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Abstract Background Homology-based recombination (HR) is the cornerstone of genetic mapping. However, a lack of sufficient sequence homology or the presence of a genomic rearrangement prevents HR through crossing, which inhibits genetic mapping in relevant genomic regions. This is particularly true in species hybrids whose genomic sequences are highly divergent along with various genome arrangements, making the mapping of genetic loci, such as hybrid incompatibility (HI) loci, through crossing impractical. We previously mapped tens of HI loci between two nematodes, Caenorhabditis briggsae and C. nigoni, through the repeated backcrossing of GFP-linked C. briggsae fragments into C. nigoni. However, the median introgression size was over 7 Mb, indicating apparent HR suppression and preventing the subsequent cloning of the causative gene underlying a given HI phenotype. Therefore, a robust method that permits recombination independent of sequence homology is desperately desired. Results Here, we report a method of highly efficient targeted recombination (TR) induced by CRISPR/Cas9 with dual guide RNAs (gRNAs), which circumvents the HR suppression in hybrids between the two species. We demonstrated that a single gRNA was able to induce efficient TR between highly homologous sequences only in the F1 hybrids but not in the hybrids that carry a GFP-linked C. briggsae fragment in an otherwise C. nigoni background. We achieved highly efficient TR, regardless of sequence homology or genetic background, when dual gRNAs were used that each specifically targeted one parental chromosome. We further showed that dual gRNAs were able to induce efficient TR within genomic regions that had undergone inversion, in which HR-based recombination was expected to be suppressed, supporting the idea that dual-gRNA-induced TR can be achieved through nonhomology-based end joining between two parental chromosomes. Conclusions Recombination suppression can be circumvented through CRISPR/Cas9 with dual gRNAs, regardless of sequence homology or the genetic background of the species hybrid. This method is expected to be applicable to other situations in which recombination is suppressed in interspecies or intrapopulation hybrids.

Background: PIWI-interacting RNAs (piRNAs) are a class of noncoding RNAs originally reported in the reproductive system of mammals and later found to be aberrantly expressed in tumors. However, the function and mechanism of piRNAs in testicular cancer are not very clear. Methods: The expression level and distribution of piR-36249 were detected by RT-qPCR and immunofluorescence staining assay. Testicular cancer cell (NT2) progression was measured by CCK8 assay, colony formation assay and wound healing assay. Cell apoptosis was assessed by flow cytometry and western blot. RNA sequencing and dual-luciferase reporter assay were conducted to identify the potential targets of piR-36249. The relationship between piR-36249 and OAS2 or DHX36 was confirmed using overexpression assay, knockdown assay, pull-down assay and RIP assay. Results: piR-36249 is significantly downregulated in testicular cancer tissues compared to tumor-adjacent tissues. Functional studies demonstrate that piR-36249 inhibits testicular cancer cell proliferation, migration and activates the cell apoptosis pathway. Mechanically, we identify that piR-36249 binds to the 3′UTR of 2′-5′-oligoadenylate synthetase 2 (OAS2) mRNA. OAS2 has been shown in the literature to be a tumor suppressor modulating the occurrence and development of some tumors. Here, we show that OAS2 knockdown also promotes testicular cancer cell proliferation and migration. Furthermore, piR-36249 interacts with DHX36, which has been reported to promote translation. DHX36 can also bind to OAS2 mRNA, and knockdown of DHX36 increases OAS2 mRNA but downregulates its protein, indicating the enhancing effect of DHX36 on OAS2 protein expression. Conclusion: All these data suggest that piR-36249, together with DHX36, functions in inhibiting the malignant phenotype of testicular cancer cells by upregulating OAS2 protein and that piR-36249 may be used as a suppressor factor to regulate the development of testicular cancer.

