Near-infrared light controlled protein degradation by photo-caged lenalidomide and pomalidomide.

Immunomodulatory drugs (e.g. thalidomide, lenalidomide and pomalidomide) have been proven highly successful in clinical treatment of multiple myeloma. However, systematic degradation of zinc finger transcriptional factors induced by these drugs could lead to severe systematic toxicity in patients. Previous reports of NVOC caged pomalidomide attempted to regulate its activity using UVA irradiation, but their application was limited by high cytotoxicity and low tissue penetration. Here, we reported red-shifted BODIPY caged lenalidomide and pomalidomide that enabled red-light controlled protein degradation with spatiotemporal precision.

Transmission in home environment associated with the second wave of COVID-19 pandemic in India.

India has suffered from the second wave of COVID-19 pandemic since March 2021. This wave of the outbreak has been more serious than the first wave pandemic in 2020, which suggests that some new transmission characteristics may exist. COVID-19 is transmitted through droplets, aerosols, and contact with infected surfaces. Air pollutants are also considered to be associated with COVID-19 transmission. However, the roles of indoor transmission in the COVID-19 pandemic and the effects of these factors in indoor environments are still poorly understood. Our study focused on reveal the role of indoor transmission in the second wave of COVID-19 pandemic in India. Our results indicated that human mobility in the home environment had the highest relative influence on COVID-19 daily growth rate in the country. The COVID-19 daily growth rate was significantly positively correlated with the residential percent rate in most state-level areas in India. A significant positive nonlinear relationship was found when the residential percent ratio ranged from 100 to 120%. Further, epidemic dynamics modelling indicated that a higher proportion of indoor transmission in the home environment was able to intensify the severity of the second wave of COVID-19 pandemic in India. Our findings suggested that more attention should be paid to the indoor transmission in home environment. The public health strategies to reduce indoor transmission such as ventilation and centralized isolation will be beneficial to the prevention and control of COVID-19.

Industrial Anomaly Detection with Skip Autoencoder and Deep Feature Extractor.

Over recent years, with the advances in image recognition technology for deep learning, researchers have devoted continued efforts toward importing anomaly detection technology into the production line of automatic optical detection. Although unsupervised learning helps overcome the high costs associated with labeling, the accuracy of anomaly detection still needs to be improved. Accordingly, this paper proposes a novel deep learning model for anomaly detection to overcome this bottleneck. Leveraging a powerful pre-trained feature extractor and the skip connection, the proposed method achieves better feature extraction and image reconstructing capabilities. Results reveal that the areas under the curve (AUC) for the proposed method are higher than those of previous anomaly detection models for 16 out of 17 categories. This indicates that the proposed method can realize the most appropriate adjustments to the needs of production lines in order to maximize economic benefits.

Patient-derived organoid (PDO) platforms to facilitate clinical decision making.

Based on recent advances in organoid research as well as the need to find more accurate models for drug screening in cancer research, patient-derived organoids have emerged as an effective in vitro model system to study cancer. Showing numerous advantages over 2D cell lines, 3D cell lines, and primary cell culture, organoids have been applied in drug screening to demonstrate the correlation between genetic mutations and sensitivity to targeted therapy. Organoids have also been used in co-clinical trials to compare drug responses in organoids to clinical responses in the corresponding patients. Numerous studies have reported the successful use of organoids to predict therapy response in cancer patients. Recently, organoids have been adopted to predict treatment response to radiotherapy and immunotherapy. The development of high throughput drug screening and organoids-on-a-chip technology can advance the use of patient-derived organoids in clinical practice and facilitate therapeutic decision-making.

One-tube SARS-CoV-2 detection platform based on RT-RPA and CRISPR/Cas12a.

