Biology subjects

Zhou, B.

Publications and source records attributed to Zhou, B..

At least 145 records · Page 8Linked to original sources

bioRxiv2021

SARS-CoV-2 hijacks neutralizing dimeric IgA for enhanced nasal infection and injury

Robust severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection in nasal turbinate (NT) accounts for high viral transmissibility, yet whether neutralizing IgA antibodies can control it remains unknown. Here, we evaluated receptor binding domain (RBD)-specific monomeric B8-mIgA1 and B8-mIgA2, and dimeric B8-dIgA1 and B8-dIgA2 against intranasal SARS-CoV-2 challenge in Syrian hamsters. These antibodies exhibited comparably potent neutralization against authentic virus by competing with human angiotensin converting enzyme-2 (ACE2) receptor for RBD binding. While reducing viruses in lungs, pre-exposure intranasal B8-dIgA1 or B8-dIgA2 led to 81-fold more infectious viruses and severer damage in NT than placebo. Virus-bound B8-dIgA1 and B8-dIgA2 could engage CD209 as an alternative receptor for entry into ACE2-negative cells and allowed viral cell-to-cell transmission. Cryo-EM revealed B8 as a class II neutralizing antibody binding trimeric RBDs in 3-up or 2-up/1-down conformation. Therefore, RBD-specific neutralizing dIgA engages an unexpected action for enhanced SARS-CoV-2 nasal infection and injury in Syrian hamsters.

microbiology↗

bioRxiv2021

Serum glycoprotein markers in non-alcoholic steatohepatitis and hepatocellular carcinoma

Fatty liver disease progresses through stages of fat accumulation and inflammation to non-alcoholic steatohepatitis (NASH), fibrosis and cirrhosis and eventually hepatocellular carcinoma (HCC). Currently available diagnostic tools for HCC lack sensitivity and specificity and deliver little value to patients. In this study, we investigated the use of circulating serum glycoproteins to identify a panel of potential prognostic markers that may be indicative of progression from the healthy state to NASH and further to HCC. Serum samples were processed using a standard pre-analytical sample preparation protocol and were analyzed using a novel high throughput glycoproteomics platform. We analyzed 413 glycopeptides, representing 57 abundant serum proteins and compared among the three phenotypes. Our initial dataset contained healthy, NASH, and HCC serum samples. We analyzed normalized abundance of common glycoforms and found 40 glycopeptides with statistically significant differences in abundances in NASH and HCC compared to controls. Summary level relative abundance of core-fucosylated, sialylated and branched glycans containing glycopeptides were higher in NASH and HCC as compared to controls. We replicated some of our findings in an independent set of samples of individuals with benign liver conditions and HCC, respectively. Our results may be of value in the management of liver diseases. TOC only O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=95 SRC="FIGDIR/small/462486v2_ufig1.gif" ALT="Figure 1"> View larger version (14K): org.highwire.dtl.DTLVardef@91672corg.highwire.dtl.DTLVardef@cad49org.highwire.dtl.DTLVardef@4496b9org.highwire.dtl.DTLVardef@103615c_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2021

Two Ubiquitin-Activating Systems Occur in Plants with One Playing a Major Role in Plant Immunity

Many plants possess two or more ubiquitin-activating enzymes (E1). However, it is unclear whether the E1s of a plant genome play equivalent roles in various pathways. Here we report that tomato and tobacco encode dual ubiquitin-activating systems (DUAS) in which the E1s UBA1 and UBA2 display differential specificities in charging four groups of E2s. The C-terminal ubiquitin-folding domain of the E1s play a major but not sole role in determining the differential specificities of charging the four groups E2s. The dual systems do not play equivalent roles in plant immunity, with silence of UBA2 only compromising host immunity. Among the differentially charged E2s, group IV members UBC32, UBC33 and UBC34 are shown to be essential for ER-associated protein degradation (ERAD) and plant immunity. Like tomato, Arabidopsis UBC32/33/34 E2 triplet are also differentially charged by its E1s and are essential for plant immunity. Loss of function in Arabidopsis UBC32, UBC33 and UBC34 does not affect flg22 and elf18-triggered inhibition of seedling growth but results in alteration of ER stress tolerance, which likely contribute to the diminished plant immunity in the mutants. Our results uncover DUAS in plants and a previously unknown E1-ERAD-associated E2 triplet module in the regulation of host immunity. One sentence summaryPlant dual ubiquitin E1 systems play distinct roles in plant immunity by differentially charging the ERAD-associated E2s for ER stress tolerance.