Summary: Mechanical allodynia and hyperalgesia are intractable symptoms lacking effective clinical treatments in patients with neuropathic pain. However, whether and how mechanically responsive non-peptidergic nociceptors are involved remains elusive. Here, we showed that von Frey-evoked static allodynia and aversion, along with mechanical hyperalgesia after spared nerve injury (SNI) were reduced by ablation of MrgprdCreERT2-marked neurons. Electrophysiological recordings revealed that SNI-opened Aβ-fiber inputs to laminae I-IIo and vIIi, as well as C-fiber inputs to vIIi, were all attenuated in Mrgprd-ablated mice. In addition, priming chemogenetic or optogenetic activation of Mrgprd+ neurons drove mechanical allodynia and aversion to low-threshold mechanical stimuli, along with mechanical hyperalgesia. Mechanistically, gated Aβ and C inputs to vIIi were opened, potentially via central sensitization by dampening potassium currents. Altogether, we uncovered the involvement of Mrgprd+ nociceptors in nerve injury-induced mechanical pain and dissected the underlying spinal mechanisms, thus providing insights into potential therapeutic targets for pain management.

Summary: The function of the cancer-associated lncRNA Malat1 during aging is as-of-yet uncharacterized. Here, we show that Malat1 interacts with Nucleophosmin (NPM) in young mouse brain, and with Lamin A/C, hnRNP C, and KAP1 with age. RNA-seq and RT-qPCR reveal a persistent expression of Malat1_2 (the 3’isoform of Malat1) in Malat1Δ1 (5’-1.5 kb deletion) mouse retinas and brains at 1/4th level of the full-length Malat1, while Malat1_1 (the 5’isoform) in Malat1Δ2 (deletion of 3’-conserved 5.7 kb) at a much lower level, suggesting an internal promoter driving the 3’ isoform. The 1774 and 496 differentially expressed genes in Malat1Δ2 and Malat1Δ1 brains, respectively, suggest the 3’ isoform regulates gene expression in trans and the 5’ isoform in cis. Consistently, Malat1Δ2 mice show increased age-dependent retinal oxidative stress and corneal opacity, while Malat1Δ1 mice show no obvious phenotype. Collectively, this study reveals a physiological function of the lncRNA Malat1 3’-isoform during the aging process.

Collaborative filtering is the most widely used method in recommendation algorithms, but it still faces the serious problem of data sparsity. Traditional collaborative filtering uses matrix decomposition to learn the latent features of users and items. As an extension model of matrix decomposition, Funk-SVD model has attracted wide attention due to its good scalability and easy implementation, but it is difficult to extract the latent features of users and items from sparse rating information because it essentially learns the linear relationship between users and items. To solve this problem, we propose a Dual auto-encoder based Rating Prediction Recommendation Algorithm (DRPRA) model. The DRPRA model uses the strong ability of deep learning in feature learning, which combines double auto-encoders with Funk-SVD. First, the auto-encoder captures the latent features of users and items respectively. Then, the Funk-SVD combines the user features with item features to reconstruct the rating matrix. After that, we minimize the error between original rating matrix and reconstructed rating matrix, and to alleviate the problem of data sparsity and improve the accuracy of rating prediction effectively. We conducted extensive experiments on Movielens-100K, Movie Tweeting-10k, and Film Trust datasets, and the results show that the rating prediction model based on dual auto-encoders has a superior recommendation performance.

Colon cancer is the fourth leading cause of cancer-related death, and exhibited clinical differences among patients of different ages, including malignancy, metastasis, and mortality rate. Few studies, however, focus on the communications between aging and colon cancer. Here we identified age-dependent differentially expressed genes (DEGs) in colon cancer using TCGA transcriptome data. Through analyzing multi-omics high throughput data, including ATAC-Seq, DNaseI-Seq and ChIP-Seq, we obtained six age-dependent transcription factors in colon cancer, and their age-dependent targets, significantly affecting patients’ overall survivals. Transcription factor ETS1 potentially functioned in both aging process and colon cancer progression through regulating its targets, RGL2 and SLC2A3. In addition, comparing with its relative lower expression levels in elderly patients, higher levels of RGL2 were detected in young patients, and significantly associated with larger tumor size, higher metastasis, and invasions of colon cancer, consistent with the clinical traits that young patients’ colon cancer exhibited late stages with more aggressiveness. Thus, these elements may serve as keys linking aging and colon cancer, and providing new insights and basis for mechanism researches, as well as diagnosis and therapies of colon cancer, especially in young patients.

