Biology subjects

Liu, T.

Publications and source records attributed to Liu, T..

At least 253 records · Page 14Linked to original sources

bioRxiv2023

Spatiotemporal Control of Genomics and Epigenomics by Ultrasound

CRISPR (clustered regularly interspaced short palindromic repeats) is a revolutionary technology for genome editing. Its derived technologies such as CRISPR activation (CRISPRa) and CRISPR interference (CRISPRi) further allow transcriptional and epigenetic modulations. Focused ultrasound (FUS) can penetrate deep in biological tissues and induce mild hyperthermia in a confined region to activate heat-sensitive genes. Here we engineer a set of CRISPR(a/i) tools containing heat-sensitive genetic modules controllable by FUS for the regulation of genome and epigenome in live cells and animals. We demonstrated the capabilities of FUS-inducible CRISPRa, CRISPRi, and CRISPR (FUS-CRISPR(a/i)) to upregulate, repress, and knockout exogenous and/or endogenous genes, respectively, in different cell types. We further targeted FUS-CRISPR to telomeres in tumor cells to induce telomere disruption, inhibiting tumor growth and enhancing tumor susceptibility to killing by chimeric antigen receptor (CAR)-T cells. FUS-CRISPR-mediated telomere disruption for tumor priming combined with CAR-T therapy demonstrated synergistic therapeutic effects in xenograft mouse models. The FUS-CRISPR(a/i) toolbox allows the remote, noninvasive, and spatiotemporal control of genomic and epigenomic reprogramming in vivo, with extended applications in cancer treatment.

bioengineering↗

bioRxiv2023

mRNA condensation fluidizes the cytoplasm

The intracellular environment is packed with macromolecules of mesoscale size, and this crowded milieu significantly influences cell physiology. When exposed to stress, mRNAs released after translational arrest condense with RNA binding proteins, resulting in the formation of membraneless RNA protein (RNP) condensates known as processing bodies (P-bodies) and stress granules (SGs). However, the impact of the assembly of these condensates on the biophysical properties of the crowded cytoplasmic environment remains unclear. Here, we find that upon exposure to stress, polysome collapse and condensation of mRNAs increases mesoscale particle diffusivity in the cytoplasm. Increased mesoscale diffusivity is required for the efficient formation of Q-bodies, membraneless organelles that coordinate degradation of misfolded peptides that accumulate during stress. Additionally, we demonstrate that polysome collapse and stress granule formation has a similar effect in mammalian cells, fluidizing the cytoplasm at the mesoscale. We find that synthetic, light-induced RNA condensation is sufficient to fluidize the cytoplasm, demonstrating a causal effect of RNA condensation. Together, our work reveals a new functional role for stress-induced translation inhibition and formation of RNP condensates in modulating the physical properties of the cytoplasm to effectively respond to stressful conditions.

cell biology↗

bioRxiv2023

SQANTI3: curation of long-read transcriptomes for accurate identification of known and novel isoforms

The emergence of long-read RNA sequencing (lrRNA-seq) has provided an unprecedented opportunity to analyze transcriptomes at isoform resolution. However, the technology is not free from biases, and transcript models inferred from these data require quality control and curation. In this study, we introduce SQANTI3, a tool specifically designed to perform quality analysis on transcriptomes constructed using lrRNA-seq data. SQANTI3 provides an extensive naming framework to describe transcript model diversity in comparison to the reference transcriptome. Additionally, the tool incorporates a wide range of metrics to characterize various structural properties of transcript models, such as transcription start and end sites, splice junctions, and other structural features. These metrics can be utilized to filter out potential artifacts. Moreover, SQANTI3 includes a Rescue module that prevents the loss of known genes and transcripts exhibiting evidence of expression but displaying low-quality features. Lastly, SQANTI3 incorporates IsoAnnotLite, which enables functional annotation at the isoform level and facilitates functional iso-transcriptomics analyses. We demonstrate the versatility of SQANTI3 in analyzing different data types, isoform reconstruction pipelines, and sequencing platforms, and how it provides novel biological insights into isoform biology. The SQANTI3 software is available at https://github.com/ConesaLab/SQANTI3.

