Biology subjects

Zhang, L.

Publications and source records attributed to Zhang, L..

At least 703 records · Page 39Linked to original sources

bioRxiv2024

Single-cell RNA sequencing reveals recruitment of the M2-like CCL8high macrophages in Lewis lung carcinoma-bearing mice following hypofractionated radiotherapy

Tumor-associated macrophages (TAMs) are a crucial factor in reprogramming the tumor microenvironment following radiotherapy. The mechanisms underlying this process remain to be elucidated. Here, we seek to investigate the effects of hypofractionated radiotherapy on macrophages dynamics in a subcutaneous Lewis lung carcinoma murine model. Utilizing single-cell RNA sequencing, we identified a distinct M2-like population of macrophages with high Ccl8 expression level post-hypofractionated radiotherapy. Remarkbly, hypofractionated radiotherapy promoted CCL8high macrophages infiltration and reprogrammed them by upregulating immunosuppressive genes and downregulating antigen-presenting genes, leading to an immunosuppressive tumor microenvironment. Further, we demonstrated that hypofractionated radiotherapy amplified the CCL signaling pathway, enhancing the pro-tumorigenic functions of CCL8high macrophages and promoting macrophages recruitment. The combination therapy of hypofractionated radiotherapy with the CCL signal inhibitor Bindarit was effective in reducing M2 macrophages infiltration and extending the duration of local tumor control. This research highlights the potential of targeting TAMs and introduces a novel combination to improve the efficacy of hypofractionated radiotherapy.

immunology↗

bioRxiv2024

Type I interferon governs immunometabolic checkpoints that coordinate inflammation during Staphylococcal infection

Fine-tuned inflammation during infection enables pathogen clearance while minimizing host damage. Inflammation regulation depends on macrophage metabolic plasticity, yet coordination of metabolic and inflammatory programs is underappreciated. Here we show that type I interferon (IFN) temporally guides metabolic control of inflammation during methicillin-resistant Staphylococcus aureus (MRSA) infection. In macrophages, staggered Toll-like receptor and type I IFN signaling permitted a transient respiratory burst followed by inducible nitric oxide synthase (iNOS)-mediated OXPHOS disruption. OXPHOS disruption promoted type I IFN, suppressing other pro-inflammatory cytokines, notably IL-{beta}. iNOS expression peaked at 24h post-infection, followed by lactate-driven iNOS repression via histone lactylation. In contrast, type I IFN pre-conditioning sustained infection-induced iNOS expression and amplified type I IFN. Cutaneous MRSA infection in mice constitutively expressing epidermal type I IFN led to elevated iNOS levels, impaired wound healing, vasculopathy, and lung infection. Thus, kinetically regulated type I IFN signaling coordinates immunometabolic checkpoints that control infection-induced inflammation. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=101 SRC="FIGDIR/small/575104v1_ufig1.gif" ALT="Figure 1"> View larger version (27K): org.highwire.dtl.DTLVardef@977b1eorg.highwire.dtl.DTLVardef@1142c6corg.highwire.dtl.DTLVardef@170776borg.highwire.dtl.DTLVardef@b02c2f_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2024

Transcriptome Dynamics in Mouse Amygdala under Acute and Chronic Stress Revealed by Thio-labeled RNA Sequencing