The target review has been proven to be able to predict the target user’s rating of the target item. However, in practice, it is difficult to obtain the target review promptly. In addition, the target review and the rating may sometimes be inconsistent (such as preference reviews and low ratings). There is currently a lack of research on the above issues. Therefore, this paper proposed a Recommendation algorithm that Simulates the generation of target review semantics and fuses the ID Information (RSII). Specifically, based on the characteristics of the target review available during the model training, this paper designed a teacher module and a review semantics learning module. The teacher module learned the semantics of the target review and guided the review semantics learning model to learn these semantics. Then, this study used the fusion module to dynamically fuse the target review semantics and the ID information, enriching the representation of predictive features, thereby, alleviating the problem of inconsistency between the target review and the rating. Finally, the RSII model was extensively tested on three public datasets. The results showed that compared with seven of the latest and most advanced models, the RSII model improved the MSE metric by 8.81% and the MAE metric by 10.29%.

It has been previously demonstrated that uniaxial cyclic stretching (UCS) induces differentiation of mesenchymal stem cells (MSCs) into osteoblasts in vitro. It is also known that interactions between cells and external forces occur at various aspects including cell–matrix, cytoskeleton, nucleus membrane, and chromatin. However, changes in chromatin landscape during this process are still not clear. The present study was aimed to determine changes of chromatin accessibility under cyclic stretch. The influence of cyclic stretching on the morphology, proliferation, and differentiation of hMSCs was characterized. Changes of open chromatin sites were determined by assay for transposase accessible chromatin with high-throughput sequencing (ATAC-seq). Our results showed that UCS induced cell reorientation and actin stress fibers realignment, and in turn caused nuclear reorientation and deformation. Compared with unstrained group, the expression of osteogenic and chondrogenic marker genes were the highest in group of 1 Hz + 8% strain; this condition also led to lower cell proliferation rate. Furthermore, there were 2022 gene loci with upregulated chromatin accessibility in 1 Hz + 8% groups based on the analysis of chromatin accessibility. These genes are associated with regulation of cell morphogenesis, cell–substrate adhesion, and ossification. Signaling pathways involved in osteogenic differentiation were found in up-regulated GO biological processes. These findings demonstrated that UCS increased the openness of gene loci associated with regulation of cell morphogenesis and osteogenesis as well as the corresponding transcription activities. Moreover, the findings also connect the changes in chromatin accessibility with cell reorientation, nuclear reorientation, and deformation. Our study may provide reference for directed differentiation of stem cells induced by mechanical microenvironments.

In the past decades, long noncoding RNAs (lncRNAs) have been indicated to play important roles in almost every biological process. Aging is a gradual and irreversible biological processes and associated with many human diseases. In this review, we will discuss the function and mechanism of lncRNAs in influencing the progression of aging-related diseases. Moreover, we will discuss the roles of lncRNAs in 3D genome organization. Keywords: Long noncoding RNAs, Aging, 3D genome

