Biology subjects

Zou, L.

Publications and source records attributed to Zou, L..

67 records · Page 4Linked to original sources

bioRxiv2020

Selective germline genome edited pigs and their long immune tolerance in Non Human Primates

Organ transplantation is the only curative treatment for patients with terminal organ failure, however, there is a worldwide organ shortage. Genetically modified pig organs and tissues have become an attractive and practical alternative solution for the severe organ shortage, which has been made possible by significant progress in xenotransplantation in recent years. The past several decades witnessed an expanding list of genetically engineered pigs due to technology advancements, however, the necessary combination of genetic modifications in pig for human organ xenotransplantation has not been determined. In the current study, we created a selective germline genome edited pig (SGGEP). The first triple xenoantigens (GGTA, B4GAL, and CAMH) knockout somatic cells were generated to serve as a prototype cells and then human proteins were expressed in the xenoantigen knockout cells, which include human complement system negative regulatory proteins (CD46, CD55, and CD59); human coagulation system negative regulatory proteins thrombomodulin (THBD); tissue factor pathway inhibitor (TFPI); CD39; macrophage negative regulatory proteins (human CD47); and natural killer cell negative regulatory human HLA-E. After the successful establishment of SGGEP by the nuclear tranfer, we engrafted SGGEP skin to NHP, up to 25 days graft survival without immunosuppressive drugs was observed. Because a pig skin graft does not impact the success of a subsequent allograft or autograft or vice versa, thus our SGGEP could have a great potential for clinical value to save severe and large area burn patients and the other human organ failure. Therefore, this combination of specific gene modifications is a major milestone and provides proof of concept to initiate investigator-initiated clinical trials (IITs) in severe burn patients with defined processes and governance measures in place and the other clinical application.

synthetic biology↗

bioRxiv2019

Human glial progenitor cells effectively remyelinate the demyelinated adult brain

Human glial progenitor cells (hGPCs) can completely myelinate the brains of congenitally hypomyelinated shiverer mice, rescuing the phenotype and extending or normalizing the lifespan of these mice. We asked if implanted hGPCs might be similarly able to broadly disperse and remyelinate the diffusely and/or multicentrically-demyelinated adult CNS. In particular, we asked if fetal hGPCs could effectively remyelinate both congenitally hypomyelinated adult axons, and axons acutely demyelinated in adulthood, using adult shiverer mice and cuprizone-demyelinated mice, respectively. We found that hGPCs broadly infiltrate the adult CNS after callosal injection, and robustly myelinate congenitally-unmyelinated axons in adult shiverer. Moreover, implanted hGPCs similarly remyelinated denuded axons after cuprizone demyelination, whether they were delivered prior to or after initial cuprizone demyelination. Extraction and FACS of hGPCs from cuprizone-demyelinated brains in which they had been resident, followed by RNA-seq of the isolated human hGPCs, revealed their activation of transcriptional programs indicating their initiation of oligodendrocyte differentiation and myelination. These data indicate the ability of transplanted hGPCs to disperse throughout the adult CNS, to myelinate dysmyelinated regions encountered during their parenchymal colonization, and to also be recruited as myelinating oligodendrocytes at later points in life, upon demyelination-associated demand.

neuroscience↗

bioRxiv2019

Bacterial metabolism rescues the inhibition of intestinal drug absorption by food and drug additives

Food and drugs contain diverse small molecule additives (excipients) with unclear impacts on human physiology. Here, we evaluate their potential impact on intestinal absorption, screening 136 unique compounds for inhibition of the key transporter OATP2B1. We identified and validated 24 potent OATP2B1 transport inhibitors, characterized by higher molecular weight and hydrophobicity compared to poor or non-inhibitors. OATP2B1 inhibitors were also enriched for dyes, including 8 azo (R-N=N-R') dyes. Pharmacokinetic studies in mice confirmed that FD&C Red No. 40, a common azo dye excipient, inhibited drug absorption; however, the human gut microbiome inactivated azo dye excipients, producing metabolites that no longer inhibit OATP2B1 transport. These results support a beneficial role for the microbiome in limiting the unintended effects of food and drug additives in the intestine. One Sentence SummaryFood and drug additives inhibit intestinal drug transporters, although some are inactivated by gut bacterial metabolism.