Both acute and chronic stress have significant impact on brain functions. The amygdala is essential in mediating stress responses, but how its transcriptomic dynamics change under stress remains elusive. To overcome the difficulties in detecting subtle stress-induced changes by evaluating total RNA using classic RNA sequencing, we conducted thiol-labeled RNA sequencing (SLAM-seq). We injected 4-thiouridine (4sU) into mouse amygdala followed by SLAM-seq to detect nascent mRNA induced by acute and chronic restraint stress, and found that SLAM-seq could label actively transcribed genes in the major neuronal and glial subtypes. We also found that acute stress induced active transcription of 6 gamma-aminobutyric acid (GABA) receptors and only 1 glutamate receptor, indicating an imminent increase of inhibitory control in the stressed amygdala. Conversely, chronic stress led to active transcription of 3 glutamate receptors and 4 GABA receptors, suggesting a release of inhibitory control and hyperactivity of the amygdala. SLAM-seq also identified genes associated with myelination under chronic stress, and this finding is confirmed by immunostaining showing increased myelination in chronically stressed amygdala. Additionally, genes detected by SLAM-seq and RNA-seq only partially overlapped, with SLAM-seq particularly sensitive to transcriptional changes in genes with high basal transcription levels. Thus, by applying SLAM-seq in vivo, we obtained a rich dataset of nascently transcribed genes in the amygdala under stress, and revealed distinct transcriptional dynamics associated with acute and chronic stress.

neuroscience↗

bioRxiv2024

Reprogramming neuroblastoma by diet-enhanced polyamine depletion

Neuroblastoma is a highly lethal childhood tumor derived from differentiation-arrested neural crest cells1,2. Like all cancers, its growth is fueled by metabolites obtained from either circulation or local biosynthesis3,4. Neuroblastomas depend on local polyamine biosynthesis, with the inhibitor difluoromethylornithine showing clinical activity5. Here we show that such inhibition can be augmented by dietary restriction of upstream amino acid substrates, leading to disruption of oncogenic protein translation, tumor differentiation, and profound survival gains in the TH-MYCN mouse model. Specifically, an arginine/proline-free diet decreases the polyamine precursor ornithine and augments tumor polyamine depletion by difluoromethylornithine. This polyamine depletion causes ribosome stalling, unexpectedly specifically at adenosine-ending codons. Such codons are selectively enriched in cell cycle genes and low in neuronal differentiation genes. Thus, impaired translation of these codons, induced by the diet-drug combination, favors a pro-differentiation proteome. These results suggest that the genes of specific cellular programs have evolved hallmark codon usage preferences that enable coherent translational rewiring in response to metabolic stresses, and that this process can be targeted to activate differentiation of pediatric cancers. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=192 SRC="FIGDIR/small/573662v1_ufig1.gif" ALT="Figure 1"> View larger version (64K): org.highwire.dtl.DTLVardef@1fdbaecorg.highwire.dtl.DTLVardef@fb03f4org.highwire.dtl.DTLVardef@1c0afa7org.highwire.dtl.DTLVardef@14c384f_HPS_FORMAT_FIGEXP M_FIG C_FIG Highlights- Extra-tumoral conversion of arginine feeds tumor ornithine via uptake from circulation in MYCN-neuroblastoma. - A proline and arginine free diet enhances pharmacological polyamine depletion via reduced ornithine substrate availability. - Polyamine depletion disrupts oncogenic translation to induce a pro-differentiation proteome causing neuroblast differentiation and prolonged survival in the TH-MYCN mouse model. - Genes of specific cellular programs have evolved codon usage preferences that enable coherent translational rewiring in response to metabolic stress, such as polyamine depletion.

cancer biology↗

bioRxiv2024

Systemic Dosing of Virus-derived Serpin Improves Survival and Immunothrombotic Damage in Murine Colitis

Inflammatory bowel disease (IBD) is potentially life-threatening, with risk of bleeding, clotting, infection, sepsis, cancer and toxic megacolon. Systemic and local immune and coagulation dysfunction increase IBD severity. Current treatments are partially effective, but there is no definitive cure. Serine protease cascades activate thrombotic, thrombolytic and complement pathways and are regulated by inhibitors, serpins. Viruses encode proteins evolved from endogenous central regulatory pathways. A purified secreted Myxomavirus-derived serpin, Serp-1, dosed as a systemic anti-inflammatory drug, has proven efficacy in vascular and inflammatory disorders. PEGylated Serp-1 protein (PEGSerp-1) has improved efficacy in lupus and SARS-CoV-2 models. We examined PEGSerp-1 treatment in a mouse Dextran Sodium Sulfate (DSS) colitis model. Prophylactic PEGSerp-1 significantly improved survival in acute severe 4-5% DSS colitis, reducing inflammation and crypt damage in acute 4-5% DSS induced colitis and when dosed as a chronic delayed treatment for recurrent 2% DSS colitis. PEGSerp-1 reduced iNOS+ M1 macrophage invasion, damage to crypt architecture and vascular inflammation with decreased uPAR, fXa, fibrinogen and complement activation. This work supports PEGSerp-1 as a tissue targeting serpin therapeutic.

