Biology subjects

Gao, S.

Publications and source records attributed to Gao, S..

At least 235 records · Page 13Linked to original sources

bioRxiv2019

Synthetic cassettes for pH-mediated sensing, counting and containment

AbstractAs pH is fundamental to all biological processes, pH-responsive bacterial genetic circuits enable precise sensing in any environment. Where unintentional release of engineered bacteria poses a concern, coupling pH sensing to expression of a toxin creates an effective bacterial containment system. Here, we present a pH-sensitive kill switch (acidic Termination of Replicating Population; acidTRP), based on the E. coli asr promoter, with a survival ratio of less than 1 in 106. We integrate acidTRP with cryodeath to produce a two-factor containment system with a combined survival ratio of less than 1 in 1011 whilst maintaining evolutionary stability. We further develop a pulse-counting circuit with single cell readout for each administered stimulus pulse. We use this pulse-counter to record multiple pH changes and combine it with acidTRP to make a two-count acid-sensitive kill switch. These results demonstrate the ability to build complex genetic systems for biological containment.

synthetic biology↗

bioRxiv2019

Considering founding and variable genomes is critical in studying polyploid evolution

A wide range of differences between the subgenomes, termed as subgenome asymmetry or SA, has been reported in various polyploids and different species seem to have different responses to polyploidization. We compared subgenome differences in gene ratio and relative diversity between artificial and natural genotypes of several allopolyploid species. Surprisingly, consistent differences in neither gene ratio nor relative diversity between the subgenomes were detected between these two types of polyploid genotypes although they differ in times exposed to evolutional selection. As expected, the estimated ratio of retained genes between a subgenome and its diploid donor was invariably higher for the artificial allopolyploid genotypes due likely to the presence of variable genome components (VGC). Clearly, the presence of VGC means that exaggerated differences between a donor and a subgenome in a polyploid are inevitable when random genotypes were used to represent species of either a polyploid or its donors. SA was also detected in genotypes before the completion of the polyploidization events as well as in those which were not formed via polyploidization. Considering that significant changes during and following polyploidization have been detected in previous studies, our results suggest that the influence of VGC needs to be considered in evaluating SA and that diploid donors may define changes in polyploid evolution.

evolutionary biology↗

bioRxiv2019

Precise annotation of human, chimpanzee, rhesus macaque and mouse mitochondrial genomes using 5’ and 3’ end small RNAs

Using 5 and 3 end small RNAs, we annotated human, chimpanzee, rhesus macaque and mouse mitochondrial genomes at 1 base-pair (bp) resolution to cover both strands of the mammalian mitochondrial genome entirely without leaving any gaps or overlaps. The precise annotation of all coding and non-coding genes (e.g. ncMT1, MDL2 and MDL1AS) led to the discovery of novel functions and mechanisms of mitochondrion. In this study, we defined the conserved sequence block (CSB) region to span five CSBs (CSB1, CSB2, CSB3, LSP and HSP) and identified the motifs of five CSBs in the mitochondrial displacement loop (D-loop) regions of 52 mammals. The conserved arrangement of these five CSBs in 17 primates inspired us to investigate the function of the mtDNA D-loop, which has been puzzling scientists for more than 50 years. We found that 5 sRNAs of MDL1AS control the expression levels of mitochondrial genes as a whole by a negative feedback mechanism. Thus, the precise annotations of three CSBs (CSB2, LSP and HSP) in more species will help to understand the function of the mtDNA D-loop. The precision annotation of animal mitochondrial genomes also provides abundant information for studying the molecular phylogenetics and evolution of animals.

bioinformatics↗

bioRxiv2019

Diverse noncanonical PAMs recognized by SpCas9 in human cells

The CRISPR/Cas9 system derived from Streptococcus pyogenes (SpCas9) provides unprecedented genome editing capabilities, but the potential for off-target mutations limits its application. In addition to NGG protospacer adjacent motif (PAM), off-target mutations are also associated with noncanonical PAMs, which have not yet been systematically evaluated. Here, we developed a highly sensitive approach that allows systematically analyzing PAM sequences in human cells, and identified multiple alternative PAMs recognized by SpCas9.

bioengineering↗

bioRxiv2019

Practical and safe method of cryopreservation for clinical application of human adipose-derived mesenchymal stem cells without a programmable freezer or serum

