Biology subjects

Xiong, Y.

Publications and source records attributed to Xiong, Y..

At least 127 records · Page 7Linked to original sources

bioRxiv2023

Alterations in the Mammary Gland and Tumor Microenvironment of Formerly Obese Mice

Obesity is a risk factor for breast cancer, and women with obesity that develop breast cancer have a worsened prognosis. Within the mammary gland, obesity causes chronic, macrophage-driven inflammation and adipose tissue fibrosis. To examine the impact of weight loss on the mammary microenvironment, mice were fed high-fat diet to induce obesity, then switched to a low-fat diet. In formerly obese mice, we observed reduced numbers of crown-like structures and fibrocytes in mammary glands, while collagen deposition was not resolved with weight loss. Following transplant of TC2 tumor cells into the mammary glands of lean, obese, and formerly obese mice, diminished collagen deposition and cancer-associated fibroblasts were observed in tumors from formerly obese mice compared to obese mice. When TC2 tumor cells were mixed with CD11b+CD34+ myeloid progenitor cells, collagen deposition within the tumors was significantly greater compared to when tumor cells were mixed with CD11b+CD34- monocytes, suggesting that fibrocytes contribute to early collagen deposition in mammary tumors of obese mice. Overall, these studies show that weight loss resolved some of the microenvironmental conditions within the mammary gland that may contribute to tumor progression.

cancer biology↗

bioRxiv2023

Building a genetic and epigenetic predictive model of breast cancer intrinsic subtypes using large-scale data and hierarchical structure learning

Breast cancer subtyping is a difficult clinical and scientific challenge. The prevalent Prediction Analysis of Microarray of 50 genes (PAM50) system and its Immunohistochemistry (IHC) surrogate showed significant inconsistencies. This is because of the limited training samples, highly variable molecular features and in-efficient strategies used in these classifiers. The rapid development of early screening technologies, especially in the field of circulating tumor DNA, has also challenged the subtyping of breast cancer at the DNA level. By integrating large-scale DNA-level data and using a hierarchical structure learning algorithm, we developed Unified Genetic and Epigenetic Subtyping (UGES), a new intrinsic subtype classifier. The benchmarks showed that the use of all classes of DNA alterations worked much better than single classes, and that the multi-step hierarchical learning is crucial, which improves the overall AUC score by 0.074 compared to the one-step multi-classification method. Based on these insights, the ultimate UGES was trained as a three-step classifier on 50831 DNA features of 2065 samples, including mutations, copy number aberrations, and methylations. UGES achieved overall AUC score 0.963, and greatly improved the clinical stratification of patients, as each stratas survival difference became statistically more significant p-value=9.7e-55 (UGES) vs 2.2e-47 (PAM50). Finally, UGES identified 52 subtype-level DNA biomarkers that can be targeted in early screening technology to significantly expand the time window for precision care. The analysis code is freely available at https://github.com/labxscut/UGES.

bioinformatics↗

bioRxiv2023

Enteric glial hub cells coordinate intestinal motility

The enteric nervous system is a complex network of neurons and glia within the gut that coordinate gut motility. By optimizing single nucleus RNA-sequencing methods and spatial transcriptomics, we generated maps of the mouse duodenum and identified distinct molecular classes of enteric glia across the intestine with unique morphological and spatial identities. Here we show enteric glial functional specialization, with one myenteric subtype directly sensing force and expressing the mechanosensory ion channel PIEZO2. Genetic reduction of PIEZO2 in enteric glial populations enriched for this mechanosensory subtype led to defects in gastrointestinal motility. These results provide insight into the multifaceted functions of distinct enteric glial cell subtypes in maintaining gut health, and emphasize the importance of considering subtype-specific roles of enteric glia in a wide range of diseases and disorders.