molecular biology↗

bioRxiv2024

Spatial characterization of interface dermatitis in cutaneous lupus reveals novel chemokine ligand-receptor pairs that drive disease

BackgroundChemokines play critical roles in the recruitment and activation of immune cells in both homeostatic and pathologic conditions. Here, we examined chemokine ligand-receptor pairs to better understand the immunopathogenesis of cutaneous lupus erythematosus (CLE), a complex autoimmune connective tissue disorder. ObjectivesOur objectives were to (1) characterize the cellular and proteomic constitution of interface dermatitis in CLE using blister biopsies, (2) map chemokine:ligand receptor pairs that govern recruitment of immune cells to form interface dermatitis in CLE, and (3) perform unbiased analyses in tandem on different clinical subtypes to identify novel genes and proteins underlying discoid versus subacute CLE. MethodsWe used suction blister biopsies to measure cellular infiltrates with spectral flow cytometry in the interface dermatitis reaction, as well as 184 protein analytes in interstitial skin fluid using 96-plex immunoassay targeted proteomics. Flow and 96-plex immunoassay data concordantly demonstrated significant increases in T cells and antigen presenting cells (APCs). We also performed spatial transcriptomics and spatial proteomics of punch biopsies using digital spatial profiling (DSP) technology on CLE skin and healthy margin controls to examine discreet locations within the tissue. ResultsSpatial and 96-plex immunoassay data confirmed elevation of interferon (IFN) and IFN-inducible CXCR3 chemokine ligands. Comparing involved versus uninvolved keratinocytes in CLE samples revealed upregulation of essential inflammatory response genes in areas near interface dermatitis, including AIM2. 96-plex immunoassay data confirmed upregulation of Caspase 8, IL-18 which is the final product of AIM2 activation, and induced chemokines including CCL8 and CXCL6 in CLE lesional samples. Chemotaxis assays using PBMCs from healthy and CLE donors revealed that T cells are equally poised to respond to CXCR3 ligands, whereas CD14+CD16+ APC populations are more sensitive to CXCL6 via CXCR1 and CD14+ are more sensitive to CCL8 via CCR2. ConclusionsTaken together, our data map a pathway from keratinocyte injury to lymphocyte recruitment in CLE via AIM2-Casp8-IL-18-CXCL6/CXCR1 and CCL8/CCR2, and IFNG/IFNL1-CXCL9/CXCL11-CXCR3, and identify potential novel biomarkers of disease. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=168 SRC="FIGDIR/small/574422v2_ufig1.gif" ALT="Figure 1"> View larger version (46K): org.highwire.dtl.DTLVardef@1e86dbeorg.highwire.dtl.DTLVardef@7d0208org.highwire.dtl.DTLVardef@1108aa8org.highwire.dtl.DTLVardef@ff255f_HPS_FORMAT_FIGEXP M_FIG C_FIG Model of chemokine systems governing recruitment of immune cell subsets to form interface dermatitis in cutaneous lupus. A. Summary of fresh tissue blister biopsy studies demonstrating increases in HLADR+ cells in nonlesional lupus biopsies as assessed by flow cytometry, and increased CXCL6 and CCL8 as assessed by 96 plex immunoassay. Lesional lupus biopsies also demonstrated significant increases in T cells and CXCL9/10/11 production. B. Model of chemokine-directed formation of interface dermatitis in cutaneous lupus. 1. Whole transcriptome atlas (WTA) digital spatial profiling (DSP) revealed increased AIM2 in keratinocytes proximal to inflammation, which is reported to be induced by environmental triggers including UV light and toxins. We also noted increased Caspase 8 (Casp8) and IL18 at the protein level, which can be induced downstream of AIM2. Chemokines including CXCL6 and CCL8 can be induced downstream of IL18, explaining how CCL8 and CXCL6 might be induced. 2. Recruitment of myeloid cell first responders by CCL8 and CXCL6. CD14+CD16+ myeloid cells, which were recently described in nonlesional lupus skin, express CXCR1 and migrate towards CXCL6, whereas CD14+CD16-myeloid cells express more CCR2 and migrate towards CCL8. 3. The CXCR3 ligands CXCL9/10/11 are expressed by keratinocytes, but more strongly in CD45+ immune cell and T cell regions of interest (ROIs). 4. T cells express CXCR3 and migrate towards CXCL9 and to a greater extent CXCL11. 5. The recruited HLADR+ myeloid populations and T cells contribute to formation of interface dermatitis. Thus, we propose a model in which keratinocyte/myeloid crosstalk can reinforce chemokine systems to optimally recruit lymphocytes and other immune cells to form interface dermatitis. Created with Biorender.com. Plain language summaryLupus skin rashes arise during flares, after exposure to medications or sunlight, or in response to other triggers of inflammation. To understand how white blood cells enter the skin to cause these rashes, we used new technologies to look at proteins that attract them into the skin. We found proteins that are expressed by skin cells in lesions that can recruit specific types of white blood cells that are thought to be the initiators of skin rashes. Once in the skin, these and other white blood cells can make additional proteins that bring in more and more cells. We hope that our findings will be used to test new topical treatments for lupus and other autoimmune skin rashes.