BackgroundAdipose-derived mesenchymal stem cells (ADSCs) have emerged as a promising therapeutic modality for cellular therapy because of their rapid proliferation and potent cellular activity compared to conventional bone marrow-derived mesenchymal stem cells (MSCs). Cosmetic lipoaspirates provide an easily obtainable source of ADSCs. Cryopreservation facilitates their clinical application due to increased transportability and pooling of sufficient numbers of cells. However, proper cryopreservation techniques have not been established yet.\n\nMethodsWe evaluated the post-thaw viability and ADSC functions after cryopreservation with three cryoprotectants (serum containing 10% dimethylsulfoxide (DMSO), serum-free: CP-1TM, DMSO-free: SCB-DFTM) at two temperature (-80{degrees}C, -150{degrees}C) and two cell densities: (1 x 106, 7 x 106 cells/mL) for up to 18 months using cryovials. After determining optimal conditions, we also tested if large quantities of ADSCs remained viable after 18 months of cryopreservation in a 100-mL cryobag. Rate-controlled freezing methods or liquid nitrogen storage were not exploited.\n\nResultsADSCs cryopreserved in serum containing 10% DMSO or CP-1TM at -150{degrees}C and 7 x 106 cells/mL were most viable (>85%) after 18 months without perturbation of MSC functions. Even suboptimal conditions (-80{degrees}C, 1 x 106 cells/mL, no DMSO) assured >80% viability when stored for up to 9 months. Large quantities of ADSCs in a cryobag were properly cryopreserved.\n\nConclusionsA programmable freezer or liquid nitrogen storage is not necessary. CP-1TM is preferable in terms of side effects. Simplified cryopreservation methods (-80{degrees}C and no DMSO) can be used for up to 9 months, resulting in reduced infusion toxicities and lower costs.

cell biology↗

bioRxiv2019

Task-Based Functional Connectomes Predict Cognitive Phenotypes Across Psychiatric Disease

ImportanceWe show that three common approaches to clinical deficits (cognitive phenotype, disease group, disease severity) each offer useful and perhaps complimentary explanations for the brains underlying functional architecture as affected by psychiatric disease.\n\nObjectiveTo understand how different clinical frameworks are represented in the brains functional connectome.\n\nDesignWe use an openly available dataset to create predictive models based on multiple connectomes built from task-based functional MRI data. We use these models to predict individual traits corresponding to multiple cognitive constructs across disease category. We also show that these same connectomes statistically differ depending on disease category and symptom burden.\n\nSettingThis was a population-based study with data collected in UCLA.\n\nParticipantsHealthy adults were recruited by community advertisements from the Los Angeles area. Participants with adult ADHD, bipolar disorder, and schizophrenia were recruited using a patient-oriented strategy involving outreach to local clinics and online portals (separate from the methods used to recruit healthy volunteers)

neuroscience↗

bioRxiv2019

Validation and delineation of a locus conferring Fusarium crown rot resistance on 1HL in barley by analysing transcriptomes from multiple pairs of near isogenic lines

BackgroundFusarium crown rot (FCR) is a chronic and severe disease in cereal production in semi-arid regions worldwide. One of the putative quantitative trait locus (QTL) designated as Qcrs.cpi-1H has been previously mapped on chromosome arm 1HL in barley.\n\nResultsIn this study, five pairs of near-isogenic lines (NILs) targeting the 1HL locus were developed. Analysing the NILs found that the resistant allele at Qcrs.cpi-1H significantly reduced FCR severity. Transcriptomic analysis was then conducted against three of the NIL pairs, which placed the Qcrs.cpi-1H locus in an interval spanning about 11 Mbp. A total of 56 expressed genes bearing SNPs were detected in this interval, which would facilitate detailed mapping as well as cloning gene(s) underlying the resistance locus. Also, five differentially expressed genes (DEGs) bearing non-synonymous SNPs were identified in the interval. Differences in DEGs regulated by Qcrs.cpi-1H those by Qcrs.cpi-4H (another known locus conferring FCR resistance) indicate that different mechanisms could be involved in their resistance.\n\nConclusionNILs developed in this study and the transcriptomic sequences obtained from them did not only allow the validation of the resistance locus Qcrs.cpi-1H and the identification of candidate genes underlying its resistance, they also allowed the delineation of the resistance locus and the development of SNPs markers which formed a solid base for detailed mapping as well as cloning gene(s) underlying the locus.

