Biology subjects

Si, L.

Publications and source records attributed to Si, L..

13 recordsLinked to original sources

bioRxiv2024

Architecture of the ATP-driven motor for protein import into chloroplasts

Thousands of nuclear-encoded proteins are transported into chloroplasts through the TOC-TIC translocon spanning the chloroplast envelope membranes. A motor complex pulls the translocated proteins out of the TOC-TIC complex into the chloroplast stroma by hydrolyzing ATP. The Orf2971-FtsHi complex was suggested to serve as the ATP-hydrolyzing motor in Chlamydomonas reinhardtii, but little is known about its architecture and assembly. Here, we report the 3.2-[A] resolution structure of the Chlamydomonas Orf2971-FtsHi complex. The 20-subunit complex spans the chloroplast inner envelope with two bulky modules protruding into the intermembrane space and stromal matrix. Six subunits form a hetero-hexamer potentially providing the pulling force through ATP hydrolysis. The remaining subunits, including potential enzymes/chaperones, likely facilitate the complex assembly and regulate its proper function. Our results provide the structural foundation for mechanistic understanding of chloroplast protein translocation.

biochemistry↗

bioRxiv2024

Combining Scalable Organ Chip Platform with Deep Learning-Based Imaging Analysis for Cancer Therapeutic Screening

Functional precision oncology represents an emerging approach to genomic approaches by testing treatment options directly on patient-derived models. Current assays, including the use of patient-derived xenograft (PDX) and patient-derived organoid (PDOs), faces major barriers in clinical use due to technical challenges, such as standardization, cost, assay time, scalability, and faithful mimicry of patient tumor microenvironment (TME). Here, we introduce an Organ Chip (OC) device constructed entirely from thermoplastic materials, free of porous membrane or other barrier structures, and optimized for high-content imaging (HCI). This automation-compatible device supports tissue-specific extracellular matrices and coculture for a wide spectrum of organ and disease types, including the TME. As a proof-of-concept, we demonstrate the growth of pancreatic, lung, and colon cancer cell lines and primary lung cancer cells and the testing of cancer drugs in these models. HCI-based phenotypic profiling enabled accurate quantification of drug response, with better performance than traditional biochemical assays. Moreover, we developed a deep-learning method for assessing drug responses using bright field images. The integration of a low-cost, scalable, and faithful OC models with automatic high-content image analysis represents a significant stride towards functional precision oncology and cancer drug discovery.

bioengineering↗

bioRxiv2024

Growth phase-dependent reorganization of cryptophyte photosystem I antennae

Photosynthetic cryptophytes are eukaryotic algae that utilize membrane-embedded chlorophyll a/c binding proteins (CACs) and lumen-localized phycobiliproteins (PBPs) as their light-harvesting antennae. Cryptophytes go through logarithmic and stationary growth phases, and may adjust their light-harvesting capability according to their particular growth state. How cryptophytes change the type/arrangement of the photosynthetic antenna proteins to regulate their light-harvesting remains unknown. Here we solved four structures of cryptophyte photosystem I (PSI) bound with CACs that show the rearrangement of CACs at different growth phases. We identified a cryptophyte-unique protein, PsaQ, which harbors three chlorophyll molecules. PsaQ specifically binds to the lumenal region of PSI during logarithmic growth phase and may assist the association of PBPs with photosystems and energy transfer from PBPs to photosystems.

plant biology↗

bioRxiv2024

DNA origami vaccine (DoriVac) nanoparticles improve both humoral and cellular immune responses to infectious diseases

Current SARS-CoV-2 vaccines have demonstrated robust induction of neutralizing antibodies and CD4+ T cell activation, however CD8+ responses are variable, and the duration of immunity and protection against variants are limited. Here we repurposed our DNA origami vaccine nanotechnology, DoriVac, for targeting infectious viruses, namely SARS-CoV-2, HIV, and Ebola. The DNA origami nanoparticle, conjugated with infectious-disease-specific heptad repeat 2 (HR2) peptides, which act as highly conserved antigens, and CpG adjuvant at precise nanoscale spacing, induced neutralizing antibodies, Th1 CD4+ T cells, and CD8+ T cells in naive mice, with significant improvement over a bolus control. Pre-clinical studies using lymph-node-on-a-chip systems validated that DoriVac, when conjugated with antigenic peptides or proteins, induced promising cellular and humoral immune responses in human cells. Moreover, DoriVac bearing full-length SARS-CoV-2 spike protein achieved immune responses comparable to current mRNA vaccine platforms while potentially reducing storage constraints. These results suggest that DoriVac holds potential as a versatile, modular vaccine platform, capable of inducing both humoral and cellular immunities, underscoring its potential utility in addressing future pandemics.