Azalomycin F5a was a polyhydroxy macrolide produced by streptomycete strains. Our preliminary researches indicated that it could kill methicillin-resistant Staphylococcus aureus (MRSA) likely by increasing the permeability of cell membrane, and that cell-membrane phospholipids were likely important targets. To confirm this, membrane permeability assay was performed and visualized by fluorescence staining, and then the detailed interactions between azalomycin F5a and model membranes prepared with 1,2-dihexadecanoyl-sn-glycero-3-phospho-(1'-rac-glycerol) (DPPG) were determined using attenuated total reflectance fourier transform infrared spectroscopy and 31P nuclear magnetic resonance techniques. The results indicated that there were strong interactions between azalomycin F5a and model membranes, especially between azalomycin F5a and the polar head of phospholipid. For further evidence and details, the molecular dynamics (MD) simulation of the interactions between azalomycin F5a and DPPG or lysyl-DPPG were performed using Amber16 software package. A strong interaction between the lactone ring of azalomycin F5a and the polar head of DPPG or lysyl-DPPG had been clearly observed. Moreover, a larger distribution probability out of phospholipid bilayer had been discovered for the guanidyl side chain of azalomycin F5a, especially when probable anion molecules anchoring on the cytoplasmic membrane occurred. Therefore, lipoteichoic acid (LTA), a vital component of gram-positive bacterial envelope, was investigated for its probable interactions with azalomycin F5a using broth microdilution method. The results showed that azalomycin F5a-induced MRSA lysis could be prevented by LTA. This deduced that there were some interactions between azalomycin F5a, more likely its guanidyl side chain, and LTA. Thereby, azalomycin F5a increasing the cell-membrane permeability of MRSA had likely achieved by the synergy of its lactone ring binding to the polar head of phospholipid and its guanidyl side chain targeting to LTA, and which had eventually led to the autolysis of MRSA cells.

Increasing studies show that long non-coding RNAs (lncRNAs) play essential roles in various fundamental biological processes. Long non-coding RNA growth arrest-specific transcript 5 (GAS5) showed differential expressions between young and old mouse brains in our previous RNA-Seq data, suggesting its potential role in senescence and brain aging. Examination using quantitative reverse transcription-polymerase chain reaction revealed that GAS5 had a significantly higher expression level in the old mouse brain hippocampus region than the young one. Cellular fractionation using hippocampus-derived HT22 cell line confirmed its nucleoplasm and cytoplasm subcellular localization. Overexpression or knockdown of GAS5 in HT22 cell line revealed that GAS5 inhibits cell cycle progression and promotes cell apoptosis. RNA-Seq analysis of GAS5-knockdown HT22 cells identified differentially expressed genes related to cell proliferation (e.g., DNA replication and nucleosome assembly biological processes). RNA pull-down assay using mouse brain hippocampus tissues showed that potential GAS5 interacting proteins could be enriched into several Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, and some of them are involved in senescence-associated diseases such as Parkinson’s and Alzheimer’s diseases. These results contribute to understand better the underlying functional network of GAS5 and its interacting proteins in senescence at brain tissue and brain-derived cell line levels. Our study may also provide a reference for developing diagnostic and clinic biomarkers of GAS5 in senescence and brain aging.

The research of the runoff structure and its influencing factors in the Xilinhe River Basin not only provides indispensable basic data for the economic development, but also has long-term significance for the protection of grasslands. Based on the runoff data of Xilinhot Hydrological Station from 1960 to 2010 and the daily meteorological data of three surrounding weather stations from 1960 to 2010, the paper calculated the potential evapotranspiration with Penman’s formula and used the combination of Mann-Kendall and Pettitt to diagnose the variation points of characteristic value of runoff distribution during the year. The cumulant slope change rate method is used to quantitatively analyze the contribution rate of climate change and human activities to the uneven distribution coefficient and the complete adjustment coefficient of runoff during the year. The results show that (1) the monthly distribution of runoff in the Xilinhe River Basin is obviously “bimodal” during the year, and the uneven coefficient, complete adjustment coefficient, and concentration in the 2000s are significantly higher than those of 60s-90s. (2) In 1998, the coefficient of uneven distribution of runoff in the Xilinhe River Basin and the coefficient of complete adjustment both showed abrupt changes. (3) Climate change and human activities contributed 11.48% and 88.52% and 9.35% and 90.65% to the uneven distribution coefficient and the complete adjustment coefficient, respectively, of the runoff in the Xilinhe River Basin. Human activities are the main driving factors for changes in the distribution of runoff in the Xilinhe River Basin during the year.