plant biology↗

bioRxiv2021

Dynamics of poly(A) tail length and non-A residues during the human oocyte-to-embryo transition

Poly(A) tail-mediated post-transcriptional regulation of maternal mRNA has been shown to be vital in the oocyte-to-embryo transition (OET) in flies, fish, frogs, and mice1-8. However, nothing is known about poly(A) tail dynamics for even a single gene during the human OET, because of the limited availability of human oocytes and embryos in combination with the low sensitivity of previous methods. Here, we systematically profiled the transcriptome-wide mRNA poly(A) tails in human oocytes at the germ-vesicle (GV), metaphase I (MI), and metaphase II (MII) stages, as well as pre-implantation embryos at the 1-cell (1C), 2-cell (2C), 4-cell (4C), 8-cell (8C), morula (MO), and blastocyst (BL) stages using single-oocyte/embryo PAIso-seq1 and PAIso-seq2 methods. We show that poly(A) tail length is highly dynamic during the OET, with BTG4 responsible for global deadenylation. Moreover, we reveal that non-A residues occur primarily in poly(A) tails of maternal RNA, which begin to increase at the MI stage, become highly abundant after fertilization (with U residues occurring in about two thirds, G residues in about one third, and C residues in about one fifth of mRNAs), and decline at the 8C stage. Importantly, we reveal that TUT4/7 can add U residues to deadenylated mRNA, which can then be re- polyadenylated to produce 5'-end and internal U residues. In addition, the re- polyadenylated mRNA can be stabilized through the addition of G residues by TENT4A/B. Finally, we demonstrate that U residues in poly(A) tails mark the maternal transcripts for quicker degradation in 8C human embryos compared to those without U residues. Together, our results not only reveal the dynamics of poly(A) tail length and non-A residues, but also provide mechanistic insights into the regulation of the length and the role of non-A residues during human OET. These findings further scientific understanding and open a new door for studying the human OET.

developmental biology↗

bioRxiv2021

The genome of stress tolerant crop wild relativePaspalum vaginatum leads to increased biomassproductivity in the crop Zea mays

A number of crop wild relatives can tolerate extreme stressed to a degree outside the range observed in their domesticated relatives. However, it is unclear whether or how the molecular mechanisms employed by these species can be translated to domesticated crops. Paspalum Paspalum vaginatum is a self-incompatible and multiply stress-tolerant wild relative of maize and sorghum. Here we describe the sequencing and pseudomolecule level assembly of a vegetatively propagated accession of P. vaginatum. Phylogenetic analysis based on 6,151 single-copy syntenic orthologous conserved in 6 related grass species placed paspalum as an outgroup of the maize-sorghum clade demonstrating paspalum as their closest sequenced wild relative. In parallel metabolic experiments, paspalum, but neither maize nor sorghum, exhibited significant increases in trehalose when grown under nutrient-deficit conditions. Inducing trehalose accumulation in maize, imitating the metabolic phenotype of paspalum, resulting in autophagy dependent increases in biomass accumulation.

plant biology↗

bioRxiv2021

Mitochondrial interactome quantitation reveals structural changes in metabolic machinery in failing murine heart