pharmacology and toxicology↗

bioRxiv2019

Crif1 Promotes Osteoporosis in Mice after Radiation

Radiation induces rapid bone loss and enhances bone resorption and RANKL expression. RANKL provides the crucial signal to induce osteoclast differentiation and plays an important role in bone resorption. However, the mechanisms of radiation-induced osteoporosis are not fully understood. Here, we show that Crif1 expression increases in bone marrow cells after radiation. Conditional Crif1 deletion in bone marrow cells causes decreases in RANKL expression and the RANKL/OPG ratio, and relieves bone loss after radiation in mice. We further demonstrated in vitro that Crif1 promotes RANKL secretion via the cAMP/PKA pathway. Moreover, protein-protein docking screening identified five compounds as Crif1 inhibitors; these compounds dramatically suppressed RANKL secretion and CREB phosphorylation when cells were exposed to forskolin. This study enriches current knowledge of the pathogenesis of osteoporosis and provides insights into potential therapeutic strategies for osteoporosis treatment.

cell biology↗

bioRxiv2019

Bacterial fucosidase enables the production of Bombay red blood cells

We present a method to produce H antigen-deficient red blood cells (RBCs) for transfusion to individuals with anti-H antibodies. A fucosidase from bacteria was heterologously expressed efficiently in E. coli and has been demonstrated to completely remove H antigens on the surface of human RBCs in a facile conversion process. The approach we describe here holds promise for making H-deficient RBCs available for a rare population beyond ABO types.

molecular biology↗

bioRxiv2019

Unraveling the functional role of the orphan solute carrier, SLC22A24 in the transport of steroid conjugates through metabolomic and genome-wide association studies

Variation in sex hormone levels has wide implications for health and disease. The genes encoding the proteins involved in steroid disposition represent key determinants of interindividual variation in steroid levels and ultimately, their effects. Beginning with metabolomic data from genome-wide association studies (GWAS), we observed that genetic variants in the orphan transporter, SLC22A24 were significantly associated with levels of androsterone glucuronide and etiocholanolone glucuronide (sentinel SNPs p-value <1x10-30). In cells over-expressing human or various mammalian orthologs of SLC22A24, we showed that steroid conjugates and bile acids were substrates of the transporter. Phylogenetic, genomic, and transcriptomic analyses suggested that SLC22A24 has a specialized role in the kidney and appears to function in the reabsorption of organic anions, and in particular, anionic steroids. Phenome-wide analysis showed that functional variants of SLC22A24 are associated with human disease such as cardiovascular diseases and acne, which have been linked to dysregulated steroid metabolism. Collectively, these functional genomic studies reveal a previously uncharacterized protein involved in steroid homeostasis, opening up new possibilities for SLC22A24 as a pharmacological target for regulating steroid levels.\n\nAuthor SummarySteroid hormones, ranging from sex steroids such as testosterone to glucocorticoids play key roles in human health and disease. Accordingly, the identification of the genes and proteins involved in their synthesis, disposition and elimination has been the subject of numerous genetic studies. We have been intrigued by recent studies demonstrating that genetic variants in or near a gene encoding SLC22A24 are strongly associated with steroid levels. SLC22A24 is an orphan transporter with no known ligands and no known biological functions. In this study, we use cellular and computational methods to show that SLC22A24 transports steroid conjugates, bile acids and other dicarboxylic acids. Based on the direction of association of a common stop codon in SLC22A24 with lower levels of steroids, our studies suggest that the transporter functions to reabsorb steroid conjugates in the kidney, a surprising finding, given that conjugation pathways generally function to polar molecules that are readily eliminated by the kidney. The absence of the transporter gene in many species and its presence in higher order primates suggest that SLC22A24 plays a specialized role in steroid homeostasis. Overall, our studies indicate that SLC22A24 functions in the reabsorption of conjugated steroids in the kidney.

pharmacology and toxicology↗

bioRxiv2019

Co-regulation and functional cooperativity of FOXM1 and RHNO1 bidirectional genes in ovarian cancer

We report that the oncogenic transcription factor FOXM1 is arranged in a head-to-head configuration with RHNO1, a gene involved in the ATR/CHK1-dependent DNA replication stress (DRS) response. FOXM1 and RHNO1 are both amplified and upregulated in high-grade serous ovarian cancer (HGSC). FOXM1 and RHNO1 expression are closely associated in normal and cancer tissues, including single cells, and a bidirectional promoter (F/R-BDP) mediates balanced expression. Targeting of FOXM1 and RHNO1 in HGSC cells using shRNA, CRISPR mutagenesis, or CRISPR interference directed to the F/R-BDP reduced DNA homologous recombination repair (HR) capacity, increased DNA damage, reduced clonogenic survival, and sensitized HGSC cells to the poly-ADP ribosylase inhibitor (PARPi) olaparib. Thus, there is functional cooperativity between FOXM1 and RHNO1 in cancer cells, and combinatorial targeting of this bidirectional gene pair may be a novel cancer therapeutic strategy. More broadly, our data provide evidence that bidirectional gene units function in human cancer.