neuroscience↗

bioRxiv2023

OmicVerse: A single pipeline for exploring the entire transcriptome universe

Single-cell sequencing is frequently marred by "interruptions" due to limitations in sequencing throughput, yet bulk RNA-seq may harbor these ostensibly "interrupted" cells. In response, we introduce the single cell trajectory blending from Bulk RNA-seq (BulkTrajBlend) algorithm, a component of the OmicVerse suite that leverages a Beta-Variational AutoEncoder for data deconvolution and graph neural networks for the discovery of overlapping community. This approach proficiently interpolates and restores the continuity of "interrupted" cells within single-cell RNA sequencing dataset. Furthermore, OmicVerse provides an extensive toolkit for bulk and single cell RNA-seq analysis, offering uniform access to diverse methodologies, streamlining computational processes, fostering exquisite data visualization, and facilitating the extraction of novel biological insights to advance scientific research.

bioinformatics↗

bioRxiv2023

Persistent biofluid small molecule alterations induced by Trypanosoma cruzi infection are not restored by antiparasitic treatment

Table of contents graphic O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=105 SRC="FIGDIR/small/543565v1_figu1.gif" ALT="Figure 1"> View larger version (25K): org.highwire.dtl.DTLVardef@10c3a0aorg.highwire.dtl.DTLVardef@1b69d6dorg.highwire.dtl.DTLVardef@199867org.highwire.dtl.DTLVardef@64b20b_HPS_FORMAT_FIGEXP M_FIG C_FIG Chagas Disease (CD), caused by Trypanosoma cruzi (T. cruzi) protozoa, is a complicated parasitic illness with inadequate medical measures for diagnosing infection and monitoring treatment success. To address this gap, we analyzed changes in the metabolome of T. cruzi-infected mice via liquid chromatography tandem mass spectrometry analysis of clinically-accessible biofluids: saliva, urine, and plasma. Urine was the most indicative of infection status, across mouse and parasite genotypes. Metabolites perturbed by infection in the urine include kynurenate, acylcarnitines, and threonylcarbamoyladenosine. Based on these results, we sought to implement urine as a tool for assessment of CD treatment success. Strikingly, it was found that mice with parasite clearance following benznidazole antiparasitic treatment had comparable overall urine metabolome to mice that failed to clear parasites. These results match with clinical trial data in which benznidazole treatment did not improve patient outcomes in late-stage disease. Overall, this study provides insights into new small molecule-based CD diagnostic methods and a new approach to assess functional treatment response.

microbiology↗

bioRxiv2023

Comparative analysis of drug-like EP300/CREBBP acetyltransferase inhibitors

The human acetyltransferase paralogs EP300 and CREBBP are master regulators of lysine acetylation whose activity has been implicated in various cancers. In the half-decade since the first drug-like inhibitors of these proteins were reported, three unique molecular scaffolds have taken precedent: an indane spiro-oxazolidinedione (A-485), a spiro-hydantoin (iP300w), and an aminopyridine (CPI-1612). Despite increasing use of these molecules to study lysine acetylation, the dearth of data regarding their relative biochemical and biological potencies makes their application as chemical probes a challenge. To address this gap, here we present a comparative study of drug-like EP300/CREBBP acetyltransferase inhibitors. First, we determine the biochemical and biological potencies of A-485, iP300w, and CPI-1612, highlighting the increased potency of the latter two compounds at physiological acetyl-CoA concentrations. Cellular evaluation shows that inhibition of histone acetylation and cell growth closely aligns with the biochemical potencies of these molecules, consistent with an on-target mechanism. Finally, we demonstrate the utility of comparative pharmacology by using it to investigate the hypothesis that increased CoA synthesis caused by knockout of PANK4 can competitively antagonize binding of EP300/CREBBP inhibitors and demonstrate proof-of-concept photorelease of a potent inhibitor molecule. Overall, our study demonstrates how knowledge of relative inhibitor potency can guide the study of EP300/CREBBP-dependent mechanisms and suggests new approaches to target delivery, thus broadening the therapeutic window of these preclinical epigenetic drug candidates.