genetics↗

bioRxiv2019

A novel QTL conferring Fusarium crown rot resistance located on chromosome arm 6HL in barley

Fusarium crown rot (FCR), caused primarily by Fusarium pseudograminearum, is a devastating disease for cereal production in semi-arid regions worldwide. To identify and characterize loci conferring FCR resistance, we assessed a landrace AWCS799 which is among the top lines identified from a screening. Genetic control of its resistance was investigated by generating and analysing two populations of recombinant inbred lines. One of the populations was used for QTL detection and the other for validation. A novel QTL, located on the long arm of chromosome 6H (designated as Qcrs.caf-6H), was consistently detected in each of the four trials conducted against the mapping population. The QTL explained up to 29.1% of the phenotypic variance and its effect was confirmed in the validation population. Significant interactions between this resistance locus and either plant height or heading date were not detected, further facilitating its manipulation in breeding programs. Key MessageThis study identified and validated a novel and large-effect QTL conferring Fusarium crown rot resistance on the long arm of chromosome 6HL in barley.

plant biology↗

bioRxiv2019

Gain-of-function mutations in the UNC-2/CaV2α channel lead to hyperactivity and excitation-dominant synaptic transmission in Caenorhabditis elegans

Mutations in pre-synaptic voltage gated calcium channels can lead to familial hemiplegic migraine type 1 (FHM1). While mammalian studies indicate that the migraine brain is hyperexcitable due to enhanced excitation or reduced inhibition, the molecular and cellular mechanisms underlying this excitatory/inhibitory (E/I) imbalance are poorly understood. We identified a gain-of-function (gf) mutation in the Caenorhabditis elegans CaV2 channel 1 subunit, UNC-2, which leads to increased calcium currents. unc-2(gf) mutants exhibit hyperactivity and seizure-like motor behaviors. Expression of the unc-2 gene with FHM1 substitutions R192Q and S218L leads to hyperactivity similar to that of unc-2(gf) mutants unc-2(gf) mutants display increased cholinergic- and decreased GABAergic-transmission. Moreover, we reveal that and increased cholinergic transmission in unc-2(gf) mutants leads to reduction of GABA synapses in a TAX-6/calcineurin dependent manner. Our studies provide mechanistic insight into how CaV2 gain-of-function mutations disrupt excitation-inhibition balance in the nervous system.

neuroscience↗

bioRxiv2018

Inhibition of tetrameric Patched1 by Sonic Hedgehog through an asymmetric paradigm

The Hedgehog (Hh) pathway controls embryonic development and postnatal tissue maintenance and regeneration. Inhibition of oligomeric Hh receptor Patched (Ptch) by the secreted and post-translationally modified ligand Hh relieves suppression of the signaling cascades. Here, we report the cryo-EM structure of tetrameric Ptch1 in complex with palmitoylated N-terminal signaling domain of human Sonic hedgehog (ShhNp). The structure shows that four Ptch1 protomers are organized as a loose dimer of dimers and each dimer binds to one ShhNp through two distinct inhibitory interfaces. Ptch1-A binds to ShhNp through the well-characterized Ca2+-mediated interface on the globular domain of ShhNp, and Ptch1-B primarily interacts with the N-terminal peptide and the palmitoyl moiety. Map comparison reveals that the cholesteryl moiety of native ShhN occupies a recently identified extracellular steroid binding pocket in Ptch1-B. Our structure elucidates the tetrameric assembly of Ptch1 and suggests asymmetric mode of actions of the Hh ligands for inhibiting the potential cholesterol transport activity of Ptch1.

biophysics↗

bioRxiv2018

Global analysis of N6-methyladenosine functions and its disease association using deep learning and network-based methods