bioinformatics↗

bioRxiv2023

RealSeq2: a software integrated with UMI identification, error correction, and methylation modifications storing

High-throughput UMI technology sequencing is widely used in early tumor screening, detection, recurrence monitoring, etc. Detecting extremely low-frequency mutations is especially important for monitoring tumor recurrence, so high-precision data, as well as high-quality data, are required. We developed RealSeq2, a new integrated data-preprocessing software based on fastp and gencore, to achieve adapter removal, quality control, UMI identification, and generate consensus reads by clustering and error correction using multithreading in high-throughput next-generation sequencing background. RealSeq2 also supports methylation data of 5-methylcytosine bisulfite-free sequencing. RealSeq2 defined a new tag in SAM for storing methylation information, which is beneficial for co-identifying methylation sites and mutation sites for downstream analysis. RealSeq2 includes three submodules: ReadsProfiler, ReadsCleaner, and ReadsRecycler. In addition, the output format file (BAM or SAM) is universal for downstream analyses. RealSeq2 is the preferred upstream analysis software for the co-detection of ultra-low frequency mutations and bisulfite-free methylation data. The error profile provides data support for downstream analysis. Additionally, XM tags will become a standard protocol for recording methylation signals.

bioinformatics↗

bioRxiv2023

Paternal CHH methylation potentiates stress responses against Pseudomonas syringae in Arabidopsis progenies

Systemic acquired resistance (SAR) is an induced immune mechanism in plants, involving epigenetic regulation by chromatin remodeling and DNA methylation, which can be inherited to progeny following stress exposure. Intersexual epigenetic conflict sometimes leads to unequal expression of maternal and paternal alleles in offspring, resulting in parent-of-origin effects of inheritance. To better understand the parental contributions of epialleles in plant defense, isogenic Arabidopsis parental lines were mock-treated (M) and Pseudomonas syringae (Pst)-treated (P) for reciprocal crosses to produce F1 progenies (MP, PM). Together with their self-fertilized F1 descendants (MM, PP), the genome-wide inherited DNA methylation and transcriptomic changes against Pst were analyzed. F1 descendants shared widespread DNA methylation and transcriptional changes at transposable elements (TEs) and genes. The confrontation of epigenomes triggers the reprogramming of DNA methylation in reciprocal crosses, resulting in transgressive segregation that also shows the parental effect of Pst treatment. Compared to PM, the MP (Pst-primed paternal genome) was found to contributes to CHH hypermethylation, which is associated with processes in plant-pathogen interaction, including carbohydrate metabolism, glutathione metabolism and stronger translation process, which potentially contribute to improved disease resistance in MP in response to Pst challenge. Our data suggested a parent-of-origin effect of defense priming that contributes differently toward improved defense response in progenies.

genetics↗

bioRxiv2023

Assessing Genetic Diversity in Critically Endangered Chieniodendron hainanense Populations within Fragmented Habitats in Hainan

AbstractHabitat fragmentation engenders a reduction in the geographic distribution of species, thereby rendering diminutive populations susceptible to extinction due to environmental, demographic, and genetic factors. Chieniodendron hainanense (henceforth C. hainanense) exemplifies a wild plant with extremely small populations (WPESP) and faces endangerment, necessitating urgent national conservation efforts. Elucidating the genetic diversity of C. hainanense is crucial for uncovering underlying mechanisms and devising protective strategies. In the present study, 35 specimens from six distinct cohort groups were genotyped utilizing genotyping-by-sequencing (GBS) and single nucleotide polymorphism (SNP) methodologies. The results indicated that C. hainanense exhibits limited genetic diversity. Observed heterozygosity within C. hainanense populations spanned from 10.79% to 14.55%, with an average value of 13.15%. The six C. hainanense populations can be categorized into two distinct groups: (1) Diaoluoshan and Baishaling, and (2) Wuzhishan, Huishan, Bawangling, and Jianfengling. The degree of genetic differentiation among C. hainanense populations is relatively weak. The observed loss of diversity can be attributed to the effects of natural selection.