cancer biology↗

bioRxiv2019

TMC1 confers a leak conductance to modulate excitability of auditory hair cells in mammals

Hearing sensation relies on the mechano-electrical transducer (MET) channel of cochlear hair cells, in which Transmembrane channel-like 1 (TMC1) and TMC2 have been proposed to be the pore-forming subunits. Meanwhile it has been reported that TMCs regulate other biological processes in a variety of lower organisms ranging from sensations to motor functions. However, it is still an open question whether TMCs play roles other than their function in MET in mammals. In this study, we report that in mouse hair cells TMC1, but not TMC2, provides a background leak conductance, with properties distinct from those of the MET channels. By cysteine substitution, 4 amino acids of TMC1 are characterized critical for the leak conductance. The leak conductance is essential for action potential firing and tonotopic along the cochlear coil. Taken together, our results suggest that TMC1 confers a background leak conductance that modulates membrane excitability in cochlear hair cells.

cell biology↗

bioRxiv2019

Streamlined low-input transcriptomics through EASY-RNAseq

High-throughput sequencing for transcriptome profiling is an increasingly accessible and important tool for biological research. However, accurate profiling of small cell populations remains challenging due to issues with gene detection sensitivity and experimental complexity. Here we describe a streamlined RNAseq protocol (EASY RNAseq) for sensitive transcriptome assessment starting from low amount of input materials. EASY RNAseq is technically robust enough for sequencing small pools of homogenous and heterogeneous cells, recovering higher numbers of genes and with a more even expression distribution pattern than other commonly used methods. Application of EASY RNAseq to single human embryos at the 8-cell stage was able to achieve detection of 70% protein-coding genes. This workflow may thus serve as a useful tool for sensitive interrogation of rare cell populations.

genomics↗

bioRxiv2019

Translational regulation of non-autonomous mitochondrial stress response promotes longevity

Inhibition of mRNA translation delays aging, but the underlying mechanisms remain underexplored. Mutations in both DAF-2 (IGF-1 receptor) and RSKS-1 (ribosomal S6 kinase/S6K) cause synergistic lifespan extension in C. elegans. To understand the roles of S6K-mediated translational regulation in this process, we performed genome-wide translational profiling and genetic screens to identify genes that are not only regulated at the translational level in the daf-2 rsks-1 mutant, but also affect lifespan. Inhibition of CYC-2.1 (cytochrome c) in the germline significantly extends lifespan through non-autonomous activation of the mitochondrial unfolded protein response (UPRmt) and AMP-activated kinase (AMPK) in the metabolic tissue. Furthermore, the RNA-binding protein GLD-1-mediated translational repression of cytochrome c in the germline is important for the non-autonomous activation of UPRmt and synergistic longevity of the daf-2 rsks-1 mutant. Together, these results illustrate a translationally regulated non-autonomous mitochondrial stress response mechanism in the modulation of lifespan by insulin-like signaling and S6K. HighlightsO_LILongevity of the daf-2 rsks-1 mutant is mediated by translational repression of ribosomal proteins and CYC-2.1/cytochrome c. C_LIO_LIGermline inhibition of cyc-2.1 non-autonomously activates UPRmt and AMPK to extend lifespan. C_LIO_LIGLD-1 represses germline cyc-2.1 translation in the daf-2 rsks-1 mutant. C_LIO_LITranslational regulation of cyc-2.1 and UPRmt contribute to the synergistic longevity of the daf-2 rsks-1 mutant. C_LI O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/533695v1_ufig1.gif" ALT="Figure 1"> View larger version (32K): org.highwire.dtl.DTLVardef@9f3ce7org.highwire.dtl.DTLVardef@576930org.highwire.dtl.DTLVardef@bb0c09org.highwire.dtl.DTLVardef@133ce4_HPS_FORMAT_FIGEXP M_FIG C_FIG