Advancements of cross-linking mass spectrometry (XL-MS) for structural analysis of proteins bridges the gap between purified systems and native tissue environments. Here, isobaric quantitative protein interaction reporter technology (iqPIR) was utilized to further extend XL-MS to the first system-wide comparative study of mitochondrial proteins from healthy and diseased murine hearts. The failing heart interactome includes 602 statistically significant cross-linked peptide pairs altered in the disease condition. Structural insight into ketone oxidation metabolons, OXPHOS machinery, and nucleotide transporter hybrid-conformations, support mitochondrial remodeling in failing heart while bringing forth new hypotheses for pathological mechanisms. Application of quantitative cross-linking technology in tissue provides molecular-level insight to complex biological systems difficult to model in cell culture, thus providing a valuable resource for study of human diseases.

systems biology↗

bioRxiv2021

STAT3 expression in dendritic cells protects mice from colitis by a gut microbiome-dependent mechanism

An imbalance in gut homeostasis results in local and systemic pathogenesis. It is still not well-understood how the immune system interacts with the gut microbiome and maintains a delicate balance. Here, we utilized a mouse model in which STAT3 expression is deleted in CD11c+ (i.e., dendritic) cells (STAT3 cKO); these mice developed an ulcerative colitis-like disease, colon carcinoma and myelodysplastic syndrome-like disease. Circulating IgE levels in STAT3 cKO mice were significantly elevated. The gut microbiome was indispensable for the observed pathogenesis, as treatment with broad-spectrum antibiotics or cross-fostering STAT3 cKO pups with mothers harboring a different microbiome prevented disease development. Gut microbiome analyses suggested that decreased commensal bacteria and increased pathogenic bacteria most likely contributed to disease. Our data suggest that STAT3 controls the manifestation of inflammation in the gut caused by the microbiome. Therefore, we conclude that a deficiency of STAT3 in DCs is sufficient to trigger uncontrolled inflammation and the development of inflammatory bowel disease.

immunology↗

bioRxiv2021

N-glycosylation profiles of the SARS-CoV-2 spike D614G mutant and its ancestral protein characterized by advanced mass spectrometry

N-glycosylation plays an important role in the structure and function of membrane and secreted proteins. The spike protein on the surface of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the virus that causes COVID-19, is heavily glycosylated and the major target for developing vaccines, therapeutic drugs and diagnostic tests. The first major SARS-CoV-2 variant carries a D614G substitution in the spike (S-D614G) that has been associated with altered conformation, enhanced ACE2 binding, and increased infectivity and transmission. In this report, we used mass spectrometry techniques to characterize and compare the N-glycosylation of the wild type (S-614D) or variant (S-614G) SARS-CoV-2 spike glycoproteins prepared under identical conditions. The data showed that half of the N-glycosylation sequons changed their distribution of glycans in the S-614G variant. The S-614G variant showed a decrease in the relative abundance of complex-type glycans (up to 45%) and an increase in oligomannose glycans (up to 33%) on all altered sequons. These changes led to a reduction in the overall complexity of the total N-glycosylation profile. All the glycosylation sites with altered patterns were in the spike head while the glycosylation of three sites in the stalk remained unchanged between S-614G and S-614D proteins.

biochemistry↗

bioRxiv2021

Discovery of nanobodies against SARS-CoV-2 and an uncommon neutralizing mechanism