N6-methyladenosine (m6A) is the most abundant methylation, existing in >25% of human mRNAs. Exciting recent discoveries indicate the close involvement of m6A in regulating many different aspects of mRNA metabolism and diseases like cancer. However, our current knowledge about how m6A levels are controlled and whether and how regulation of m6A levels of a specific gene can play a role in cancer and other diseases is mostly elusive. We propose in this paper a computational scheme for predicting m6A-regulated genes and m6A-associated disease, which includes Deep-m6A, the first model for detecting condition-specific m6A sites from MeRIP-Seq data with a single base resolution using deep learning and a new network-based pipeline that prioritizes functional significant m6A genes and its associated diseases using the Protein-Protein Interaction (PPI) and gene-disease heterogeneous networks. We applied Deep-m6A and this pipeline to 75 MeRIP-seq human samples, which produced a compact set of 709 functionally significant m6A-regulated genes and nine functionally enriched subnetworks. The functional enrichment analysis of these genes and networks reveal that m6A targets key genes of many critical biological processes including transcription, cell organization and transport, and cell proliferation and cancer-related pathways such as Wnt pathway. The m6A-associated disease analysis prioritized five significantly associated diseases including leukemia and renal cell carcinoma. These results demonstrate the power of our proposed computational scheme and provide new leads for understanding m6A regulatory functions and its roles in diseases.\n\nAuthor summaryThe goal of this work is to identify functional significant m6A-regulated genes and m6A-associated diseases from analyzing an extensive collection of MeRIP-seq data. To achieve this, we first developed Deep-m6A, a CNN model for single-base m6A prediction. To our knowledge, this is the first condition-specific single-base m6A site prediction model that combines mRNA sequence feature and MeRIP-Seq data. The 10-fold cross-validation and test on an independent dataset showthat Deep-m6A outperformed two sequence-based models. We applied Deep-m6A followed by network-based analysis using HotNet2 and RWRH to 75 human MeRIP-Seq samples from various cells and tissue under different conditions to globally detect m6A-regulated genes and further predict m6A mediated functions and associated diseases. This is also to our knowledge the first attempt to predict m6A functions and associated diseases using only computational methods in a global manner on a large number of human MeRIP-Seq samples. The predicted functions and diseases show considerable consistent with those reported in the literature, which demonstrated the power of our proposed pipeline to predict potential m6A mediated functions and associated diseases.

bioinformatics↗

bioRxiv2018

Dimethylarsenic acid (DMA) accumulation positively correlates with realgar-induced subchronic toxicity in rats

The toxicity of realgar depends largely on different arsenic species accumulation and distribution in the body. Here, after continuous oral administration of different doses of realgar for 90 days and subsequent 60-day withdrawal period, clinical observations, food consumption, body weights, blood biochemistry, hematology, and histomorphological examination of rats were performed. Realgar 40mg{middle dot}kg-1{middle dot}d-1 and 170 mg{middle dot}kg-1{middle dot}d-1 of realgar (which is equivalent to 40-fold and 100-fold the maximum clinical dose, respectively) can cause toxicity in rats, including degreased body weight, peripheral blood neutrality abnormal ratio of granulocytes and lymphocytes, hypercoagulability of the blood, liver and kidney tissue damage, liver and kidney may be the main toxic target organs of realgar. The no observed adverse effect level (NOAEL) dose is 10 mg{middle dot}kg-1. At the same time, the content and distribution of arsenic species in tissues were determined. The content of total arsenic (tAs) and Dimethylarsenic acid (DMA) in the tissues of the realgar group was significantly higher than those of the control group. After 60 days of discontinuation, the DMA content in the realgar group decreased, but it was still higher than that in the control group, and liver and kidney damage occurred during the administration period basically returned to normal. Therefore, the authors speculated that when the DMA content in the tissue exceeds a certain range, liver and kidney toxicity will be induced. However, when the DMA content is lower than the above threshold after drug withdrawal, the liver and kidney lesions can return to normal.

pharmacology and toxicology↗

bioRxiv2018

Quantitative mass spectrometry to interrogate proteomic heterogeneity in metastatic lung adenocarcinoma and validate a novel somatic mutation CDK12-G879V