biochemistry↗

bioRxiv2023

Spinal Cord Injury regulates circular RNA expression in axons

IntroductionNeurons transport mRNA and translational machinery to axons for local translation. After spinal cord injury (SCI), de novo translation is assumed to enable neurorepair. Knowledge of the identity of axonal mRNAs that participate in neurorepair after SCI is limited. We sought to identify and understand how axonal RNAs play a role in axonal regeneration. MethodsWe obtained preparations enriched in axonal mRNAs from control and SCI rats by digesting spinal cord tissue with cold-active protease (CAP). The digested samples were then centrifuged to obtain a supernatant that were then sequenced. We used bioinformatics analyses to identify DEGS and map them to various biological processes. We validated the DEGs by RT-qPCR and RNA-scope. ResultsThe supernatant fraction was highly enriched for axonal mRNA. Using Gene Ontology, the second most significant pathway for all differentially expressed genes (DEGs) was axonogenesis. Among the DEGs was Rims2, which is predominately a circular RNA (circRNA) in the CNS. We show that Rims2 RNA within spinal cord axons is circular. We found an additional 200 putative circRNAs in the axonal-enriched fraction. Knockdown in primary rat cortical neurons of the RNA editing enzyme ADAR1, which inhibits formation of circRNAs, significantly increased axonal outgrowth. Focusing on Rims2 we used Circular RNA Interactome to predict that several of the miRNAs that bind to circRims2 also bind to the 3UTR of GAP-43, PTEN or CREB1, all known regulators of axonal outgrowth. Axonally-translated GAP-43 supports axonal elongation and we detect GAP-43 mRNA in the rat axons by RNAscope. DiscussionBy using our method for enrichment of axonal RNA, we detect SCI induced DEGs, including circRNA such as Rims2. Ablation of ADAR1, the enzyme that regulates circRNA formation, promotes axonal outgrowth of cortical neurons. We developed a pathway model using Circular RNA Interactome that indicates that Rims2 through miRNAs can regulate the axonal translation GAP-43 a known regulator of axonal regeneration indicating that axonal mRNA contribute to regeneration.

neuroscience↗

bioRxiv2023

Designer DNA NanoGripper

DNA has shown great biocompatibility, programmable mechanical properties, and structural addressability at the nanometer scale, making it a versatile material for building high precision nanorobotics for biomedical applications. Herein, we present design principle, synthesis, and characterization of a DNA nanorobotic hand, called the "NanoGripper", that contains a palm and four bendable fingers as inspired by human hands, bird claws, and bacteriophages evolved in nature. Each NanoGripper finger has three phalanges connected by two flexible and rotatable joints that are bendable in response to binding to other entities. Functions of the NanoGripper have been enabled and driven by the interactions between moieties attached to the fingers and their binding partners. We showcase that the NanoGripper can be engineered to interact with and capture various objects with different dimensions, including gold nanoparticles, gold NanoUrchins, and SARS-CoV-2 virions. When carrying multiple DNA aptamer nanoswitches programmed to generate fluorescent signal enhanced on a photonic crystal platform, the NanoGripper functions as a sensitive viral biosensor that detects intact SARS-CoV-2 virions in human saliva with a limit of detection of [~] 100 copies/mL, providing RT-PCR equivalent sensitivity. Additionally, we use confocal microscopy to visualize how the NanoGripper-aptamer complex can effectively block viral entry into the host cells, indicating the viral inhibition. In summary, we report the design, synthesis, and characterization of a complex nanomachine that can be readily tailored for specific applications. The study highlights a path toward novel, feasible, and efficient solutions for the diagnosis and therapy of other diseases such as HIV and influenza. One-sentence summaryDesign, synthesis, characterization, and functional showcase of a human-hand like designer DNA nanobot

bioengineering↗

bioRxiv2023

A Quadrivalent mRNA immunization elicits potent immune responses against vaccinia and monkeypox viral antigens - a step closer to a broad orthopoxvirus vaccine