immunology↗

bioRxiv2024

Deep Learning Analysis on Images of iPSC-derived Motor Neurons Carrying fALS-genetics Reveals Disease-Relevant Phenotypes

Amyotrophic lateral sclerosis (ALS) is a devastating condition with very limited treatment options. It is a heterogeneous disease with complex genetics and unclear etiology, making the discovery of disease-modifying interventions very challenging. To discover novel mechanisms underlying ALS, we leverage a unique platform that combines isogenic, induced pluripotent stem cell (iPSC)-derived models of disease-causing mutations with rich phenotyping via high-content imaging and deep learning models. We introduced eight mutations that cause familial ALS (fALS) into multiple donor iPSC lines, and differentiated them into motor neurons to create multiple isogenic pairs of healthy (wild-type) and sick (mutant) motor neurons. We collected extensive high-content imaging data and used machine learning (ML) to process the images, segment the cells, and learn phenotypes. Self-supervised ML was used to create a concise embedding that captured significant, ALS-relevant biological information in these images. We demonstrate that ML models trained on core cell morphology alone can accurately predict TDP-43 mislocalization, a known phenotypic feature related to ALS. In addition, we were able to impute RNA expression from these image embeddings, in a way that elucidates molecular differences between mutants and wild-type cells. Finally, predictors leveraging these embeddings are able to distinguish between mutant and wild-type both within and across donors, defining cellular, ML-derived disease models for diverse fALS mutations. These disease models are the foundation for a novel screening approach to discover disease-modifying targets for familial ALS.

neuroscience↗

bioRxiv2024

The Nrf2/Id2 Axis in Vascular Smooth Muscle Cells: Novel Insights into the Interplay between Vascular Calcification and Aging