ecology↗

bioRxiv2023

Pharyngeal Mechanosensory Neurons Control Food Swallow in Drosophila melanogaster

As the early step of food ingestion, the swallow is under rigorous sensorimotor control. Nevertheless, the mechanisms underlying swallow control at a molecular and circuitry level remain largely unknown. Here, we find that mutation of the mechanotransduction channel genes nompC, Tmc, or piezo impairs the regular pumping rhythm of the cibarium during feeding of the fruit fly Drosophila melanogaster. A group of multi-dendritic mechanosensory neurons, which co-express the three channels, wrap the cibarium and are crucial for coordinating the filling and emptying of the cibarium. Inhibition of them causes difficulty in food emptying in the cibarium, while their activation leads to difficulty in cibarium filling. Synaptic and functional connections are detected between the pharyngeal mechanosensory neurons and the motor circuit that controls swallow. This study elucidates the role of mechanosensation in swallow, and provides insights for a better understanding of the neural basis of food swallow.

neuroscience↗

bioRxiv2023

Dysregulated Wnt and NFAT signaling in a Parkinson's disease LRRK2 G2019S knock-in model

BackgroundParkinsons disease (PD) is a progressive late-onset neurodegenerative disease leading to physical and cognitive decline. Mutations of leucine-rich repeat kinase 2 (LRRK2) are the most common genetic cause of PD. LRRK2 is a complex scaffolding protein with known regulatory roles in multiple molecular pathways. Two prominent examples of LRRK2-modulated pathways are Wingless/Int (Wnt) and nuclear factor of activated T-cells (NFAT) signaling. Both are well described key regulators of immune and nervous system development as well as maturation. The aim of this study was to establish the physiological and pathogenic role of LRRK2 in Wnt and NFAT signaling in the brain, as well as the potential contribution of the non-canonical Wnt/Calcium pathway. MethodsIn vivo cerebral Wnt and NFATc1 signaling activity was quantified in LRRK2 G2019S mutant knock-in (KI) and LRRK2 knockout (KO) male and female mice with repeated measures over 28 weeks, employing lentiviral luciferase biosensors, and analyzed using a mixed-effect model. To establish spatial resolution, we investigated tissues, and primary neuronal cell cultures from different brain regions combining luciferase signaling activity, immunohistochemistry, qPCR and western blot assays. Results were analyzed by unpaired t-test with Welchs correction or 2-way ANOVA with post hoc corrections. ResultsIn vivo Wnt signaling activity in LRRK2 KO and LRRK2 G2019S KI mice was increased significantly [~]3-fold, with a more pronounced effect in males ([~]4-fold) than females ([~]2-fold). NFATc1 signaling was reduced [~]0.5-fold in LRRK2 G2019S KI mice. Brain tissue analysis showed region-specific expression changes in Wnt and NFAT signaling components. These effects were predominantly observed at the protein level in the striatum and cerebral cortex of LRRK2 KI mice. Primary neuronal cell culture analysis showed significant genotype-dependent alterations in Wnt and NFATc1 signaling under basal and stimulated conditions. Wnt and NFATc1 signaling was primarily dysregulated in cortical and hippocampal neurons respectively. ConclusionsOur study further built on knowledge of LRRK2 as a Wnt and NFAT signaling protein. We identified complex changes in neuronal models of LRRK2 PD, suggesting a role for mutant LRRK2 in the dysregulation of NFAT, and canonical and non-canonical Wnt signaling.

neuroscience↗

bioRxiv2023

Matrin3 mediates differentiation through stabilizing chromatin accessibility and chromatin loop-domain interactions, and YY1 mediated enhancer-promoter interactions