developmental biology↗

bioRxiv2018

Construction of a germline-specific RNAi tool in C. elegans

Analysis of complex biological functions usually requires tissue-specific genetic manipulations in multicellular organisms. The C. elegans germline plays regulatory roles not only in reproduction, but also in metabolism, stress response and aging. Previous studies have used mutants of rrf-1, which encodes an RNA-directed RNA polymerase, as a germline-specific RNAi tool. However, the rrf-1 mutants showed RNAi efficiency in somatic tissues. Here we constructed a germline-specific RNAi strain by combining an indel mutation of rde-1, which encodes an Argonaute protein that functions cell autonomously to ensure RNAi efficiency, and a single-copy rde-1 transgene driven by the sun-1 germlinespecific promoter. The germline RNAi efficiency and specificity are confirmed by RNAi phenocopy of known mutations, knockdown of GFP reporter expression, as well as quantitative RT-PCR measurement of tissue-specific mRNAs upon RNAi knockdown. The germline-specific RNAi strain shows no obvious deficiencies in reproduction, lipid accumulation, thermo-tolerance and life span compared to wild-type animals. By screening an RNAi sub-library of phosphatase genes, we identified novel regulators of thermo-tolerance. Together, we have created a useful tool that can facilitate the genetic analysis of germline-specific functions in C. elegans.

genetics↗

bioRxiv2018

Dietary lysozyme supplement alters serum biochemical makers and milk metabolite profile of sows via gut microbiota

Lysozyme is an important antimicrobial agent with promising future in replacing antibiotics in livestok production. The aim of current study was to determine variations in sows gut microbiota, serum immunity and breast milk metabolite profile mediated by lysozyme supplementation.Thirty-six pregnant sows were assigned to a control group without supplementation and two treatments with 0.5 g/kg and 1.0 g/kg lysozyme provided in formula feed for 21days. Microbiota analysis based on 16s RNA high-throughput sequencing and untargeted liquid chromatography tandem mass spectrometry were applied and combined in analysis. Serum biochemical indicators and immunoglobulins were also determined. Sows received 1.0kg/t lyszoyme treatment shown significant redution in microbial diversity. Spirochaetes, Euryarchaeota and Actinobacteria significantly increased while Firmicutes showed a remarkable reduction in 1.0kg/t treated group compared with control. Pyrimidine metabolism,Purine metabolism and Amino acid related enzymes were significantly upregulated in 1.0kg/t lysozyme treated group. The richness of gram-positive bacteria were significantly down-regulated by lysozyme treatments. Serum aspartate transaminase (AST) activity was significantly un-regulated. Serum IgM levels were significantly higher in the 1.0 kg/t group compared with control, while IgA levels was significantly lower in 1.0kg/t group. Over thirty metabolites from sows breast milk including L-Glutamine,creatine and L-Arginine were sigficantly altered by lysozyme treatment. There existed crucial correlations among gut microbiota, serum immunity and breast milk metabolites where lactobacillus and prevotella may play a key role in lysozyme mediated host-microbial interactions. Overall, lysozyme supplementation could effectively improve the composition, metabolic functions and phenotypes of sows gut microbiota and it also benefit sows with better immune status and breast milk composition.\n\nImportanceEnteric infections caused by pathogens have a significant negative effect on neonatal survival and animal health in swine production. The application of antibiotics in feeds at subtherapeutic levels could improve performance and overall health and is used extensively throughout the industry. However, abuse of antibiotics is contributing to the high level of drug resistance in microbial communities and rising concerns regarding human health. Here, we revealed that lysozyme supplementation could effectively improve the composition, metabolic functions and phenotypes of sows gut microbiota and it also benefit sows with better immune status and breast milk composition. These findings confirmed that lysozyme could be a suitable alternative to antibiotics in swine production.

microbiology↗

bioRxiv2017

Molecular evolution, diversity and adaptation of H7N9 influenza A viruses in China

A novel H7N9 avian influenza virus has caused five human epidemics in China since 2013. The substantial increase in prevalence and the emergence of antigenically divergent or highly pathogenic (HP) H7N9 strains during the current outbreak raises concerns about the epizootic-potential of these viruses. Here, we investigate the evolution and adaptation of H7N9 by combining publicly available data with newly generated virus sequences isolated in Guangdong between 2015-2017. Phylogenetic analyses show that currently-circulating H7N9 viruses belong to distinct lineages with differing spatial distributions. Using ancestral sequence reconstruction and structural modelling we have identified parallel amino-acid changes on multiple separate lineages. Furthermore, we infer mutations in HA primarily occur at sites involved in receptor-recognition and/or antigenicity. We also identify seven new HP strains, which likely emerged from viruses circulating in eastern Guangdong around March 2016 and is further associated with a high rate of adaptive molecular evolution.

evolutionary biology↗