OBJECTIVEVascular calcification (VC) significantly contributes to cardiovascular morbidity and mortality and escalates with age. However, effective pharmaceutical interventions are lacking, and the molecular mechanisms linking aging to VC remain elusive. This study explores the role of nuclear factor erythroid 2-related factor 2 (NRF2) in VC, specifically focusing on the interplay between vascular senescence and oxidative stress (OS). APPROACH AND RESULTSUsing a chronological aging mouse model, we noted a significant decline in the expression and activity of Nrf2 in the aortas of aged mice, coinciding with increased medial VC. Administering NRF2 activators effectively reduced this calcification. In the vascular smooth muscle cell (VSMC)-specific Nrf2 knockout (Nrf2SMCKO) models, created using adenine and Vitamin D, there was an increase in calcium deposition within the aortic medial layer, alongside heightened VSMC senescence and OS. Furthermore, NRF2 knockout exacerbated VC in aortic rings and primary VSMCs in high-phosphate conditions, while Nrf2 overexpression in VSMCs inhibited calcium deposition by alleviating cell senescence and OS. RNAseq analysis of the aortas from Nrf2SMCKO and control mice highlighted a significant downstream regulator of Inhibitor of DNA Binding 2 (ID2). Our results show that reduced NRF2 levels lead to increased VSMC senescence, characterized by heightened p16 expression and diminished ID2 expression. Inhibiting ID2 negated NRF2s protective effects against VSMC senescence and VC. CONCLUSIONThis study emphasizes the critical role of NRF2 dysfunction in the nexus of vascular senescence, OS, and VC. It proposes the NRF2-ID2 axis in VSMCs as a promising therapeutic target for reducing VC and mitigating age-related cardiovascular diseases. HighlightsO_LIDecline in NRF2 activity is linked to increased vascular calcification (VC) and aging. C_LIO_LIVSMC-specific Nrf2 knockout causes a remarkable exacerbation of arterial calcification. C_LIO_LINRF2 alleviates VC by inhibiting oxidative stress and VSMC senescence. C_LIO_LIID2 contributes to the protective role of NRF2 in VSMC senescence and VC. C_LI

molecular biology↗

bioRxiv2023

Quantification of cell phenotype transition manifolds with information geometry

Cell phenotype transition (CPT) is crucial in development and other biological processes. Advances in single-cell sequencing reveal that CPT dynamics are confined to low-dimensional manifolds, yet current methods cannot directly quantify these manifolds. We present SCIM (single cell information manifolds), a geometry-guided approach using information geometry. SCIM embeds single-cell gene expression profiles as Gaussian distributions, defining the Fisher metric naturally in this space. We compute each cells coarse Ricci curvature, finding that low-curvature cells mark critical transitions. By further calculating each cells information velocity from RNA velocity, we find that regions with high information velocity coincide with low curvature, suggesting that geometry guides cellular dynamics on CPT manifolds. SCIM uncovers invariant features of CPT manifolds and provides a general framework for quantifying dynamics on these manifolds.

biophysics↗

bioRxiv2023

Auditory High Entropy Response (A-HER): A Novel Paradigm for Studying Brain Processing of Uncertain Information

This paper introduces a novel experimental paradigm - Auditory High Entropy Response (A-HER), which maximizes the information entropy of auditory stimulus sequences. This allows us to study how the brain processes complex information, rather than isolated individual events. Our analysis of the frequency response of the frontal theta rhythm induced by A-HER indicated a significant increase in signal-to-noise ratio and repeatability compared to zero-entropy Auditory Steady-State Response (A-SSR) and low-entropy mismatch negativity (MMN). We further investigated whether the A-HER response was induced by stimulus sequence differences or uncertainty, and studied its propagation rules. Different principles between evoked and entrained were found in A-HER and A-SSR. In conclusion, the A-HER paradigm, by maximizing stimulus sequence uncertainty, offers a new approach to analyzing how the brain processes uncertain information. It has potential for diagnosing and researching neurological and mental diseases, and for brain-computer interfaces, thus potentially impacting neuroscience, cognitive science, and psychology.

bioengineering↗

bioRxiv2023

Genomic evidence for hybridization and introgression between blue peafowl and green peafowl and selection for white plumage