SARS-CoV-2 and its variants continue to threaten public health. The virus recognizes the host cell by attaching its Spike receptor-binding domain (RBD) to the host receptor ACE2. Therefore, RBD is a primary target for neutralizing antibodies and vaccines. Here we report the isolation, and biological and structural characterization of two single-chain antibodies (nanobodies, DL4 and DL28) from RBD-immunized alpaca. Both nanobodies bind Spike with affinities that exceeded the detection limit (picomolar) of the biolayer interferometry assay and neutralize the original SARS-CoV- 2 strain with IC50 of 0.086 g mL-1 (DL4) and 0.385 g mL-1 (DL28). DL4 and a more potent, rationally designed mutant, neutralizes the Alpha variant as potently as the original strain but only displays marginal activity against the Beta variant. By contrast, the neutralizing activity of DL28, when in the Fc-fused divalent form, was less affected by the mutations in the Beta variant (IC50 of 0.414 g mL-1 for Alpha, 1.060 g mL-1 for Beta). Crystal structure studies reveal that DL4 blocks ACE2-binding by direct competition, while DL28 neutralizes SARS-CoV-2 by an uncommon mechanism through which DL28 distorts the receptor-binding motif in RBD and hence prevents ACE2-binding. Our work provides two neutralizing nanobodies for potential therapeutic development and reveals an uncommon mechanism to design and screen novel neutralizing antibodies against SARS-CoV-2.

biochemistry↗

bioRxiv2021

Shallow neural networks trained to detect collisions recover features of visual loom-selective neurons

Animals have evolved sophisticated visual circuits to solve a vital inference problem: detecting whether or not a visual signal corresponds to an object on a collision course. Such events are detected by specific circuits sensitive to visual looming, or objects increasing in size. Various computational models have been developed for these circuits, but how the collision-detection inference problem itself shapes the computational structures of these circuits remains unknown. Here, inspired by the distinctive structures of LPLC2 neurons in the visual system of Drosophila, we build an anatomically-constrained shallow neural network model and train it to identify visual signals that correspond to impending collisions. Surprisingly, the optimization arrives at two distinct, opposing solutions, only one of which matches the actual dendritic weighting of LPLC2 neurons. The LPLC2-like solutions are favored when a population of units is trained on the task, but not when units are trained in isolation. The trained model reproduces experimentally observed LPLC2 neuron responses for many stimuli, and reproduces canonical tuning of loom sensitive neurons, even though the models are never trained on neural data. These results show that LPLC2 neuron properties and tuning are predicted by optimizing an anatomically-constrained neural network to detect impending collisions.

neuroscience↗

bioRxiv2021

Emerging single cell endothelial heterogeneity supports sprouting tumour angiogenesis and growth

Blood vessels supplying tumors are often dysfunctional and generally heterogeneous. The mechanisms underlying this heterogeneity remain poorly understood. Here, using multicolor lineage tracing, in vivo time-lapse imaging and single cell RNA sequencing in a mouse glioma model, we identify tumour-specific blood endothelial cells that originate from cells expressing the receptor for colony stimulating factor 1, Csf1r, a cytokine which controls macrophage biology. These Csf1r lineage endothelial cells (CLECs) form up to 10% of the tumour vasculature and express, besides classical blood endothelial cell markers, a gene signature that is distinct from brain endothelium but shares similarities with lymphatic endothelial cell populations. in silico analysis of pan-cancer single cell RNAseq datasets highlights the presence of a comparable subpopulation in the endothelium of a wide spectrum of human tumours. We show that CLECs actively contribute to sprouting and remodeling of tumour blood vessels and that selective depletion of CLECs reduces vascular branching and tumour growth. Our findings indicate that a non-tumour resident Csf1r-positive population is recruited to tumours, differentiates into blood endothelial cells to contribute to vascularization and, thereby, tumour growth.

cancer biology↗

bioRxiv2021

Immunogenicity and In vivo protection of a variant nanoparticle vaccine that confers broad protection against emerging SARS-CoV-2 variants