Although emerging evidence indicates that alterations in proteins within nuclear compartments elicit changes in chromosomal architecture and differentiation, the underlying mechanisms are not well understood. Here we investigate the direct role of the abundant nuclear complex protein Matrin3 (Matr3) in chromatin architecture and development in the context of myogenesis. Using an acute targeted protein degradation platform (dTAG-Matr3), we reveal the dynamics of development-related chromatin reorganization. Upon acute depletion of Matr3, gains in chromatin accessibility and MyoD binding were observed, prior to widespread loss in the steady-state Matr3-knockout. These initial changes correlated with gene expression changes later in development. High-throughput chromosome conformation capture (Hi-C) experiments revealed substantial chromatin loop rearrangements soon after Matr3 depletion. Notably, YY1 binding was detected in close proximity to enhancer and promoter regions, accompanied by the emergence of novel YY1-mediated enhancer-promoter loops, which occurred concurrently with changes in histone modifications and chromatin-level binding patterns. Overall, our results suggest that Matr3 mediates differentiation through stabilizing chromatin accessibility and chromatin loop-domain interactions, and highlight a conserved and direct role for Matr3 in maintenance of chromosomal architecture.

molecular biology↗

bioRxiv2023

The first genome assembly of the amphibian nematode parasite (Aplectana chamaeleonis)

Cosmocercoid nematodes are common parasites of the digestive tract of amphibians. Genomic resources are important for understanding the evolution of a species and the molecular mechanisms of parasite adaptation. So far, no genome resource of Cosmocercoid has been reported. In 2020, a massive Cosmocercoid infection was found in the small intestine of a toad, causing severe intestinal blockage. We morphologically identified this parasite as A. chamaeleonis. Here, we report the first A. chamaeleonis genome with a genome size of 1.04 Gb. The repeat content of this A. chamaeleonis genome is 72.45 %, and the total length is 751 Mb. This resource is fundamental for understanding the evolution of Cosmocercoid and provides the molecular basis for Cosmocercoid infection and control.

genomics↗

bioRxiv2023

What can we learn when fitting a complex gene expressionmodel to a simple telegraph model?

In experiments, the distributions of mRNA or protein numbers in single cells are often fitted to the random telegraph model which includes synthesis and decay of mRNA or protein, and switching of the gene between active and inactive states. While commonly used, this model does not describe how fluctuations are influenced by crucial biological mechanisms such as feedback regulation, non-exponential gene inactivation durations, and multiple gene activation pathways. Here we investigate the dynamical properties of four relatively complex gene expression models by fitting their steady-state mRNA or protein number distributions to the simple telegraph model. We show that despite the underlying complex biological mechanisms, the telegraph model with three effective parameters can accurately capture the steady-state gene product distributions, as well as the conditional distributions in the active gene state, of the complex models. Some effective parameters are reliable and can reflect realistic dynamic behaviors of the complex models, while others may deviate significantly from their real values in the complex models. The effective parameters can also be applied to characterize the capability for a complex model to exhibit multimodality. Using additional information such as single-cell data at multiple time points, we provide an effective method of distinguishing the complex models from the telegraph model. Furthermore, using measurements under varying experimental conditions, we show that fitting the mRNA or protein number distributions to the telegraph model may even reveal the underlying gene regulation mechanisms of the complex models. The effectiveness of these methods is confirmed by analysis of single-cell data for E. coli and mammalian cells. All these results are robust with respect to cooperative transcriptional regulation and extrinsic noise. In particular, we find that faster relaxation speed to the steady state results in more precise parameter inference under large extrinsic noise.

biophysics↗

bioRxiv2023

RNA Sensing and Innate Immunity Constitutes a Barrier for Interspecies Chimerism

Interspecies chimera formation with human pluripotent stem cells (PSCs) holds great promise to generate humanized animal models and provide donor organs for transplant. However, the approach is currently limited by low levels of human cells ultimately represented in chimeric embryos. Different strategies have been developed to improve chimerism by genetically editing donor human PSCs. To date, however, it remains unexplored if human chimerism can be enhanced in animals through modifying the host embryos. Leveraging the interspecies PSC competition model, here we discovered retinoic acid-inducible gene I (RIG-I)-like receptor (RLR) signaling, an RNA sensor, in "winner" cells plays an important role in the competitive interactions between co-cultured mouse and human PSCs. We found that genetic inactivation of Ddx58/Ifih1-Mavs-Irf7 axis compromised the "winner" status of mouse PSCs and their ability to outcompete PSCs from evolutionarily distant species during co-culture. Furthermore, by using Mavs-deficient mouse embryos we substantially improved unmodified donor human cell survival. Comparative transcriptome analyses based on species-specific sequences suggest contact-dependent human-to-mouse transfer of RNAs likely plays a part in mediating the cross-species interactions. Taken together, these findings establish a previously unrecognized role of RNA sensing and innate immunity in "winner" cells during cell competition and provides a proof-of-concept for modifying host embryos, rather than donor PSCs, to enhance interspecies chimerism.