The blue peafowl (Pavo cristatus) and the green peafowl (Pavo muticus) have significant public affection due to their stunning appearance, although the green peafowl is currently endangered. Some studies have suggested introgression between these the two species, although evidence is mixed. In this study, we successfully assembled a high-quality chromosome-level reference genome of the blue peafowl, including the autosomes, Z and W sex chromosomes as well as a complete mitochondria DNA sequence. Data from 77 peafowl whole genomes, 76 peafowl mitochondrial genomes and 33 peahen W chromosomes genomes provide the first substantial genetic evidence for recent hybridization between green and blue peafowl. We found three hybrid green peafowls in zoo samples rather than in the wild samples, with blue peafowl genomic content of 16-34%. Maternal genetic analysis showed that two of the hybrid female green peafowls contained complete blue peafowl mitochondrial genomes and W chromosomes. Hybridization of endangered species with its relatives is extremely detrimental to conservation. Some animal protection agencies release captive green peafowls in order to maintain the wild population of green peafowls. Therefore, in order to better protect the endangered green peafowl, we suggest that purebred identification must be carried out before releasing green peafowls from zoos into the wild in order to preventing the hybrid green peafowl from contaminating the wild green peafowl. In addition, we also found that there were historical introgression events of green peafowl to blue peafowl in four Zoo blue peafowl individuals. The introgressed genomic regions contain IGFBP1 and IGFBP2 genes that could affect blue peafowl body size. Finally, we identified that the nonsense mutation (g.4:12583552G>A) in the EDNRB2 gene is the genetic causative mutation for white feather color of blue peafowl (also called white peafowl), which prevents melanocytes from being transported into feathers, such that melanin cannot be deposited.

genomics↗

bioRxiv2023

EryDB: a transcriptomic profile database for erythropoiesis and erythroid-related diseases

Erythropoiesis is a finely regulated and complex process that involves multiple transformations from hematopoietic stem cells to mature red blood cells at hematopoietic sites from the embryonic to the adult stages. Investigations into its molecular mechanisms have generated a wealth of expression data, including bulk and single-cell RNA sequencing data. A comprehensively integrated and well-curated erythropoiesis-specific database will greatly facilitate the mining of gene expression data and enable large-scale research of erythropoiesis and erythroid related diseases. Here, we present EryDB, an open-access and comprehensive database dedicated to the collection, integration, analysis, and visualization of transcriptomic data for erythropoiesis and erythroid-related diseases. Currently, the database includes expertly curated quality-assured data of 3,803 samples and 1,187,119 single cells derived from 107 public studies of three species (Homo sapiens, Mus musculus, and Danio rerio), nine tissue types, and five diseases. EryDB provides users with the ability to not only browse the molecular features of erythropoiesis between tissues and species, but also perform computational analyses of single-cell and bulk RNA sequencing data, thus serving as a convenient platform for customized queries and analyses. EryDB v1.0 is freely accessible at https://ngdc.cncb.ac.cn/EryDB/home.

bioinformatics↗

bioRxiv2023

Phosphorylated transcription factor PuHB40 is involved in ROS-dependent anthocyanin biosynthesis in pear exposed to high-light stress

