Biology subjects

Fang, X.

Publications and source records attributed to Fang, X..

At least 127 records · Page 7Linked to original sources

bioRxiv2020

Molecular mechanisms underlying the extreme mechanical anisotropy of the flaviviral exoribonuclease-resistant RNAs (xrRNAs)

Mechanical anisotropy is an essential property for many biomolecules to assume their structures, functions and applications, however, the mechanisms for their direction-dependent mechanical responses remain elusive. Herein, by using single-molecule nanopore sensing technique, we explore the mechanisms of directional mechanical stability of the xrRNA1 RNA from ZIKA virus (ZIKV), which forms a complex ring-like architecture. We reveal extreme mechanical anisotropy in ZIKV xrRNA1 which highly depends on Mg2+ and the key tertiary interactions. The absence of Mg2+ and disruption of the key tertiary interactions strongly affect the structural integrity and attenuate mechanical anisotropy. The significance of ring structure in RNA mechanical anisotropy is further supported by steered molecular dynamics simulations on ZIKV xrRNA1 and another two RNAs with ring structures, the HCV IRES and THF riboswitch. We anticipate the ring structures can be used as key elements to build RNA-based nanostructures with controllable mechanical anisotropy for biomaterial and biomedical applications.

biophysics↗

bioRxiv2020

Site-specific covalent labeling of large RNAs with nanoparticles empowered by expanded genetic alphabet transcription

Conjugation of RNAs with nanoparticles is of significant importance for its numerous applications in biology and medicine, which however remains challenging, especially for large ones. So far, the majority of RNA labeling rely on solid-phase chemical synthesis, which is generally limited to RNAs smaller than 100 nts. We here present an efficient and generally applicable labeling strategy for site-specific covalent conjugation of large RNAs with gold nanoparticle (AuNP) empowered by expanded genetic alphabet transcription. We synthesize an amine-derivatized TPT3 (TPT3A), which are site-specifically incorporated into a 97-nt 3SL RNA and a 719-nt mini genomic RNA (DENV-mini) from Dengue virus serotype 2 (DENV2) by standard in vitro transcription with expanded genetic alphabet containing the A-T, G-C natural base pairs and the TPT3-NaM unnatural base pair. TPT3 modification cause minimal structural perturbations to the RNAs by small angle X-ray scattering. The purified TPT3A-modified RNAs are covalently conjugated with mono-Sulfo-NHS-Nanogold nanoparticles via the highly selective amine-NHS ester reaction and purified under non-denaturing conditions. We demonstrate the application of the AuNP-RNA conjugates in large RNA structural biology by an emerging molecular ruler, X-ray scattering interferometry (XSI). The inter-nanoparticle distance distributions in the 3SL and DENV-mini RNAs derived from XSI measurements support the hypothetical model of flavivirus genome circularization, thus validate the applicability of this novel labeling strategy. The presented strategy overcomes the size constraints in conventional RNA labeling strategies, and is expected to have wide applications in large RNA structural biology and RNA nanotechnology. Significance StatementWe present a site-specific labeling strategy for large RNAs by T7 transcription with expanded genetic alphabet containing TPT3-NaM unnatural base pair. The applicability of this labeling strategy is validated by X-ray scattering interferometry measurements on a 97-nt and a 719-nt RNAs. This strategy can be applicable to natural RNAs or artificial RNA nanostructures with sizes from tens up to thousands of nucleotides, or covalent conjugation of RNAs with other metal nanoparticles. The usage of a far upstream forward primer during PCR enables easy purification of RNA from DNA templates, the non-denaturing conditions for conjugation reactions and purification avoids potential large RNA misfolding. This labeling strategy expands our capability to site-specifically conjugate RNA with nanoparticles for many applications.

biophysics↗

bioRxiv2020

Fibrinolytic niche is requested for alveolar type 2 cell-mediated alveologenesis and injury repair

COVID-19, SARS, and MERS are featured by fibrinolytic dysfunction. To test the role of the fibrinolytic niche in the regeneration of alveolar epithelium, we compared the self-renewing capacity of alveolar epithelial type 2 (AT2) cells and its differentiation to AT1 cells between wild type (wt) and fibrinolytic niche deficient mice (Plau-/- and Serpine1Tg). A significant reduction in both proliferation and differentiation of deficient AT2 cells was observed in vivo and in 3D organoid cultures. This decrease was mainly restored by uPA derived A6 peptide, a binding fragment to CD44 receptors. The proliferative and differential rate of CD44+ AT2 cells was greater than that of CD44- controls. There was a reduction in transepithelial ion transport in deficient monolayers compared to wt cells. Moreover, we found a marked suppression in total AT2 cells and CD44+ subpopulation in lungs from brain dead patients with acute respiratory distress syndrome (ARDS) and a mouse model infected by influenza viruses. Thus, we demonstrate that the fibrinolytic niche can regulate AT2-mediated homeostasis and regeneration via a novel uPA-A6-CD44+-ENaC cascade.

cell biology↗

bioRxiv2020

Precision engineering of the transcription factor Cre1 in Hypocrea jecorina (Trichoderma reesei) for efficient cellulase production in the presence of glucose