Summary: Three-dimensional chromatin structures undergo dynamic reorganization during mammalian spermatogenesis; however, their impacts on gene regulation remain unclear. Here, we focused on understanding the structure-function regulation of meiotic chromosomes by Hi-C and other omics techniques in mouse spermatogenesis across five stages. Beyond confirming recent reports regarding changes in compartmentalization and reorganization of topologically associating domains (TADs), we further demonstrated that chromatin loops are present prior to and after, but not at, the pachytene stage. By integrating Hi-C and RNA-seq data, we showed that the switching of A/B compartments between spermatogenic stages is tightly associated with meiosis-specific mRNAs and piRNAs expression. Moreover, our ATAC-seq data indicated that chromatin accessibility per se is not responsible for the TAD and loop diminishment at pachytene. Additionally, our ChIP-seq data demonstrated that CTCF and cohesin remain bound at TAD boundary regions throughout meiosis, suggesting that dynamic reorganization of TADs does not require CTCF and cohesin clearance. : Male Reproductive Endocrinology; Genomics; Transcriptomics Subject Areas: Male Reproductive Endocrinology, Genomics, Transcriptomics

In western region of China, the water-sand inrush across overlying strata fissures caused by mining threatens the mine safety production seriously. In order to study the development of water-sand inrush across overlying strata fissures caused by mining, a simulated test system consisted of load support bracket, laboratory module, confined water module, coal seam mining simulator, storage tank, and control system is developed. The combination of coal bearing strata in the south of Shendong mining area is looked on as the engineering background, and a series of new nonhydrophilic composite materials with lower intensity are developed to simulate the coal measure strata. The excavation of physical model can reproduce the whole process of water-sand inrush across overlying strata fissures caused by mining to the life. Under the action of mining and water pressure, after the fourth excavation, the mining-induced vertical fractures of overlying strata pass through the entire bedrock and connect the unconsolidated sand bed, which serves as pathways between the unconsolidated sand bed and working face, triggering water-sand inrush. The water pump suddenly accelerates, and the water yield suddenly increases to the extreme value of 150 L/h. The water pressure rapidly drops to 0 MPa, and a small amount of colored sand enters into the fractures of overlying strata and flows out with the water. The distribution of support pressure around the working face can be divided into 4 areas obviously, that is, the original stress area, the stress-concentrated area, the stress-released area, and the stress restoration area. Test results show that the system is stable and reliable, which have important significance for studying the formation mechanism of water-sand inrush across overlying strata fissures further.

Abstract Background Excessive nerve growth factor (NGF) is commonly found in the follicular fluid of patients with polycystic ovary syndrome (PCOS). Furthermore, oocytes from PCOS patients exhibit lower developmental competence. The purpose of this study was to explore the association between excessive NGF and low oocyte competence in vitro. Methods Excessive NGF was added to mouse cumulus oocyte complexes (COCs) cultured in vitro to investigate meiotic maturation of the oocyte. After culture, mRNA expression levels of Pfkp and Ldha genes in cumulus cells (CCs) and Gdf9, Bmp15 and Fgf8 genes in oocytes, were determined by real-time quantitative polymerase chain reaction (qPCR). We also investigated the mRNA content of Pfkp and Ldha in CCs from PCOS and non-PCOS patients. Results Excessive NGF significantly inhibited oocyte meiotic maturation. The inhibitory effect was mediated by the NGF high-affinity receptor, NTRK1. mRNA content of Pfkp and Ldha genes in CCs was significantly reduced by excessive NGF stimulation. Moreover, the expression levels of Gdf9, Bmp15 and Fgf8 were also decreased in oocytes, and was induced by excessive NGF-stimulated CCs. In addition, lower expression levels of Pfkp and Ldha in CCs were identified in Chinese PCOS patients with excessive NGF (PCOS, 22 ± 2.63 ng/ml, n = 13; non-PCOS, 7.18 ± 2.42 ng/ml, n = 9; p