BACKGROUND: COVID-19 has spread rapidly around the world, affecting a large percentage of the population. When lifting certain mandatory measures for an economic restart, robust surveillance must be established and implemented, with nucleic acid detection for SARS-CoV-2 as an essential component. METHODS: We tried to develop a one-tube detection platform based on RT-RPA (Reverse Transcription and Recombinase Polymerase Isothermal Amplification) and DNA Endonuclease-Targeted CRISPR Trans Reporter (DETECTR) technology, termed OR-DETECTR, to detect SARS-CoV-2. We designed RT-RPA primers of the RdRp and N genes following the SARS-CoV-2 gene sequence. We optimized reaction components so that the detection process could be carried out in one tube. Specificity was demonstrated by detecting nucleic acid samples from pseudoviruses from seven human coronaviruses and Influenza A (H1N1). Clinical samples were used to validate the platform and all results were compared to rRT-PCR. RNA standards and pseudoviruses diluted by different gradients were used to demonstrate the detection limit. Additionally, we have developed a lateral flow assay based on OR-DETECTR for detecting COVID-19. RESULTS: The OR-DETECTR detection process can be completed in one tube, which takes approximately 50 min. This method can specifically detect SARS-CoV-2 from seven human coronaviruses and Influenza A (H1N1), with a low detection limit of 2.5 copies/µl input (RNA standard) and 1 copy/µl input (pseudovirus). Results of six samples from SARS-CoV-2 patients, eight samples from patients with fever but no SARS-CoV-2 infection, and one mixed sample from 40 negative controls showed that OR-DETECTR is 100% consistent with rRT-PCR. The lateral flow assay based on OR-DETECTR can be used for the detection of COVID-19, and the detection limit is 2.5 copies/µl input. CONCLUSIONS: The OR-DETECTR platform for the detection of COVID-19 is rapid, accurate, tube closed, easy-to-operate, and free of large instruments.

Single-cell profiling of long noncoding RNAs and their cell lineage commitment roles via RNA-DNA-DNA triplex formation in mammary epithelium.

Long noncoding RNAs (lncRNAs), which are crucial for organ development, exhibit cell-specific expression. Thus, transcriptomic analysis based on total tissue (bulk-seq) cannot accurately reflect the expression pattern of lncRNAs. Here, we used high-throughput single-cell RNA-seq data to investigate the role of lncRNAs using the hierarchical model of mammary epithelium. With our comprehensive annotation of the mammary epithelium, lncRNAs showed much greater cell-lineage specific expression than coding genes. The lineage-specific lncRNAs were functionally correlated with lineage commitment through the coding genes via the cis- and trans-effects of lncRNAs. For the working mechanism, lncRNAs formed a triplex structure with the DNA helix to regulate downstream lineage-specific marker genes. We used lncRNA-Carmn as an example to validate the above findings. Carmn, which is specifically expressed in mammary gland stem cells (MaSCs) and basal cells, positively regulated the Wnt signaling ligand Wnt10a through formation of a lncRNA-DNA-DNA triplex, and thus controlled the stemness of MaSCs. Our study suggests that lncRNAs play essential roles in cell-lineage commitment and provides an approach to decipher lncRNA functions based on single-cell RNA-seq data.

Long non-coding RNA NEAT1-centric gene regulation.

Nuclear paraspeckle assembly transcript 1 (NEAT1) is a long non-coding RNA that is widely expressed in a variety of mammalian cell types. An increasing number of studies have demonstrated that NEAT1 plays key roles in various biological and pathological processes; therefore, it is important to understand how its expression is regulated and how it regulates the expression of its target genes. Recently, we found that NEAT1 expression could be regulated by signal transducer and activator of transcription 3 and that altered NEAT1 expression epigenetically regulates downstream gene transcription during herpes simplex virus-1 infection and Alzheimer's disease, suggesting that NEAT1 acts as an important sensor and effector during stress and disease development. In this review, we summarize and discuss the molecules and regulatory patterns that control NEAT1 gene expression and the molecular mechanism via which NEAT1 regulates the expression of its target genes, providing novel insights into the central role of NEAT1 in gene regulation.

Modulation of SRSF2 expression reverses the exhaustion of TILs via the epigenetic regulation of immune checkpoint molecules.

The elevated expression of immune checkpoints by the tumor microenvironment is associated with poor prognosis in several cancers due to the exhaustion of tumor-infiltrating lymphocytes (TILs), and the effective suppression of the expression of these genes is key to reversing the exhaustion of TILs. Herein, we determined that serine/arginine-rich splicing factor 2 (SRSF2) is a target for blocking the tumor microenvironment-associated immunosuppressive effects. We found that the expression of SRSF2 was increased in exhausted T cells and that SRSF2 was involved in multiple immune checkpoint molecules mediating TILs' exhaustion. Furthermore, SRSF2 was revealed to regulate the transcription of these immune checkpoint genes by associating with an acyl-transferases P300/CBP complex and altering the H3K27Ac level near these genes, thereafter influencing the recruitment of signal transducer and activator of transcription 3 (STAT3) to these gene promoters. Collectively, our data indicated that SRSF2 functions as a modulator of the anti-tumor response of T cells and may be a therapeutic target for reversing the exhaustion of TILs.

ACME: pan-specific peptide-MHC class I binding prediction through attention-based deep neural networks.