The global outbreak of the 2022 monkeypox virus infection of human raised the public health concerns of the threat of human-to-human transmission of zoonotic diseases. Given the evidence that other orthopoxviruses including cowpox and camelpox were also reported infectious to human, and that the reemerging risk of smallpox as a bioterrorist or accidental laboratory escape exists, there is an urgent need to develop a poxvirus vaccine with a broad protection of orthopoxviruses to stockpile for future emergency. Extensive studies of vaccinia virus (VACV) suggested that multiple VACV antigens, such as A27, L1, A33 and B5, showed high level similarity in terms of immunogenicity to their respective homologous antigens of other orthopoxviruses. These findings paved the ground for VACV antigens to be used as potential vaccine targets for development of a universal poxvirus vaccine. In this study, we construct a novel poxvirus vaccine candidate, mRNA-ALAB-LNP, encoding four vaccinia viral antigens A27, L1, A33 and B5. Strong anti-L1-specific antibody and moderate anti-A33-, anti-A27- and anti-B5-specific antibody responses were induced in mice after a single immunization. The antibody responses to all four antigens were significantly boosted after the second shot with all IgG titers >5 logs and highest being anti-A33 IgG. The high level of binding antibodies showed potent neutralizing capability against vaccinia virus. Specific IFN-{gamma} responses were detected to all four antigens with the highest cellular response being that induced by the same antigen, A33. When evaluating the cross reactivity, equivalent or better serum IgG responses were seen in responses to corresponding monkeypox antigens A35, M1, A29 and B6, in comparison to vaccinia antigens. Apparently, the mRNA vaccine encoding four vaccinia antigens induced immunity not only to vaccinia virus but also to monkeypox, suggesting that the mRNA-ALAB may be a candidate for potential vaccine development against infection of monkeypox, smallpox and possibly other orthopoxviruses.

immunology↗

bioRxiv2023

Discovery of microRNA-derived RNAs and proteins in human cells

MicroRNAs (miRNAs) are short noncoding RNAs that can regulate gene expression through the binding of their 5-ends to mRNAs. However, the biological effects of miRNA binding via their 3-ends remain unclear. Here, we discover that the exact reverse pairing of the 3-ends of miRNAs or miRNA-like RNAs, collectively termed microsized RNAs (msRNAs), with template RNAs can initiate the production of msRNA-derived RNAs (msdRs), which can subsequently be translated into polypeptides (msdPs). Using 2,632 human msRNAs from miRBase, 11,121 msdRs and 1,239 msdPs were predicted based on a 15-nucleotide pairing threshold, and the presence of representative msdRs and msdPs was confirmed in human cells. Of clinical relevance, msdP0188 is highly expressed in human lung and breast cancers, and its corresponding msRNAs and msdRs represent novel anti-cancer targets. Notably, inhibiting telomerase reverse transcriptase, a putative RNA-dependent RNA polymerase identified by bioinformatic screening, led to reduced levels of msdP0188 in human cancer cells. Our findings reveal a novel "msRNA [->] msdR [->] msdP" axis, expanding the classical central dogma and predicting many previously unrecognized RNAs and polypeptides with potential biological functions in human cells and other systems.