Lung cancer is the leading cause of cancer death both in men and women. Tumor heterogeneity is an impediment to targeted treatment of all cancers, including lung cancer. Here, we sought to characterize changes in tumor proteome and phosphoproteome by longitudinal, prospective collection of tumor tissue of an exceptional responder lung adenocarcinoma patient who survived with metastatic lung adenocarcinoma for more than seven years with HER2-directed therapy in combination with chemotherapy. We employed \"Super-SILAC\" and TMT labeling strategies to quantify the proteome and phosphoproteome of a lung metastatic site and ten different metastatic progressive lymph nodes collected across a span of seven years, including five lymph nodes procured at autopsy. We identified specific signaling networks enriched in lung compared to the lymph node metastatic sites. We correlated the changes in protein abundance with changes in copy number alteration (CNA) and transcript expression. To further interrogate the mass spectrometry data, patient-specific database was built incorporating all the somatic variants identified by whole genome sequencing (WGS) of genomic DNA from the lung, one lymph node metastatic site and blood. An extensive validation pipeline was built for confirmation of variant peptides. We validated 360 spectra corresponding to 55 germline and 6 somatic variant peptides. Targeted MRM assays demonstrated expression of two novel variant somatic peptides, CDK12-G879V and FASN-R1439Q, with expression in lung and lymph node metastatic sites, respectively. CDK12 G879V mutation likely results in a nonfunctional CDK12 kinase and chemotherapy susceptibility in lung metastatic sites. Knockdown of CDK12 in lung adenocarcinoma cells results in increased chemotherapy sensitivity, explaining the complete resolution of the lung metastatic sites in this patient.

cancer biology↗

bioRxiv2018

Extensive Expansion of the Speedy gene Family in Homininae and Functional Differentiation in Humans

BackgroundThe cell cycle plays important roles in physiology and disease. The Speedy/RINGO family of atypical cyclins regulates the cell cycle. However, the origin, evolution and function of the Speedy family are not completely understood. Understanding the origins and evolution of Speedy family would shed lights on the evolution of complexity of cell cycles in eukaryotes.\n\nResultsHere, we performed a comprehensive identification of Speedy genes in 258 eukaryotic species and found that the Speedy subfamily E was extensively expanded in Homininae, characterized by emergence of a low-Spy1-identify domain. Furthermore, the Speedy gene family show functional differentiation in humans and have a distinct expression pattern, different regulation network and co-expressed gene networks associated with cell cycle and various signaling pathways. Expression levels of the Speedy gene family are prognostic biomarkers among different cancer types.\n\nConclusionsOverall, we present a comprehensive view of the Speedy genes and highlight their potential function.

evolutionary biology↗

bioRxiv2018

Deep clustering of protein folding simulations

We examine the problem of clustering biomolecular simulations using deep learning techniques. Since biomolecular simulation datasets are inherently high dimensional, it is often necessary to build low dimensional representations that can be used to extract quantitative insights into the atomistic mechanisms that underlie complex biological processes. In this paper, we use a convolutional variational autoencoder (CVAE) to learn low dimensional, biophysically relevant latent features from long time-scale protein folding simulations in an unsupervised manner. We demonstrate our approach on three model protein folding systems, namely the Fs-peptide (14 s aggregate sampling), villin head piece (single trajectory of 125 s) and the mixed {beta}-{beta}- (BBA) protein (223 + 102 s sampling across two independent trajectories). In these systems, we show that the CVAE latent features learned correspond to distinct conformational substates along the protein folding pathways. The CVAE model predicts nearly 89% of all contacts within the folding trajectories correctly, while being able to extract folded, unfolded and potentially misfolded states in an unsupervised manner. Further, the CVAE model can be used to learn latent features of protein folding that can be applied to other independent trajectories, making it particularly attractive for identifying intrinsic features that correspond to conformational substates that share similar structural features. Together, we show that the CVAE model can quantitatively describe complex biophysical processes such as protein folding.