As sessile organisms, plants are increasingly vulnerable to environmental stresses because of global warming and climate change. Stress-induced reactive oxygen species (ROS) accumulation results in plant cell damages and even cell death. Anthocyanins are important antioxidants that scavenge ROS to maintain ROS homeostasis. However, the mechanism underlying ROS-induced anthocyanin accumulation is unclear. In this study, we determined that in pear the HD-Zip I family member PuHB40 mediates ROS-dependent anthocyanin biosynthesis under high-light stress. Specifically, PuHB40 is a transcription factor that induces PuMYB123-like/PubHLH3 complex for anthocyanin biosynthesis. The transcriptional activation by PuHB40 depends on its phosphorylation level, which is regulated by protein phosphatase 2A (PP2A). High ROS contents maintain the phosphorylation of PuHB40 at a high level, while also enhancing PuHB40-induced PuMYB123-like transcription by decreasing the transcription of PuPP2AA2, ultimately leading to increased anthocyanin biosynthesis. Our study revealed the pathway regulating ROS-induced anthocyanin biosynthesis in pear, further clarifying the mechanism underlying abiotic stress-induced anthocyanin biosynthesis, which may have implications for improving plant stress tolerance. IN A NUTSHELLO_ST_ABSBackgroundC_ST_ABSVarious abiotic stresses, including high-light intensity, promote the accumulation of anthocyanins in plants, while also activating the production of reactive oxygen species (ROS). Anthocyanins can attenuate the negative effects of high-light stress by acting as antioxidants that restrict ROS accumulation. Several reports have shown that ROS can stimulate anthocyanin accumulation, but whether high-light stress-induced anthocyanin accumulation depends on ROS is undetermined. Additionally, the mechanism underlying ROS-dependent anthocyanin biosynthesis remains unclear. QuestionDoes high-light stress-induced anthocyanin biosynthesis depend on ROS? What is the molecular basis of high-light stress-induced anthocyanin biosynthesis? FindingsHigh-light stress-induced anthocyanin biosynthesis in pear seedlings is dependent on ROS accumulation. PuMYB123-like is the key MYB transcription factor for anthocyanin biosynthesis, while PuHB40 activates anthocyanin biosynthesis in response to ROS under high-light stress. Specifically, PuHB40 activates the transcription of PuMYB123-like, with the encoded protein combining with PubHLH3 to form an MBW complex that promotes anthocyanin biosynthesis. The transcriptional activation by PuHB40 depends on its phosphorylation status, which is regulated by protein phosphatase 2A (PP2A). High ROS levels inhibit the transcription of PuPP2AA2, thereby maintaining the phosphorylation of PuHB40, enhancing the transcriptional activation by PuHB40, inducing PuMYB123-like transcription, and ultimately leading to increased anthocyanin biosynthesis. Next stepsOur future research will focus on whether ROS and the MYB123-like- PuHB40-PP2A regulatory module are also involved in the anthocyanin biosynthesis induced by other abiotic and biotic stresses, which will provide insights into biotic stress-induced anthocyanin biosynthesis and form the theoretical basis for improving fruit coloration.

plant biology↗

bioRxiv2023

Connecting Hippo pathway and cytoophidia in Drosophila posterior follicle cells

CTP synthase (CTPS), the rate-limiting enzyme in de novo synthesis of CTP, assembles into filamentous structures termed cytoophidia. Here we study the impact of Hippo pathway on the posterior follicle cells (PFCs) in Drosophila egg chambers. We find that the inactivation of Hippo pathway correlates with a reduction in cytoophidium length and number within PFCs. During the overexpression of CTPS, the presence of Hippo mutations also reduces the length of cytoophidia in PFCs. In addition, we observe that knocking down CTPS mitigates hpo (Hippo)-associated over-proliferation. In summary, our results suggest a connection between the Hippo pathway and the nucleotide biosynthesis enzyme CTPS in PFCs.

genetics↗

bioRxiv2023

Imaging the voltage of neurons distributed across entire brains of larval zebrafish

Neurons interact in networks distributed throughout the brain. Although much effort has focused on whole-brain calcium imaging, recent advances in genetically encoded voltage indicators (GEVIs) raise the possibility of imaging voltage of neurons distributed across brains. To achieve this, a microscope must image at high volumetric rate and signal-to-noise ratio. We present a remote scanning light-sheet microscope capable of imaging GEVI-expressing neurons distributed throughout entire brains of larval zebrafish at a volumetric rate of 200.8 Hz. We measured voltage of [~]1/3 of the neurons of the brain, distributed throughout. We observed that neurons firing at different times during a sequence were located at different brain locations, for sequences elicited by a visual stimulus, which mapped onto locations throughout the optic tectum, as well as during stimulus-independent bursts, which mapped onto locations in the cerebellum and medulla. Whole-brain voltage imaging may open up frontiers in the fundamental operation of neural systems.