immunology↗

bioRxiv2023

Rapid Prototyping of Thermoplastic Microfluidic 3D Cell Culture Devices by Creating Regional Hydrophilicity Discrepancy

Microfluidic three-dimensional cell culture devices that enable the recapitulation of key aspects of organ structures and functions in vivo represent a promising preclinical platform to improve translational success during drug discovery. Essential to these engineered devices is the spatial patterning of cells from different tissue types within a confined microenvironment. Traditional fabrication strategies lack the scalability, cost-effectiveness, and rapid prototyping capabilities required for industrial applications, especially for processes involving thermoplastic materials. Here, we introduce an approach to pattern fluid guides inside microchannels by establishing differential hydrophilicity using pressure-sensitive adhesives as masks and a subsequent selective coating with a biocompatible polymer. We identified optimal coating conditions using polyvinylpyrrolidone, which resulted in rapid and consistent hydrogel flow in both the open-chip prototype and the fully bonded device containing additional features for medium perfusion. We tested the suitability of our device for dynamic 3D cell culture by growing human hepatocytes in the device under controlled fluid flow for a 14-day period. Additionally, we demonstrated the potential of using our device for pharmaceutical high-throughput screening applications, such as predicting drug-induced liver injury. Our approach offers a facile strategy of rapid prototyping thermoplastic microfluidic organ chips with varying geometries, microstructures, and substrate materials.

bioengineering↗

bioRxiv2021

Self-assembling short immunostimulatory duplex RNAs with broad spectrum antiviral activity

The current COVID-19 pandemic highlights the need for broad-spectrum antiviral therapeutics. Here we describe a new class of self-assembling immunostimulatory short duplex RNAs that potently induce production of type I and type III interferon (IFN-I and IFN-III), in a wide range of human cell types. These RNAs require a minimum of 20 base pairs, lack any sequence or structural characteristics of known immunostimulatory RNAs, and instead require a unique conserved sequence motif (sense strand: 5-C, antisense strand: 3-GGG) that mediates end-to-end dimer self-assembly of these RNAs by Hoogsteen G-G base-pairing. The presence of terminal hydroxyl or monophosphate groups, blunt or overhanging ends, or terminal RNA or DNA bases did not affect their ability to induce IFN. Unlike previously described immunostimulatory siRNAs, their activity is independent of TLR7/8, but requires the RIG-I/IRF3 pathway that induces a more restricted antiviral response with a lower proinflammatory signature compared with poly(I:C). Immune stimulation mediated by these duplex RNAs results in broad spectrum inhibition of infections by many respiratory viruses with pandemic potential, including SARS-CoV-2, SARS-CoV, MERS-CoV, and influenza A, as well as the common cold virus HCoV-NL63 in both cell lines and human Lung Chips that mimic organ-level lung pathophysiology. These short dsRNAs can be manufactured easily, and thus potentially could be harnessed to produce broad-spectrum antiviral therapeutics at low cost.

immunology↗

bioRxiv2021

Mechanical control of innate immune responses against viral infection revealed in a human Lung Alveolus Chip

Mechanical forces associated with breathing play a fundamental role in lung development and disease but the molecular pathways remain largely unknown. Here, we used a mechanically actuatable Human Lung Alveolus Chip that recapitulates human lung alveolar type I and type II cell differentiation, alveolar-capillary interface formation, and genome-wide gene expression profiles characteristic of the distal lung to investigate the role of physical forces associated with cyclic breathing motions in lung innate immune responses to viral infection. When the mechanically active Alveolus Chips are infected with the influenza H3N2 virus, a cascade of host responses is elicited on-chip, including increased production of cytokines and expression of inflammation-associated genes in pulmonary epithelial and endothelial cells, resulting in enhanced recruitment of circulating immune cells as occurs during viral infection in vivo. Surprisingly, studies carried out in parallel with static chips revealed that physiological breathing motions suppress viral replication by activating protective innate immune responses in epithelial and endothelial cells. This is mediated at least in part through upregulation of S100 calcium-binding protein A7 (S100A7), which binds to the Receptor for Advanced Glycation End Products (RAGE), an inflammatory mediator that is most highly expressed in the lung alveolus in vivo. This mechano-immunological control mechanism is further supported by the finding that existing RAGE inhibitor drugs can suppress the production of inflammatory cytokines in response to influenza virus infection in this model. S100A7-RAGE interactions and modulation of mechanical ventilation parameters could therefore serve as new targets for therapeutic intervention in patients infected with influenza and other potential pandemic viruses that cause life-threatening lung inflammation.