cell biology↗

bioRxiv2023

Netrin-1 directs vascular patterning and maturity in the developing kidney

Blood filtering by the kidney requires the establishment of an intricate vascular system that works to support body fluid and organ homeostasis. Despite these critical roles, little is known about how vascular architecture is established during kidney development. More specifically, how signals from the kidney influence vessel maturity and patterning remains poorly understood. Netrin-1 (Ntn1) is a secreted ligand critical for vessel and neuronal guidance. Here, we demonstrate that Ntn1 is expressed by stromal progenitors in the developing kidney, and conditional deletion of Ntn1 from Foxd1+ stromal progenitors (Foxd1GC/+;Ntn1fl/fl) results in hypoplastic kidneys that display extended nephrogenesis. Despite expression of the netrin-1 receptor Unc5c in the adjacent nephron progenitor niche, Unc5c knockout kidneys develop normally. The netrin-1 receptor Unc5b is expressed by embryonic kidney endothelium and therefore we interrogated the vascular networks of Foxd1GC/+;Ntn1fl/fl kidneys. Wholemount, 3D analyses revealed the loss of a predictable vascular pattern in mutant kidneys. As vascular patterning has been linked to vessel maturity, we investigated arterialization in these mutants. Quantification of the CD31+ endothelium at E15.5 revealed no differences in metrics such as the number of branches or branch points, whereas the arterial vascular smooth muscle metrics were significantly reduced at both E15.5 and P0. In support of these results, whole kidney RNA-seq showed upregulation of angiogenic programs and downregulation of muscle-related programs which included smooth muscle-associated genes. Together, our findings highlight the significance of netrin-1 to proper vascularization and kidney development.

developmental biology↗

bioRxiv2023

Development, Characterization, and Structural Analysis of a Genetically Encoded Red Fluorescent Peroxynitrite Biosensor

Boronic acid-containing fluorescent molecules have been widely used to sense hydrogen peroxide and peroxynitrite, which are important reactive oxygen and nitrogen species in biological systems. However, it has been challenging to gain specificity. Our previous studies developed genetically encoded, green fluorescent peroxynitrite biosensors by genetically incorporating a boronic acid-containing noncanonical amino acid (ncAA), p-boronophenylalanine (pBoF), into the chromophore of circularly permuted green fluorescent proteins (cpGFPs). In this work, we introduced pBoF to amino acid residues spatially close to the chromophore of an enhanced circularly permuted red fluorescent protein (ecpApple). Our effort has resulted in two responsive ecpApple mutants: one bestows reactivity toward both peroxynitrite and hydrogen peroxide, while the other, namely pnRFP, is a selective red fluorescent peroxynitrite biosensor. We characterized pnRFP in vitro and in live mammalian cells. We further studied the structure and sensing mechanism of pnRFP using X-ray crystallography, 11B-NMR, and computational methods. The boron atom in pnRFP adopts an sp2-hybridization geometry in a hydrophobic pocket, and the reaction of pnRFP with peroxynitrite generates a product with a twisted chromophore, corroborating the observed "turn-off" fluorescence response. Thus, this study extends the color palette of genetically encoded peroxynitrite biosensors, provides insight into the response mechanism of the new biosensor, and demonstrates the versatility of using protein scaffolds to modulate chemoreactivity.

biochemistry↗

bioRxiv2023

Protein language model powers accurate and fast sequence search for remote homology

Homologous protein search is one of the most commonly used methods for protein annotation and analysis. Compared to structure search, detecting distant evolutionary relationships from sequences alone remains challenging. Here we propose PLMSearch (Protein Language Model), a homologous protein search method with only sequences as input. With deep representations from a pre-trained protein language model to predict similarity, PLMSearch can capture the remote homology information hidden behind the sequences. Extensive experimental results show that PLMSearch can search millions of query-target protein pairs in seconds like MMseqs2 while increasing the sensitivity by more than threefold, and is comparable to state-of-the-art structure search methods. In particular, unlike traditional sequence search methods, PLMSearch can recall most remote homology pairs with low sequence similarity but sharing similar structures. PLMSearch is freely available at https://dmiip.sjtu.edu.cn/PLMSearch.

bioinformatics↗

bioRxiv2023

Vascular Smooth Muscle Cell Mechanotransduction Through Serum and Glucocorticoid Inducible Kinase-1 Promotes Interleukin-6 Production and Macrophage Accumulation in Murine Hypertension