biophysics↗

bioRxiv2018

Pleotropic effects of PPARD accelerate colorectal tumor progression and invasion

Colorectal carcinogenesis (CRC) progression requires additional molecular mechanisms to APC mutations/aberrant {beta}-catenin signaling. PPARD is a druggable ligand-activated nuclear receptor that regulates essential genes involved in cell fate. PPARD is upregulated in intestinal epithelial cells (IECs) of human colorectal adenomas and adenocarcinomas. The mechanistic significance of PPARD upregulation in CRC remains unknown. Here we show that targeted PPARD overexpression in IECs of mice strongly augmented {beta}-catenin activation via BMP7/TAK1 signaling, promoted intestinal tumorigenesis in Apcmin mice, and accelerated CRC progression and invasiveness in mice with IEC-targeted Apc{Delta}580 mutation. Human CRC invasive fronts had higher PPARD expression than their paired adenomas. A PPARD agonist (GW501516) enhanced APC{Delta}580 mutation-driven CRC, while a PPARD antagonist (GSK3787) suppressed it. Functional proteomics analyses and subsequent validation studies uncovered PPARD upregulation of multiple pro-invasive pathways that drive CRC progression (e.g. PDGFR{beta}, AKT1, CDK1 and EIF4G1). Our results identify novel mechanisms by which PPARD promotes CRC invasiveness and provide the rational for the development of PPARD antagonists to suppress CRC.

cancer biology↗

bioRxiv2018

Bifunctional enzyme SpoT is involved in biofilm formation of Helicobacter pylori with multidrug resistance by upregulating efflux pump Hp1174 (gluP)

The drug resistance of Helicobacter pylori (H. pylori) is gradually becoming a serious problem. Biofilm formation is an important factor that leads to multidrug resistance in bacteria. The ability of H. pylori to form biofilms on the gastric mucosa has been known. However, there are few studies on the regulation mechanisms of H. pylori biofilm formation and multidrug resistance. Guanosine 3-diphosphate 5-triphosphate and guanosine 3,5-bispyrophosphate [(p)ppGpp] are global regulatory factors and are synthesized in H. pylori by the bifunctional enzyme SpoT. It has been reported that (p)ppGpp is involved in the biofilm formation and multidrug resistance of various bacteria. In this study, we found that SpoT also plays an important role in H. pylori biofilm formation and multidrug resistance. Therefore, it is necessary to carry out some further studies regarding its regulatory mechanism. Considering that efflux pumps are of great importance in the biofilm formation and multidrug resistance of bacteria, we tried to find if efflux pumps controlled by SpoT participate in these activities. Then, we found that Hp1174 (glucose/galactose transporter, gluP), an efflux pump of the MFS family, is highly expressed in biofilm-forming and multi-drug resistance (MDR) H. pylori and is upregulated by SpoT. Through further research, we determined that gluP involved in H. pylori biofilm formation and multidrug resistance. Furthermore, the average expression level of gluP in clinical MDR strains was considerably higher than that in clinical drug-sensitive strains. Taken together, our results revealed a novel molecular mechanism of H. pylori tolerance to multidrug.

microbiology↗

bioRxiv2018

Integrated proteogenomic analysis of metastatic thoracic tumors identifies APOBEC mutagenesis and copy number alterations as drivers of proteogenomic tumor evolution and heterogeneity

Elucidation of the proteogenomic evolution of metastatic tumors may offer insight into the poor prognosis of patients harboring metastatic disease. We performed whole-exome and transcriptome sequencing, copy number alterations (CNA) and mass spectrometry-based quantitative proteomics of 37 lung adenocarcinoma (LUAD) and thymic carcinoma (TC) metastases obtained by rapid autopsy and found evidence of patient-specific, multi-dimensional heterogeneity. Extreme mutational heterogeneity was evident in a subset of patients whose tumors showed increased APOBEC-signature mutations and expression of APOBEC3 region transcripts compared to patients with lesser mutational heterogeneity. TP53 mutation status was associated with APOBEC hypermutators in our cohort and in three independent LUAD datasets. In a thymic carcinoma patient, extreme heterogeneity and increased APOBEC3AB expression was associated with a high-risk germline APOBEC3AB variant allele. Patients with CNA occurring late in tumor evolution had corresponding changes in gene expression and protein abundance indicating genomic instability as a mechanism of downstream transcriptomic and proteomic heterogeneity between metastases. Across all tumors, proteomic heterogeneity was greater than copy number and transcriptomic heterogeneity. Enrichment of interferon pathways was evident both in the transcriptome and proteome of the tumors enriched for APOBEC mutagenesis despite a heterogeneous immune microenvironment across metastases suggesting a role for the immune microenvironment in the expression of APOBEC transcripts and generation of mutational heterogeneity. The evolving, heterogeneous nature of LUAD and TC, through APOBEC-mutagenesis and CNA illustrate the challenges facing treatment outcomes.

cancer biology↗