MOTIVATION: Prediction of peptide binding to the major histocompatibility complex (MHC) plays a vital role in the development of therapeutic vaccines for the treatment of cancer. Algorithms with improved correlations between predicted and actual binding affinities are needed to increase precision and reduce the number of false positive predictions. RESULTS: We present ACME (Attention-based Convolutional neural networks for MHC Epitope binding prediction), a new pan-specific algorithm to accurately predict the binding affinities between peptides and MHC class I molecules, even for those new alleles that are not seen in the training data. Extensive tests have demonstrated that ACME can significantly outperform other state-of-the-art prediction methods with an increase of the Pearson correlation coefficient between predicted and measured binding affinities by up to 23 percentage points. In addition, its ability to identify strong-binding peptides has been experimentally validated. Moreover, by integrating the convolutional neural network with attention mechanism, ACME is able to extract interpretable patterns that can provide useful and detailed insights into the binding preferences between peptides and their MHC partners. All these results have demonstrated that ACME can provide a powerful and practically useful tool for the studies of peptide-MHC class I interactions. AVAILABILITY AND IMPLEMENTATION: ACME is available as an open source software at https://github.com/HYsxe/ACME. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Magnetic bead-enzyme assemble for triple-parameter telomerase detection at single-cell level.

In this work, we developed a triple-parameter strategy for the detection of telomerase activity from cancer cells and urine samples. This strategy was developed based on magnetic bead-enzyme hybrids combined with fluorescence analysis, colorimetric assay, or adenosine triphosphate (ATP) meter as readout. The application of magnetic bead-enzyme hybrids has the advantages of magnetic separation and signal amplification. These detection methods can be used individually or in combination to achieve the optimal sensing performance and make the results more convincing. Among them, the ATP meter with portable size had easy operation and low cost, and this response strategy provided a higher sensitivity at the single-cell level. The designed strategy was suitable as naked-eye sensor and point-of-care testing (POCT) for rapid assaying of telomerase activity. Graphical abstract Magnetic bead-enzyme assemble for triple-parameter telomerase detection.

Quantum Dot Nanobeacons for Single RNA Labeling and Imaging.

Detection and imaging RNAs in live cells is in high demand. Methodology for such a purpose is still a challenge, particularly for single RNA detection and imaging in live cells. In this study, a type of quantum dot (QD) nanobeacon with controllable valencies was constructed by precisely conjugating the black hole quencher (BHQ1) and phosphorothioate comodified DNA onto CdTe:Zn2+ QDs via a one-pot hydrothermal method. The nanobeacon with only one conjugated DNA was used to label and detect low-abundance nucleic acids in live cells, and single HIV-1 RNAs were detected and imaged in live HIV-1 integrated cells. Additionally, QD nanobeacon-labeled HIV-1 genomic RNAs were encapsulated in progeny viral particles, which can be used to track the uncoating process of single viruses. The current study provides a platform for nucleic acid labeling and imaging with high sensitivity, being especially meaningful for tracking of individual RNAs in live cells.

Directing cellular information flow via CRISPR signal conductors.

The complex phenotypes of eukaryotic cells are controlled by decision-making circuits and signaling pathways. A key obstacle to implementing artificial connections in signaling networks has been the lack of synthetic devices for efficient sensing, processing and control of biological signals. By extending sgRNAs to include modified riboswitches that recognize specific signals, we can create CRISPR-Cas9-based 'signal conductors' that regulate transcription of endogenous genes in response to external or internal signals of interest. These devices can be used to construct all the basic types of Boolean logic gates that perform logical signal operations in mammalian cells without needing the layering of multiple genetic circuits. They can also be used to rewire cellular signaling events by constructing synthetic links that couple different signaling pathways. Moreover, this approach can be applied to redirect oncogenic signal transduction by controlling simultaneous bidirectional (ON-OFF) gene transcriptions, thus enabling reprogramming of the fate of cancer cells.

High expression of enhancer RNA MARC1 or its activation by DHT is associated with the malignant behavior in bladder cancer.

Enhancer RNAs (eRNAs), a subclass of noncoding RNA from enhancers, have biological functions in gene expression. However, their potential role in bladder cancer (BCa) remains largely unknown. The present study investigated the functional role of androgen-associated androgen receptor (AR) mediated-eRNA MARC1 (eMARC1) in BCa progression. Cell proliferation, migration, and apoptosis of BCa cell lines (5637 and T24) with different eMARC1 expression levels or treated with 5α-dehydrotestosterone (DHT) were investigated. In the current study, we discovered that eMARC1 was highly expressed in BCa tissues and cell lines, and eMARC1 overexpression promoted the progression of BCa cells, while knockdown of eMARC1 suppressed tumorigenesis. DHT treatment significantly elevated eMARC1 expression levels, which also facilitated cell proliferation, motility, and inhibited cell apoptosis. We further found that eMARC1 silencing impaired the androgenic effect of DHT in BCa cells. These results suggested that eMARC1 exerted its effects on BCa cell progression, and DHT promoted bladder cancer progression by activating eMARC1.

iCatch: a new strategy for capturing large DNA fragments using homing endonucleases.