The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) continues to spread globally. As SARS-CoV-2 has transmitted from person to person, variant viruses have emerged with elevated transmission rates and higher risk of infection for vaccinees. We present data showing that a recombinant prefusion-stabilized Spike (rS) protein based on the B.1.351 sequence (rS-B.1.351) was highly immunogenic in mice and produced neutralizing antibodies against SARS-CoV-2/WA1, B.1.1.7, and B.1.351. Mice vaccinated with our prototype vaccine NVX-CoV2373 (rS-WU1) or rS-B.1.351 alone, in combination, or as a heterologous prime boost, were protected when challenged with live SARS-CoV-2/B.1.1.7 or SARS-CoV-2/B.1.351. Virus titer was reduced to undetectable levels in the lungs post-challenge in all vaccinated mice, and Th1-skewed cellular responses were observed. A strong anamnestic response was demonstrated in baboons boosted with rS-B.1.351 approximately one year after immunization with NVX-CoV2373 (rS-WU1). An rS-B.1.351 vaccine alone or in combination with prototype rS-WU1 induced protective antibody- and cell-mediated responses that were protective against challenge with SARS-CoV-2 variant viruses.

immunology↗

bioRxiv2021

Combination Respiratory Vaccine Containing Recombinant SARS-CoV-2 Spike and QuadrivalentSeasonal Influenza Hemagglutinin Nanoparticles with Matrix-M Adjuvant

The 2019 outbreak of a severe respiratory disease caused by an emerging coronavirus, SARS-CoV-2, has spread globally with high morbidity and mortality. Co-circulating seasonal influenza has greatly diminished recently, but expected to return with novel strains emerging, thus requiring annual strain adjustments. We have developed a recombinant hemagglutinin (HA) quadrivalent nanoparticle influenza vaccine (qNIV) produced using an established recombinant insect cell expression system to produce nanoparticles. Influenza qNIV adjuvanted with Matrix-M was well-tolerated and induced robust antibody and cellular responses, notably against both homologous and drifted A/H3N2 viruses in Phase 1, 2, and 3 trials. We also developed a full-length SARS-CoV-2 spike protein vaccine which is stable in the prefusion conformation (NVX-CoV2373) using the same platform technology. In phase 3 clinical trials, NVX-CoV2373 is highly immunogenic and protective against the prototype strain and B.1.1.7 variant. Here we describe the immunogenicity and efficacy of a combination quadrivalent seasonal flu and COVID-19 vaccine (qNIV/CoV2373) in ferret and hamster models. The combination qNIV/CoV2373 vaccine produces high titer influenza hemagglutination inhibiting (HAI) and neutralizing antibodies against influenza A and B strains. The combination vaccine also elicited antibodies that block SARS-CoV-2 spike protein binding to the human angiotensin converting enzyme-2 (hACE2) receptor. Significantly, hamsters immunized with qNIV/CoV2373 vaccine and challenged with SARS-CoV-2 were protected against weight loss and were free of replicating SARS-CoV-2 in the upper and lower respiratory tract with no evidence of viral pneumonia. This study supports evaluation of qNIV/CoV2373 combination vaccine as a preventive measure for seasonal influenza and CoVID-19. HighlightsO_LICombination qNIV/CoV2373 vaccine induced protective hemagglutination inhibition (HAI) responses to seasonal influenza A and B unchanged when formulated with recombinant spike. C_LIO_LICombination qNIV/CoV2373 vaccine maintained clinical and virologic protection against experimental challenge with SARS-CoV-2. C_LIO_LICombination qNIV/CoV2373 vaccine showed no clinical or histological sign of enhanced disease following experimental challenge with SARS-CoV-2. C_LIO_LICombination qNIV/CoV2373 vaccine induced antibodies against SARS-CoV-2 neutralizing epitopes common between US-WA and B.1.352 variant. C_LI

immunology↗

bioRxiv2021

Comprehensive epigenomic profiling of human alveolar epithelial differentiation identifies key epigenetic states and transcription factor co-regulatory networks for maintenance of distal lung identity