cell biology↗

bioRxiv2023

Integrated single-cell analysis defines the epigenetic basis of castration-resistant prostate luminal cells

Understanding prostate response to castration and androgen receptor signaling inhibitors (ARSI) is critical to improving long-term prostate cancer (PCa) patient survival. Here we use a multi-omics approach on 229,794 single cells to create a mouse single-cell reference atlas better suited to interpreting mouse prostate biology and castration response. Our reference atlas refines single-cell annotations and provides chromatin context, which, when coupled with mouse lineage tracing demonstrates that the castration-resistant luminal cells are distinct from the pre-existent urethra- proximal stem/progenitor cells. Molecular pathway analysis and therapeutic studies further implicate JUN/FOS, WNT/{beta}-Catenin, FOXQ1, NF{kappa}B, and JAK/STAT pathways as the major drivers of castration- resistant luminal populations with high relevance to human PCa. Importantly, we demonstrate the utility of our datasets, which can be explored through an interactive portal (https://visportal.roswellpark.org/data/tang/), to aid in developing novel combination treatments with ARSI for advanced PCa patients. O_FIG O_LINKSMALLFIG WIDTH=164 HEIGHT=200 SRC="FIGDIR/small/530998v1_ufig1.gif" ALT="Figure 1"> View larger version (62K): org.highwire.dtl.DTLVardef@8bd384org.highwire.dtl.DTLVardef@1ebc004org.highwire.dtl.DTLVardef@8b4dbforg.highwire.dtl.DTLVardef@19d7052_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2023

Reprogramming an RNA-guided archaeal endonuclease for genome editing

RNA-guided endonucleases developed for genome editing have been identified exclusively from bacteria until now. Recently, the RNA-guided TnpB endonucleases encoded by bacterial IS200/IS605 and IS607 families have been reprogrammed for genome editing. In fact, these insertion sequence families are also widely distributed in archaea. However, whether archaea-derived TnpB could be used for genome editing remains unclear. Here, we found that TnpB derived from Sulfolobus islandicus REY15A (SisTnpB1) was an Mg2+/Mn2+-dependent RNA-guided endonuclease, and it could cleave DNA next to the 5 TTTAA transposon-associated motif (TAM) to generate double-stranded DNA breaks at the 3-end. We also found that SisTnpB1 was active at 37-85{degrees}C and tolerant to variations of TAM. Moreover, SisTnpB1 could be reprogrammed to edit bacterial cells with an editing efficiency of 100% at 37{degrees}C or 45{degrees}C. In summary, our finding expanded the resources of RNA-guided endonucleases for genome editing, especially those derived from archaea, the third domain of life.

molecular biology↗

bioRxiv2023

Chemogenetic activation of corticotropin-releasing factor-expressing neurons in the anterior bed nucleus of the stria terminalis reduces effortful motivation behaviors

Corticotropin-releasing factor (CRF) in the anterior bed nucleus of the stria terminalis (aBNST) is associated with chronic stress and avoidance behavior. However, CRF+ BNST neurons project to reward- and motivation-related brain regions, suggesting a potential role in motivated behavior. We used chemogenetics to selectively activate CRF+ aBNST neurons in male and female CRF-ires-Cre mice during an effort-related choice task and a concurrent choice task. In both tasks, mice were given the option either to exert effort for high value rewards or to choose freely available low value rewards. Acute chemogenetic activation of CRF+ aBNST neurons reduced barrier climbing for a high value reward in the effort-related choice task in both males and females. Furthermore, acute activation of CRF+ aBNST neurons also reduced effortful lever pressing in high-performing males in the concurrent choice task. These data suggest a novel role for CRF+ aBNST neurons in effort-based decision and motivated behavior.