In this paper, the inhibition effect of an alternative current (AC) electric field on ice crystallization in 0.9 wt % NaCl aqueous solution was confirmed thermodynamically with characterization. An innovative experimental and analytical method, combining differential scanning calorimeter (DSC) measurement with an externally applied electric field was created by implanting microelectrodes in a sample crucible. It was found that the ice crystallization, including pure ice and salty ice, was obviously inhibited after field cooling with an external AC electric field in a frequency range of 100 k–10 MHz, and the crystallization ratio was related to frequency. Compared with non-field cooling, the crystallization ratio of ice crystals was reduced to less than 20% when E = 57.8 kV/m and f = 1 MHz. The dielectric spectrum results show that this inhibition effect of an alternating electric field on ice crystal growth is closely related to the dielectric relaxation process of hydrated ions.

PTPN12 is an important tumor suppressor that plays critical roles in various physiological processes. However, the molecular basis underlying the substrate specificity of PTPN12 remains uncertain. Here, enzymological and crystallographic studies have enabled us to identify two distinct structural features that are crucial determinants of PTPN12 substrate specificity: the pY+1 site binding pocket and specific basic charged residues along its surface loops. Key structurally plastic regions and specific residues in PTPN12 enabled recognition of different HER2 phosphorylation sites and regulated specific PTPN12 functions. In addition, the structure of PTPN12 revealed a CDK2 phosphorylation site in a specific PTPN12 loop. Taken together, our results not only provide the working mechanisms of PTPN12 for desphosphorylation of its substrates but will also help in designing specific inhibitors of PTPN12.

Abstract Meiotic recombination is carried out through a specialized pathway for the formation and repair of DNA double-strand breaks (DSBs) made by the Spo11 protein. The present study shed light on the functional role of cyclin, CYC2, in Tetrahymena thermophila which has transcriptionally high expression level during meiosis process. Knocking out the CYC2 gene results in arrest of meiotic conjugation process at 2.5–3.5 h after conjugation initiation, before the meiosis division starts, and in company with the absence of DSBs. To investigate the underlying mechanism of this phenomenon, a complete transcriptome profile was performed between wild-type strain and CYC2 knock-out strain. Functional analysis of RNA-Seq results identifies related differentially expressed genes (DEGs) including SPO11 and these DEGs are enriched in DNA repair/mismatch repair (MMR) terms in homologous recombination (HR), which indicates that CYC2 could play a crucial role in meiosis by regulating SPO11 and participating in HR.

Antimicrobial resistance has been seriously threatening human health, and discovering new antimicrobial agents from the natural resource is still an important pathway among various strategies to prevent resistance. Guanidine-containing polyhydroxyl macrolides, containing a polyhydroxyl lactone ring and a guanidyl side chain, can be produced by many actinomycetes and have been proved to possess many bioactivities, especially broad-spectrum antibacterial and antifungal activities. To explore the potential of these compounds to be developed into new antimicrobial agents, a review on their structural diversities, spectroscopic characterizations, bioactivities, acute toxicities, antimicrobial mechanisms, and the structure-activity relationship was first performed based on the summaries and analyses of related publications from 1959 to 2019. A total of 63 guanidine-containing polyhydroxyl macrolides were reported, including 46 prototype compounds isolated from 33 marine and terrestrial actinomycetes and 17 structural derivatives. Combining with their antimicrobial mechanisms, structure-activity relationship analyses indicated that the terminal guanidine group and lactone ring of these compounds are vital for their antibacterial and antifungal activities. Further, based on their bioactivities and toxicity analyses, the discovery of guanidyl side-chain targeting to lipoteichoic acid of Staphylococcus aureus indicated that these compounds have a great potency to be developed into antimicrobial and anti-inflammatory drugs.

In the original publication, the funding information was incorrectly published. The correct funding information is provided in this correction. This work is supported by grants from the Projects of International Cooperation and Exchanges Ministry of Science and Technology of China (2013DFG32390) and the National Natural Science Foundation of China (31472059) to X.S. X.S is a recipient of the Young Thousand Talents program (KJ2070000026).