Disruption of alveolar epithelial cell (AEC) differentiation is implicated in peripheral lung diseases strongly impacting morbidity and mortality worldwide, such as chronic obstructive pulmonary disease, idiopathic pulmonary fibrosis, and lung adenocarcinoma. Elucidating underlying disease pathogenesis requires a mechanistic molecular understanding of AEC differentiation. However, to date no study has comprehensively characterized the dynamic epigenomic alterations that facilitate this critical process in humans. We comprehensively profiled the epigenomic states of human AECs during type 2 to type 1-like cell differentiation, including the methylome and chromatin functional domains, and integrated this with transcriptome-wide RNA expression. Enhancer regions were drastically altered during AEC differentiation. Transcription factor binding analysis within enhancer regions revealed diverse interactive networks with enrichment for dozens of transcription factors, including NKX2-1 and FOXA family members, as well as transcription factors with previously uncharacterized roles in lung differentiation, such as members of the MEF2, TEAD, and AP1 families. Additionally, associations between transcription factors changed during differentiation, implicating a complex network of heterotrimeric complex switching may be involved in facilitating differentiation. Integration of AEC enhancer states with the catalog of enhancer elements in the Roadmap Epigenomics Mapping Consortium and Encyclopedia of DNA Elements (ENCODE) revealed that human mammary epithelial cells (HMEC) have a similar epigenomic structure to alveolar epithelium, with NKX2-1 serving as a distinguishing feature of distal lung differentiation. Taken together, our results suggest that enhancer regions with dynamic transcription factor interactions are hotspots of epigenomic alteration that help to facilitate AEC differentiation. Author SummaryHuman health and disease states are heavily influenced by the critical cellular processes that regulate and protect our genomes. One of these safeguards is the epigenome; the coordinated set of signals overlaid on top of our DNA that controls what can happen to a given stretch DNA. Hence, epigenomic signatures play a critical role in the development and maintenance of cellular fate and function. To determine the relationship between epigenomic alterations and cellular fates of distal lung cells in humans during the process that regenerates the human lung epithelial layer after injury, we performed comprehensive genome-wide profiling of many epigenetic modifications that have roles in regulating the function of the underlying DNA. We found that changes to enhancer regions, which act to turn on associated gene expression, were the major alterations to the epigenome during distal lung differentiation, and that within those regions dynamic changes in transcription factor associations were occurring to facilitate this process. We then characterize what was similar and distinct to the enhancers of distal lung from among other epithelial tissues and describe a novel role for specific transcription factors in this process that previously had no known role in normal lung repair.

genomics↗

bioRxiv2021

A specialized bone marrow microenvironment for fetal haematopoiesis

Local signals provided by cells in specialized niche microenvironments regulate stem cell behaviour in many different organs and species. In adult mammalian bone marrow (BM), leptin receptor-positive (LepR+) reticular cells express secreted factors that control the function of haematopoietic stem and progenitor cells (HSPCs). During fetal development, the developing skeletal system is colonized by c-Kit+ haematopoietic cells de novo after a transient phase of liver haematopoiesis. The cellular and molecular mechanisms regulating de novo haematopoietic cell colonization and expansion remain largely unknown. Here, we report that fetal and adult BM exhibit fundamental differences both in terms of cellular composition and molecular interactions by single cell RNA sequencing (scRNA-seq) analysis. While LepR+ reticular cells are almost completely absent in fetal femur, arterial endothelial cells (AECs) are a source of signals controlling the initial HSPC expansion during BM development. Long-term haematopoietic stem cells (HSCs) and other c-Kit+ HSPCs are reduced when Wnt ligand secretion by AECs is genetically blocked. We identify Wnt2 as an AEC-derived signal that directly activates {beta}-catenin dependent proliferation of fetal HSPCs. Treatment of HSPCs ex vivo with Wnt2 promotes their proliferation and improves engraftment in vivo after transplantation. Our work reveals a fundamental switch in the cellular organization and molecular regulation of BM niches in the embryonic and adult organism.

developmental biology↗

bioRxiv2021

Collaboration between the Fab and Fc contribute to maximal protection against SARS-CoV-2 in nonhuman primates following NVX-CoV2373 subunit vaccine with Matrix-M™ vaccination