Natural genetic materials contain many biosynthetic gene clusters encoding potentially valuable natural products, many of which can be used directly without codon optimization or other manipulations. With the development of synthetic biology, several DNA assembly standards have been proposed, conveniently facilitating the reuse of natural materials. Among these standards, the iBrick assembly standard was developed by our laboratory to manipulate large DNA fragments, employing two homing endonucleases. Considering the difficulty of cloning large iBrick parts using conventional endonuclease-mediated restriction and ligation methods, we herein present a new method, known as iCatch, which readily captures biosynthetic gene clusters. As the clusters cloned by iCatch have the prefix and suffix of the iBrick standard, they serve as new iBrick parts and are therefore conducive to further editing and assembly with the iBrick standard. iCatch employs the natural homologous recombination system to flank the region of interest with I-SceI and PI-PspI recognition sites, after which the genome is digested with I-SceI or PI-PspI and the fragments are then self-ligated to clone the target DNA fragments. We used this method to successfully capture the actinorhodin biosynthetic cluster from Streptomyces coelicolor and then heterologously expressed this cluster in a thermophilic Streptomyces strain. We propose that iCatch can be used for the cloning of DNA sequences that are dozens of kilobases in length, facilitating the heterologous expression of microbial natural products. Moreover, this cloning methodology can be a complementary tool for the iBrick standard, especially in applications requiring the manipulation of large DNA fragments.

Oestrogen promotes tumorigenesis of bladder cancer by inducing the enhancer RNA-eGREB1.

In recent years, studies have shown that enhancer RNAs (eRNAs) can be transcribed from enhancers. Increasing evidence has revealed that eRNAs play critical roles in the development of various cancers. Oestrogen-associated eRNAs are closely related to breast cancer. In view of the gender differences in bladder cancer (BCa), we suppose that oestrogen-associated eRNAs are also involved in tumorigenesis of BCa. In our study, we first demonstrated that eGREB1 derived from the enhancer of an oestrogen-responsive gene-GREB1 was up-regulated in BCa tissues, and the expression level of eGREB1 is positively associated with the histological grade and TNM stage of BCa. Knockdown of eGREB1 by CRISPR-Cas13a could inhibit cell proliferation, migration and invasion and induce apoptosis in BCa cells T24 and 5637. Besides, we exhibited the promoting effect of oestrogen on BCa cells. What's more, down-regulation of eGREB1 could improve the malignant biological characteristics of BCa cells induced by oestrogen. In conclusion, our data indicated that eGREB1 plays oncogenic role and oestrogen may promote the occurrence and progression of BCa by inducing eGREB1 production. Our findings provide new insights into the prevention of BCa and develop a novel therapeutic target for the treatment of BCa.

Verteporfin inhibits YAP-induced bladder cancer cell growth and invasion via Hippo signaling pathway.

The highly conserved Hippo signaling pathway is one of the most important pathways involved in tumorigenesis and progress. Previous studies show that YAP, the transcriptional coactivator of Hippo pathway, is expressed highly in many clinical bladder cancer tissues and plays crucial role on bladder cancer progress. To find the YAP-specific target drug and its molecular mechanism in bladder cancer, we apply Verteporfin (VP), a YAP specific inhibitor to function as anti-bladder cancer drug and discover that VP is able to inhibit bladder cancer cell growth and invasion in a dosage dependent manner. Moreover, we demonstrate that VP may inhibit bladder cancer cell growth and invasion via repressing target genes' expression of the Hippo signaling pathway. In further study, we provide evidence that VP is able to inhibit excessive YAP induced bladder cancer cell growth and invasion. To address the repressive function of VP against YAP in bladder cancer, we check the target genes' expression and find VP can dramatically repress YAP overexpression induced Hippo pathway target genes' expression. Taken together, we discover that VP inhibits YAP-induced bladder cancer cell growth and invasion via repressing the target genes' expression of Hippo signaling pathway.

A nonsense mutation in Ccdc62 gene is responsible for spermiogenesis defects and male infertility in repro29/repro29 mice.