ObjectiveThe objective of this investigation was to demonstrate that in vivo induction of hypertension (HTN) and in vitro cyclic stretch of aortic VSMCs can cause SGK-1-dependent production of cytokines to promote macrophage accumulation as agents of vascular remodeling. MethodsHTN was induced in C57Bl/6 mice with AngiotensinII (AngII) infusion (1.46mg/kg/day x 21 days) with or without systemic infusion of EMD638683 (2.5mg/kg/day x 21 days), a selective SGK-1 inhibitor. Systolic blood pressure (SBP) was recorded on days 0 and 21. At terminal study, abdominal aortas were harvested to quantify SGK-1 activity (pSGK-1:SGK-1) by immunoblot. Additional replicates were digested and analyzed by flow cytometry for abundance of CD11b+/F4-80+ cells (macrophages). Plasma was analyzed by ELISA to quantify IL-6 and MCP-1. Aortic VSMCs from wild-type (WT) mice were subjected to 12% biaxial cyclic stretch for 3 or 12 hours +/- EMD638683 (10M) and +/- SGK-1siRNA with subsequent QPCR for IL-6 and MCP-1 expression. Culture media was analyzed by ELISA for IL-6 and MCP-1. Aortic VSMCs from SGK-1flox+/+ mice were transfected with Cre-Adenovirus to knockout SGK-1 (SGK-1KO VSMCs) and underwent parallel tension experimentation. Computational modeling was employed to simulate VSMC signaling due to mechanical strain and AngII. Statistical analysis included ANOVA with significance at p<0.05. ResultsSGK-1 activity (pSGK-1:SGK-1) was upregulated in the abdominal aorta of mice with HTN and significantly reduced by treatment with EMD638683. Concurrently, increased CD11b+/F4-80+ cells and plasma IL-6 levels in the HTN group and reduction with EMD638683 was observed. This mirrored the increased abundance of IL-6 in media from Stretch WT VSMCs, and attenuation of the effect with EMD638683. Treating WT VSMCs with SGK-1siRNA likewise inhibited IL-6 expression. MCP-1 also demonstrated increased expression and secretion into the media in WT VSMCs with Stretch. Further supporting the integral role of mechanical signaling through SGK-1, target gene expression and cytokine secretion was unchanged in SGK-1KO VSMCs with Stretch, and computer modeling confirmed SGK-1 as an intersecting node of signaling due to mechanical strain and AngII. In summation, this data suggests a biomechanical link between aortic VSMC mechanotransduction and cytokine production to promote macrophage accumulation, mediated in-part by SGK-1 activation. ConclusionMechanotransduction through SGK-1 is instrumental in pro-inflammatory cytokine production and aortic macrophage accumulation in systemic HTN, therefore further investigation into targeting this kinase may present opportunities to modulate hypertensive vascular remodeling.

physiology↗

bioRxiv2023

Inhibiting Bet1-mediated transport of MMP14 to plasma membrane impaired GBM cell invasion

PurposeGlioblastoma (GBM) is the most aggressive and common form of brain cancer in adults. GBM is characterised by poor survival as the lack of effective therapies. This research aims to detect the roles of SNAREs in GBM and improve our knowledge of targeting therapy for GBM. Materials and methodsthe expression of SNAREs and their correlation with overall survival (OS) in GBM are investigated using the GEPIA. The level of BET1 in GBM cell lines was tested by RT-qPCR, and its biological functions in GBM cells were tested by Transwell assay and CCK8 kit. The effect of BET1 on the location of MMP14 is identified by Immunofluorescence. ResultsThe expression profile of SNARE family members in GBM tissue is changed dramatically. Among them, the mRNA levels of BET1 and VAMP3 are up-regulated, and their expression negatively correlates with OS. BET1 is also increased in GBM Cell Lines, and it is required for efficient GBM cell migration and invasion partly because it mediates the transport of MMP14 to the plasma membrane. ConclusionGBM has highly diffusive and infiltrative ability in nature, making complete surgical resection almost impossible. Our data shows that BET1 is highly expressed in GBM tissue, negatively correlated with OS, and essential for GBM cell migration and invasion. These results indicate that SNARE BET1 may present a potential target for GBM treatment.