immunology↗

bioRxiv2020

A novel viral protein translation mechanism reveals mitochondria as a target for antiviral drug development

The ongoing Severe Acute Respiratory Syndrome coronavirus 2 (SARS-CoV-2) pandemic has acutely highlighted the need to identify new treatment strategies for viral infections. Here we present a pivotal molecular mechanism of viral protein translation that relies on the mitochondrial translation machinery. We found that rare codons such as Leu-TTA are highly enriched in many viruses, including SARS-CoV-2, and these codons are essential for the regulation of viral protein expression. SARS-CoV-2 controls the translation of its spike gene by hijacking host mitochondria through 5 leader and 3UTR sequences that contain mitochondrial localization signals and activate the EGR1 pathway. Mitochondrial-targeted drugs such as lonidamine and polydatin significantly repress rare codon-driven gene expression and viral replication. This study identifies an unreported viral protein translation mechanism and opens up a novel avenue for developing antiviral drugs. One Sentence SummaryMitochondria are a potential target for antiviral therapy

microbiology↗

bioRxiv2020

In situ structure determination using single particle cryo-electron microscopy images

Cryo-electron microscopy (cryo-EM) tomography is a powerful tool for in situ structure determination. However, this method requires the acquisition of tilt series, and its time consuming throughput of acquiring tilt series severely slows determination of in situ structures. By treating the electron densities of non-target protein as non-Gaussian distributed noise, we developed a new target function that greatly improves the efficiency of the recognition of the target protein in a single cryo-EM image without acquiring tilt series. Moreover, we developed a sorting function that effectively eliminates the false positive detection, which not only improves the resolution during the subsequent structure refinement procedure but also allows using homolog proteins as models to recognize the target protein. Together, we developed an in situ single particle analysis (isSPA) method. Our isSPA method was successfully applied to solve structures of glycoproteins on the surface of a non-icosahedral virus and Rubisco inside the carboxysome. The cryo-EM data from both samples were collected within 24 hours, thus allowing fast and simple structural determination in situ.

biophysics↗

bioRxiv2020

Ratio of the interferon-γ signature to the immunosuppression signature predicts anti-PD-1 therapy response in melanoma

Immune checkpoint inhibitor (ICI) treatments produce clinical benefit in many patients. However, better pretreatment predictive biomarkers for ICI are still needed to help match individual patients to the treatment most likely to be of benefit. Existing gene expression profiling (GEP)-based biomarkers for ICI are primarily focused on measuring a T cell-inflamed tumour microenvironment that contributes positively to the response to ICI. Here, we identified an immunosuppression signature (IMS) through analysing RNA sequencing data from a combined discovery cohort (n = 120) consisting of three publicly available melanoma datasets. Using the ratio of an established IFN-{gamma} signature and IMS led to consistently better prediction of the ICI therapy outcome compared to a collection of nine published GEP signatures from the literature on a newly generated internal validation cohort (n = 55) and three published datasets of metastatic melanoma treated with anti-PD-1 (n = 48) and anti-CTLA-4 (n = 42) as well as in patients with gastric cancer treated with antiPD-1 (n = 45), demonstrating the potential utility of IMS as a predictive/prognostic biomarker that complements existing GEP signatures for immunotherapy.

immunology↗

bioRxiv2020

Human organs-on-chips as tools for repurposing approved drugs as potential influenza and COVID19 therapeutics in viral pandemics