Phenotype-driven mutagenesis is an unbiased method to identify novel genes involved in spermatogenesis and other reproductive processes. Male repro29/repro29 mice generated by the Reproductive Genomics Program at the Jackson Laboratory were infertile with deformed sperm and poor motility. Using selected exonic capture and massively parallel sequencing technologies, we identified a nonsense mutation in the exon 6 of coiled-coil domain-containing 62 gene (Ccdc62), which results in a formation of a premature stop codon and a truncated protein. Among the tissues examined, CCDC62 was found to be expressed at the highest level in mouse testis by reverse transcriptase-PCR (RT-PCR) and Western blot analysis. With immunofluorescent staining, we demonstrated that CCDC62 was expressed in the cytoplasm and the developing acrosome in the spematids of mouse testis, and was specifically localized at the acrosome in mature sperm. The complementation analysis by mating repro29/+ mice with Ccdc62 -/- mice (generated by CRISPR-Cas9 strategy) further provided genetic proof that the infertility of repro29/repro29 mice was caused by Ccdc62 mutation. Finally, it was found that intracellular colocalization and interaction of CCDC62 and Golgi-associated PDZ and coiled-coil motif-containing protein may be important for acrosome formation. Taken together, this study identified a nonsense mutation in Ccdc62, which directly results in male infertility in repro29/repro29 mice.

SPRY4-IT1: A novel oncogenic long non-coding RNA in human cancers.

Long non-coding RNAs are classified as a kind of RNA, which are longer than 200 nucleotides in length and cannot be translated into proteins. Multiple studies have demonstrated that long non-coding RNAs are involved in various cellular processes, including proliferation, differentiation, cell death, and metastasis. Among numerous long non-coding RNAs, we focus on Sprouty4-Intron 1 (SPRY4-IT1), a well-known long non-coding RNA that is overexpressed in various kinds of tumor tissues and cell lines. Accumulating evidences show that SPRY4-IT1 was dysregulated in various cancers, including melanoma, breast cancer, esophageal squamous cell carcinoma, non-small cell lung cancer, gastric cancer, colon cancer, and hepatocellular carcinoma, and amplification of SPRY4-IT1 was associated with different clinicopathological features of cancer patients. Importantly, SPRY4-IT1 exerts important roles in tumor progression and metastasis. However, detailed molecular mechanisms of SPRY4-IT1 in cancer progression and metastasis were poorly understood. In this review, we have focused on the characteristics of SPRY4-IT1 and illustrated the biological function and mechanism of SPRY4-IT1 in cancer development.

Deficiency of SPATA46, a Novel Nuclear Membrane Protein, Causes Subfertility in Male Mice.

Teratozoospermia is generally associated with clinical infertility. Despite numerous studies, the molecular mechanisms underlying male infertility are still poorly understood. In the present study, we demonstrated that deletion of Spata46, a gene encoding a novel protein of unknown function found in mouse testis, was responsible for male subfertility, and the cause of subfertility was characterized as abnormal sperm head shape and a failure of sperm-egg fusion. We also demonstrated that SPATA46 was expressed predominantly in condensed spermatids, with a highly specific localization restricted to the subacrosomal area; the protein is located at the nuclear membrane due to a transmembrane region in the N-terminus of the protein. At the subcellular level, SPATA46-deficient condensed spermatids displayed structural defects consisting of a discontinuous nuclear envelope and a cavity in the nucleus associated with an abnormal nuclear shape. Additionally, in vitro, we determined that the absence of SPATA46 led to accumulation of sperm around the perivitelline space of eggs, and the same phenomenon was also observed for natural sperm incubated with an anti-SPATA46 antibody, suggesting functional relevance of SPATA46 for sperm-egg fusion. Taken together, these results indicated that SPATA46 is a novel protein involved in reshaping of the sperm head and sperm-egg fusion.

Synthetic Tet-inducible small hairpin RNAs targeting hTERT or Bcl-2 inhibit malignant phenotypes of bladder cancer T24 and 5637 cells.

Small hairpin RNA (shRNA) can inhibit the malignant phenotypes of tumor cell through ribonucleic acid interference (RNAi). However, it is hardly to be regulated and it may induce few phenotypic changes. Here, we build a type of tetracycline (Tet)-inducible vectors which can achieve regulatable expression of shRNA in a time-dependent manner by using synthetic biology approach. In order to prove the effectiveness of this device, we chose hTERT and Bcl-2 as target genes and test the utility of the device on 5637 and T24 cell lines. The experiments show that the Tet-inducible small hairpin RNA can effectively suppress their target genes and generate anti-cancer effects on both 5637 and T24 cell lines. The device we build not only can inhibit proliferation but also can induce apoptosis and suppress migration of the bladder cancer cell lines 5637 and T24. The Tet-inducible small hairpin RNAs may provide a novel strategy for the treatment of human bladder cancer in the future.