neuroscience↗

bioRxiv2023

CCT3 drives Sorafenib resistance by inhibiting TFRC-mediated iron uptake in HCC

Sorafenib is commonly utilized in the management of advanced hepatocellular carcinoma (HCC). However, its efficacy in extending patients survival is hindered by the development of drug resistance. By employing protein posttranslational modification (PTM) omics, including acetylome, phosphoproteome, and ubiquitinome, in conjunction with genome-wide CRISPR/Cas9 knockout library screening, we have successfully identified chaperonin containing TCP1 subunit 3 (CCT3) as a key factor contributing to Sorafenib resistance. Furthermore, we observed a reduction in the ubiquitination of CCT3 at lysine 21 (K21) subsequent to Sorafenib treatment. This study provides evidence that CCT3 hinders the recycling of transferrin receptor protein 1 (TFRC) by interacting with alpha-actinin-4 (ACTN4), which is influenced by K6-linked ubiquitination on K21. Depleting CCT3 increased the susceptibility of cells to Sorafenib-induced ferroptosis, while reintroducing CCT3 through transfection restored resistance to ferroptosis. Additionally, impairing ACTN4 or TFRC depletion compromised CCT3s ability to inhibit Sorafenib-induced ferroptosis. In summary, targeting CCT3 presents a potential strategy for overcoming Sorafenib resistance in HCC.

cancer biology↗

bioRxiv2023

Selected humanization of yeast U1 snRNP leads to global suppression of pre-mRNA splicing and mitochondrial dysfunction in the budding yeast

The recognition of 5 splice site (5 ss) is one of the earliest steps of pre-mRNA splicing. To better understand the mechanism and regulation of 5 ss recognition, we selectively humanized components of the yeast U1 snRNP to reveal the function of these components in 5 ss recognition and splicing. We targeted U1C and Luc7, two proteins that interact with and stabilize the yeast U1 (yU1) snRNA and the 5 ss RNA duplex. We replaced the Zinc-Finger (ZnF) domain of yU1C with its human counterpart, which resulted in cold-sensitive growth phenotype and moderate splicing defects. Next, we added an auxin-inducible degron to yLuc7 protein and found that Luc7-depleted yU1 snRNP resulted in the concomitant loss of PRP40 and Snu71 (two other essential yeast U1 snRNP proteins), and further biochemical analyses suggest a model of how these three proteins interact with each other in the U1 snRNP. The loss of these proteins resulted in a significant growth retardation accompanied by a global suppression of pre-mRNA splicing. The splicing suppression led to mitochondrial dysfunction as revealed by a release of Fe2+ into the growth medium and an induction of mitochondrial reactive oxygen species. Together, these observations indicate that the human U1C ZnF can substitute that of yeast, Luc7 is essential for the incorporation of the Luc7-Prp40-Snu71 trimer into yeast U1 snRNP, and splicing plays a major role in the regulation of mitochondria function in yeast.

biochemistry↗

bioRxiv2023

Towards establishing a rapid constant temperature detection method for canine parvovirus based on endonuclease activities

Canine parvovirus can cause a high morbidity and mortality rate in puppies, posing a significant threat to both pet dogs and the breeding industry. Rapid, accurate and convenient detection methods are important for the early intervention and treatment of canine parvovirus. In this study, we propose a visual CPV detection system called nucleic acid mismatch enzyme digestion (NMED). This system combines reverse transcription-loop mediated isothermal amplification (RT-LAMP), endonuclease for gene mismatch detection and colloidal gold lateral chromatography. We demonstrated that NMED can induce the binding of the amplicon from the sample to the specific labeling probe, which in turn triggers digestion by the endonuclease. The sensitivity and visual visibility of RT-LAMP were increased by combining of endonuclease and colloidal gold lateral chromatography assisted by a simple temperature-controlled device. The sensitivity of the NMED assay was 1 copy/L, which was consistent with qPCR. The method was validated with 10 clinical samples that potentially had CPV infection and achieved 100% accuracy. As a rapid, accurate and convenient molecular diagnostic method, NMED has great potential for application in the field of pathogenic microorganism detection.

molecular biology↗