neuroscience↗

bioRxiv2023

MGA deletion leads to Richter's transformation via modulation of mitochondrial OXPHOS

Richters transformation (RT) is a progression of chronic lymphocytic leukemia (CLL) to aggressive lymphoma. MGA (Max gene associated), a functional MYC suppressor, is mutated at 3% in CLL and 36% in RT. However, genetic models and molecular mechanisms of MGA deletion driving CLL to RT remain elusive. We established a novel RT mouse model by knockout of Mga in the Sf3b1/Mdr CLL model via CRISPR-Cas9 to determine the role of Mga in RT. Murine RT cells exhibit mitochondrial aberrations with elevated oxidative phosphorylation (OXPHOS). We identified Nme1 (Nucleoside diphosphate kinase) as a Mga target through RNA sequencing and functional characterization, which drives RT by modulating OXPHOS. As NME1 is also a known MYC target without targetable compounds, we found that concurrent inhibition of MYC and ETC complex II significantly prolongs the survival of RT mice in vivo. Our results suggest that Mga-Nme1 axis drives murine CLL-to-RT transition via modulating OXPHOS, highlighting a novel therapeutic avenue for RT. Statement of SignificanceWe established a murine RT model through knockout of Mga in an existing CLL model based on co-expression of Sf3b1-K700E and del(13q). We determined that the MGA/NME1 regulatory axis is essential to the CLL-to-RT transition via modulation of mitochondrial OXPHOS, highlighting this pathway as a novel target for RT treatment.

cancer biology↗

bioRxiv2023

DPHL v2: An updated and comprehensive DIA pan-human assay library for quantifying more than 14,000 proteins.

A comprehensive pan-human spectral library is critical for biomarker discovery using mass spectrometry (MS)-based proteomics. DPHL v1, a previous pan-human library built from 1096 data-dependent acquisition (DDA) MS data of 16 human tissue types, allows quantifying 10,943 proteins. However, a major limitation of DPHL v1 is the lack of semi-tryptic peptides and protein isoforms, which are abundant in clinical specimens. Here, we generated DPHL v2 from 1608 DDA-MS data acquired using Orbitrap mass spectrometers. The data included 586 DDA-MS newly acquired from 17 tissue types, while 1022 files were derived from DPHL v1. DPHL v2 thus comprises data from 24 sample types, including several cancer types (lung, breast, kidney, and prostate cancer, among others). We generated four variants of DPHL v2 to include semi-tryptic peptides and protein isoforms. DPHL v2 was then applied to a publicly available colorectal cancer dataset with 286 DIA-MS files. The numbers of identified and significantly dysregulated proteins increased by at least 21.7% and 14.2%, respectively, compared with DPHL v1. Our findings show that the increased human proteome coverage of DPHL v2 provides larger pools of potential protein biomarkers.

bioinformatics↗

bioRxiv2023

An axon - T cell feedback loop enhances inflammation and axon degeneration

Inflammation is closely associated with many neurodegenerative disorders. Yet whether inflammation causes or exacerbates neurodegeneration has been challenging to define because the two processes are so closely linked. Here we disentangle inflammation from the axon damage it causes by individually blocking cytotoxic T cell function and axon degeneration. We model inflammatory damage in mouse skin, a barrier tissue that, despite frequent inflammation, must maintain proper functioning of a dense array of axon terminals. We show that sympathetic axons control skin inflammation through release of norepinephrine, which suppresses activation of gamma delta T cells via the {beta}2 adrenergic receptor. Strong inflammatory stimulation in the form of the toll like receptor 7 (TLR7) agonist imiquimod (IMQ) causes progressive gamma delta T cell-mediated, Sarm-1-dependent loss of these immunosuppressive sympathetic axons, a positive feedback loop that removes a physiological brake on T cells, resulting in enhanced inflammation and inflammatory axon damage.

neuroscience↗