cell biology↗

bioRxiv2023

RAMEN identifies effective indicators for severe COVID and Long COVID patients

The COVID-19 pandemic has significantly altered global socioeconomic structures and individual lives. Understanding the disease mechanisms and facilitating diagnosis requires comprehending the complex interplay among clinical factors like demographics, symptoms, comorbidities, treatments, lab results, complications, and other metrics, and their relation to outcomes such as disease severity and long term outcomes (e.g., post-COVID-19 condition/long COVID). Conventional correlational methods struggle with indirect and directional connections among these factors, while standard graphical methods like Bayesian networks are computationally demanding for extensive clinical variables. In response, we introduced RAMEN, a methodology that integrates Genetic Algorithms with random walks for efficient Bayesian network inference, designed to map the intricate relationships among clinical variables. Applying RAMEN to the Biobanque quebecoise de la COVID-19 (BQC19) dataset, we identified critical markers for long COVID and varying disease severity. The Bayesian Network, corroborated by existing literature and supported through multi-omics analyses, highlights significant clinical variables linked to COVID-19 outcomes. RAMENs ability to accurately map these connections contributes substantially to developing early and effective diagnostics for severe COVID-19 and long COVID.

bioinformatics↗

bioRxiv2023

T492I mutation alters SARS-CoV-2 properties via modulating viral non-structural proteins

The historically dominant SARS-CoV-2 Delta variants and the currently dominant Omicron variants carry a T492I substitution within the non-structural protein 4 (NSP4). Based on a combination of in silico analyses, we predicted that the T492I mutation increases the transmissibility and adaptability of the virus. We confirmed this hypothesis by performing competition experiments in hamsters and in human airway tissue culture models. Furthermore, we show that the T492I mutation also increases the replication capacity and infectiveness of the virus, and improves its ability to evade antibody neutralization induced by previous variants. Mechanistically, the T492I mutation increases cleavage efficiency of the viral main protease NSP5 by enhancing enzyme-substrate binding, resulting in increased production of nearly all non-structural proteins processed by NSP5. Importantly, T492I mutation suppresses the viral RNA associated chemokines in monocytic macrophages, which may contribute to the attenuated pathogenicity of Omicron variants. Our results highlight the importance of the NSP4 mutation in the evolutionary dynamics of SARS-CoV-2 and identify a novel target for the development of broad-spectrum antiviral agents.

microbiology↗

bioRxiv2022

Target Recycling Amplification Process for Digital Detection of Exosomal MicroRNAs Through Photonic Resonator Absorption Microscopy

Exosomal microRNAs (miRNAs) have considerable potential as pivotal biomarkers to monitor cancer development, dis-ease progression, treatment effects and prognosis. Here, we report an efficient target recycling amplification process (TRAP) for the digital detection of exosomal miRNAs using photonic resonator absorption microscopy (PRAM). Through toehold-mediated DNA strand displacement reactions, we achieve multiplex digital detection with sub-attomolar sensitivity in 20 minutes, robust selectivity for single nucleotide variants, and a broad dynamic range from 1 aM to 1 pM. We then applied our TRAP system to quantify miRNA in exosomal total RNAs isolated from human cancer cell lines. Compared with traditional qRT-PCR methods, TRAP showed similar accuracy in profiling exosomal miRNAs derived from cancer cells, but also exhibited at least 31-fold and 61-fold enhancement in the limits of miRNA-375 and miRNA-21 detection, respectively. The TRAP approach is ideal for exosomal or circulating miRNA biomarker quantification, where the miRNAs are present in low concentrations or sample volume, with potentials for frequent, low-cost, and minimally invasive point-of-care testing.

bioengineering↗