The rising threat of pandemic viruses, such as SARS-CoV-2, requires development of new preclinical discovery platforms that can more rapidly identify therapeutics that are active in vitro and also translate in vivo. Here we show that human organ-on-a-chip (Organ Chip) microfluidic culture devices lined by highly differentiated human primary lung airway epithelium and endothelium can be used to model virus entry, replication, strain-dependent virulence, host cytokine production, and recruitment of circulating immune cells in response to infection by respiratory viruses with great pandemic potential. We provide a first demonstration of drug repurposing by using oseltamivir in influenza A virus-infected organ chip cultures and show that co-administration of the approved anticoagulant drug, nafamostat, can double oseltamivirs therapeutic time window. With the emergence of the COVID-19 pandemic, the Airway Chips were used to assess the inhibitory activities of approved drugs that showed inhibition in traditional cell culture assays only to find that most failed when tested in the Organ Chip platform. When administered in human Airway Chips under flow at a clinically relevant dose, one drug - amodiaquine - significantly inhibited infection by a pseudotyped SARS-CoV-2 virus. Proof of concept was provided by showing that amodiaquine and its active metabolite (desethylamodiaquine) also significantly reduce viral load in both direct infection and animal-to-animal transmission models of native SARS-CoV-2 infection in hamsters. These data highlight the value of Organ Chip technology as a more stringent and physiologically relevant platform for drug repurposing, and suggest that amodiaquine should be considered for future clinical testing.

microbiology↗

bioRxiv2019

Discovery of influenza drug resistance mutations and host therapeutic targets using a human airway chip

Here we demonstrate that influenza virus replication, host responses to infection, evolution through mutation or gene reassortment, and clinical efficacy of antiviral drugs can be reconstituted in a human Airway Chip microfluidic culture device. Modeling human-to-human transmission of infection in the continued presence of antiviral drugs on chips led to the emergence of clinically prevalent mutations responsible for amantadine- and oseltamivir-resistance, as well as the discovery of new resistance mutations. Analysis of infection responses resulted in identification of host therapeutic targets and demonstration that existing non-antiviral drugs may be repurposed to inhibit viral replication and synergize with antiviral therapeutics by targeting the host response to infection rather than the virus itself. This Influenza Chip may represent an alternative preclinical tool for development of new antiviral drugs and vaccines.\n\nOne Sentence SummaryNew drug resistance mutations and potential tolerance-inducing therapeutics were discovered using an organ chip model of influenza infection.

bioengineering↗

bioRxiv2019

Rapidly increasing prevalence of overweight and obesity in older Ghanaian adults from 2007-2015: evidence from WHO-SAGE Waves 1 & 2

BackgroundStudies on changes in the prevalence and determinants of obesity in older adults living in sub-Saharan Africa are limited. We examined recent changes in obesity prevalence and associated factors for older adults in Ghana between 2007/08 and 2014/15.\n\nMethodsData on adults aged 50 years and older in Ghana were collected in the WHO SAGE Wave 1 (2007/08; n=4158) and Wave 2 (2014/15; n=1663). The weighted prevalence of obesity, overweight, normal weight and underweight, and of high central adiposity was compared in Waves 1 and 2. Multinomial and binomial logistic regressions were used to examine whether the determinants of weight status based on objectively measured body mass index and waist circumference changed between the two survey periods.\n\nResultsThe prevalence of obesity (Wave 1=10.2%, 95% CI: 8.9-11.7%; Wave 2=15.0%, 95% CI: 12.6-17.7%) and overweight (Wave 1=19.6%, 95% CI: 18.0-21.4%; Wave 2=24.5%, 95% CI: 21.7-27.5%) was higher in Wave 2 than Wave 1 and more than half of the population had high central adiposity (Wave 1=57.7%, 95% CI: 55.4-60.1%; Wave 2=66.9%, 95% CI: 63.7-70.0%) in both study periods. Obesity prevalence was 16% lower in males and 55% higher in females comparing Wave 1 to Wave 2. Female sex, urban residence, and high household wealth were associated with higher odds of overweight/obesity and high central adiposity. Those aged 70+ years had lower odds of obesity in both study waves. In Wave 2, females with physical activity level were more likely to be obese.\n\nConclusionOver the 7-year period between survey waves, the population prevalence of overweight and obesity increased by 25% and 47%, respectively, while underweight reduced by 43%. These findings differed considerably by sex, which points to differential impacts of past initiatives to reduce overweight/obesity, potential high-risk groups in Ghana, and the need to increase surveillance.

epidemiology↗