antagonizes BRCA1-dependent homologous recombination (HR) and promotes non-homologous end joining (NHEJ) in an unclear manner. Here we show that REV7 forms a complex with two proteins, FAM35A and C20ORF196. We demonstrate that FAM35A preferentially binds single-strand DNA (ssDNA) in vitro, and is recruited to DSBs as a complex with C20ORF196 and REV7 downstream of RIF1 in vivo. Epistasis analysis shows that both proteins act in the same pathway as RIF1 in NHEJ. The defects in HR pathway to repair DSBs and the reduction in resection of broken DNA ends in BRCA1-mutant cells can be largely suppressed by inactivating FAM35A or C20ORF196, indicating that FAM35A and C20ORF196 prevent end resection in these cells. Together, our data identified a REV7–FAM35A–C20ORF196 complex that binds and protects broken DNA ends to promote the NHEJ pathway for DSB repair.

RNA topoisomerase activity has recently been detected in multiple Type IA DNA topoisomerases from all three domains of life: bacteria, archaea, and eukarya. Many, but not all, Type IA topoisomerases are found to possess activities for not only DNA, but also RNA, suggesting that they may solve topological problems for both types of nucleic acids. Here we describe a detailed assay used by our group to detect RNA topoisomerase activity for many Type IA topoisomerases. We discuss the strategy, experimental procedures, troubleshooting, and limitations for this assay.

The cellular pathways that restart stalled replication forks are essential for genome stability and tumor prevention. However, how many of these pathways exist in cells and how these pathways are selectively activated remain unclear. Here, we describe two major fork restart pathways, and demonstrate that their selection is governed by 53BP1 and BRCA1, which are known to control the pathway choice to repair double-strand DNA breaks (DSBs). Specifically, 53BP1 promotes a fork cleavage-free pathway, whereas BRCA1 facilitates a break-induced replication (BIR) pathway coupled with SLX-MUS complex-mediated fork cleavage. The defect in the first pathway, but not DSB repair, in a 53BP1 mutant is largely corrected by disrupting BRCA1, and vice versa. Moreover, PLK1 temporally regulates the switch of these two pathways through enhancing the assembly of the SLX-MUS complex. Our results reveal two distinct fork restart pathways, which are antagonistically controlled by 53BP1 and BRCA1 in a DSB repair-independent manner. Copyright Xu et al. This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited.

Topoisomerases solve critical topological problems in DNA metabolism and have long been regarded as the "magicians" of the DNA world. Here we present views from 2 of our recent studies indicating that Type IA topoisomerases from all domains of life often possess dual topoisomerase activities for both DNA and RNA. In animals, one of the 2 Type IA topoisomerases, Top3β, contains an RNA-binding domain, possesses RNA topoisomerase activity, binds mRNAs, interacts with mRNA-binding proteins, and associates with active mRNA translation machinery. The RNA-binding domain is required for Top3β to bind mRNAs and promote normal neurodevelopment. Top3β forms a highly conserved complex with Tudor-domain-containing 3 (TDRD3), a protein known to interact with translation factors, histones, RNA polymerase II, single stranded DNA and RNA. Top3β requires TDRD3 for its association with the mRNA translation machinery. We suggest that Type IA topoisomerases can be "magicians" for not only DNA, but also RNA; and they may solve topological problems for both nucleic acids in all domains of life. In animals, Top3β-TDRD3 is a dual-activity topoisomerase complex that can act on DNA to stimulate transcription, and on mRNA to promote translation.

Summary: Eukaryotic replisomes are driven by the mini chromosome maintenance (MCM [M]) helicase complex, an offset ring locked around the template for leading strand synthesis by CDC45 (C) and GINS (G) proteins. Although the CDC45 MCM GINS (CMG) structure implies that interstrand crosslinks (ICLs) are absolute blocks to replisomes, recent studies indicate that cells can restart DNA synthesis on the side of the ICL distal to the initial encounter. Here, we report that restart requires ATR and is promoted by FANCD2 and phosphorylated FANCM. Following introduction of genomic ICLs and dependent on ATR and FANCD2 but not on the Fanconi anemia core proteins or FAAP24, FANCM binds the replisome complex, with concomitant release of the GINS proteins. In situ analysis of replisomes proximal to ICLs confirms the ATR-dependent release of GINS proteins while CDC45 is retained on the remodeled replisome. The results demonstrate the plasticity of CMG composition in response to replication stress. : Replication of the mammalian genome is driven by the replisome complex, which unwinds DNA and must overcome many impediments. Zhang et al. find that the encounter of the CMG with a strong block triggers a change in replisome composition that is important for restart of replication past the obstruction. Keywords: replication traverse, interstrand crosslink, ICL, ATR, FANCM, FANCD2, CMG, GINS