Recently approved vaccines have already shown remarkable protection in limiting SARS-CoV-2 associated disease. However, immunologic mechanism(s) of protection, as well as how boosting alters immunity to wildtype and newly emerging strains, remain incompletely understood. Here we deeply profiled the humoral immune response in a cohort of non-human primates immunized with a stable recombinant full-length SARS-CoV-2 spike (S) glycoprotein (NVX-CoV2373) at two dose levels, administered as a single or two-dose regimen with a saponin-based adjuvant Matrix-M. While antigen dose had some effect on Fc-effector profiles, both antigen dose and boosting significantly altered overall titers, neutralization and Fc-effector profiles, driving unique vaccine-induced antibody fingerprints. Combined differences in antibody effector functions and neutralization were strongly associated with distinct levels of protection in the upper and lower respiratory tract, pointing to the presence of combined, but distinct, compartment-specific neutralization and Fc-mechanisms as key determinants of protective immunity against infection. Moreover, NVX-CoV2373 elicited antibodies functionally target emerging SARS-CoV-2 variants, collectively pointing to the critical collaborative role for Fab and Fc in driving maximal protection against SARS-CoV-2. Collectively, the data presented here suggest that a single dose may prevent disease, but that two doses may be essential to block further transmission of SARS-CoV-2 and emerging variants. HighlightsO_LINVX-CoV2373 subunit vaccine elicits receptor blocking, virus neutralizing antibodies, and Fc-effector functional antibodies. C_LIO_LIThe vaccine protects against respiratory tract infection and virus shedding in non-human primates (NHPs). C_LIO_LIBoth neutralizing and Fc-effector functions contribute to protection, potentially through different mechanisms in the upper and lower respiratory tract. C_LIO_LIBoth macaque and human vaccine-induced antibodies exhibit altered Fc-receptor binding to emerging mutants. C_LI

immunology↗

bioRxiv2021

SARS-CoV-2 susceptibility of cell lines and substrates commonly used in diagnosis and isolation of influenza and other viruses

Coinfection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and other viruses is inevitable as the COVID-19 pandemic continues. This study aimed to evaluate cell lines commonly used in virus diagnosis and isolation for their susceptibility to SARS-CoV-2. While multiple kidney cell lines from monkeys were susceptible and permissive to SARS-CoV-2, many cell types derived from human, dog, mink, cat, mouse, or chicken were not. Analysis of MDCK cells, which are most commonly used for surveillance and study of influenza viruses, demonstrated that they were insusceptible to SARS-CoV-2 and that the cellular barrier to productive infection was due to low expression level of the angiotensin converting enzyme 2 (ACE2) receptor and lower receptor affinity to SARS-CoV-2 spike, which could be overcome by over-expression of canine ACE2 in trans. Moreover, SARS-CoV-2 cell tropism did not appear to be affected by a D614G mutation in the spike protein.

microbiology↗

bioRxiv2020

Development of novel in vitro human alveolar epithelial cell models to study distal lung biology and disease

Many acute and chronic lung diseases affect the distal lung alveoli. Although airway-derived human cell lines exist, alveolar epithelial cell (AEC)-derived lines are needed to better model these diseases. We have generated and characterized novel immortalized cell lines derived from human AECs. They grow as epithelial monolayers expressing lung progenitor markers SOX9 and SOX2, with little to no expression of mature AEC markers. Co-cultured in 3-dimensions (3D) with lung fibroblasts, the cells form NKX2-1+ organoids expressing mature AEC markers AQP5 and GPRC5A. Single-cell RNA sequencing of an AEC line in 2D versus 3D revealed increased cellular heterogeneity and induction of cytokine and lipoprotein signaling, consistent with organoid formation. Activating WNT and FGF pathways resulted in larger organoids. Our approach appears to yield lung progenitor lines that retain a genetic and structural memory of their alveolar cell lineage despite long-term expansion and whose differentiation may be modulated under various 3D conditions. These cell lines provide a valuable new system to model the distal lung in vitro.

cell biology↗