In Trichoderma reesei, carbon catabolite repression (CCR) significantly downregulates the transcription of cellulolytic enzymes, which is usually mediated by the zinc finger protein Cre1. It was found that there is a conserved region at the C-terminus of Cre1/CreA in several cellulase-producing fungi that contains up to three continuous S/T phosphorylation sites. Here, S387, S388, T389, and T390 at the C-terminus of Cre1 in T. reesei were mutated to valine for mimicking an unphosphorylated state, thereby generating the transformants Tr_Cre1S387V, Tr_Cre1S388V, Tr_Cre1T389V, and Tr_Cre1T390V, respectively. Transcription of cel7a in Tr_ Cre1S388V was markedly higher than that of the parent strain when grown in glucose-containing media. Under these conditions, both filter paperase (FPase) and p-nitrophenyl-{beta}-D-cellobioside (pNPCase) activities, as well as soluble proteins from Tr_Cre1S388V were significantly increased by up to 2- to 3-fold compared with that of other transformants and the parent strain. To our knowledge, this is the first report demonstrating an improvement of cellulase production in fungal species under CCR by mimicking dephosphorylation at the C-terminus of Cre1. Taken together, we developed a precision engineering strategy based on the modification of phosphorylation sites of Cre1 transcription factor to enhance the production of cellulase in fungal species under CCR.

bioengineering↗

bioRxiv2020

Dampened PI3K/AKT signaling contributes to cancer resistance of the naked mole rat

Mammalian species have a dramatically different susceptibility to cancer. However, how cancer-resistant species resist oncogenic transformation is not fully understood. Here, we performed a comprehensive analysis of oncogene-induced transcriptional changes in the fibroblasts of a cancer-prone species, the mouse, and three cancer-resistant species, the human, the blind mole rat, and the naked mole rat. We report that multiple cellular processes are more refractory to oncogene-induced transcriptional changes in blind mole-rat, naked mole-rat, or human cells compared to mouse cells, such as cell division, cell adhesion, extracellular matrix organization, and metabolism. Strikingly, naked mole rat cells are more resistant to Ras-induced transcriptional changes compared to the other three species. As a mechanism, we found that critical genes in the PI3K pathway including Akt1 and Pik3ca are downregulated in naked mole rat cells. Activating the PI3K/AKT pathway in the naked mole rat cells renders them susceptible to tumorigenic transformation. This study provides multiple new insights into anti-cancer mechanisms in cancer-resistant species of mammals. Significance statementAnimal species differ greatly in their cancer susceptibility. Cancer rates in the mouse range from 50-90%, while two other rodent species, the naked mole rat and the blind mole rat have only a few cancer cases ever reported. Here we examined the mechanisms responsible these differences by comparing changes in transcription patters in response to oncoproteins in the mouse, naked mole rat, blind mole rat and human cells. The most striking finding was that the naked mole rat cells were resistant to transcriptional changes induced by oncogenic Ras. We found that pathways downstream of Ras were naturally attenuated in the naked mole rat. This finding identifies a novel mechanism that evolved to provide tumor resistance to the naked mole rat.

cancer biology↗

bioRxiv2019

In vivo mRNA structure regulates miRNA cleavage in Arabidopsis

MicroRNA (miRNA)-mediated cleavage is involved in numerous essential cellular pathways. miRNAs recognize target RNAs via sequence complementarity. In addition to complementarity, in vitro and in silico studies have suggested that RNA structure may influence the accessibility of mRNAs to miRNA-Induced Silencing Complexes (miRISCs), thereby affecting RNA silencing. However, the regulatory mechanism of mRNA structure in miRNA cleavage remains elusive. Here, we investigated the role of in vivo RNA secondary structure in miRNA cleavage by developing the new CAP-STRUCTURE-seq method to capture the intact mRNA structurome in Arabidopsis thaliana. This approach revealed that miRNA target sites were not structurally accessible for miRISC binding prior to cleavage in vivo. Instead, the unfolding of the target site structure is the primary determinant for miRISC activity in vivo. Notably, we found that the single-strandedness of the two nucleotides immediately downstream of the target site, named Target Adjacent structure Motif (TAM), can promote miRNA cleavage but not miRNA binding, thus decoupling target site binding from cleavage. Our findings demonstrate that mRNA structure in vivo can regulate miRNA cleavage, providing evidence of mRNA structure-dependent regulation of biological processes.

molecular biology↗

bioRxiv2019

Extensive Mammalian Germline Genome Engineering

Xenotransplantation, specifically the use of porcine organs for human transplantation, has long been sought after as an alternative for patients suffering from organ failure. However, clinical application of this approach has been impeded by two main hurdles: 1) risk of transmission of porcine endogenous retroviruses (PERVs) and 2) molecular incompatibilities between donor pigs and humans which culminate in rejection of the graft. We previously demonstrated that all 25 copies of the PERV elements in the pig genome could be inactivated and live pigs successfully generated. In this study, we improved the scale of porcine germline editing from targeting a single repetitive locus with CRISPR to engineering 13 different genes using multiple genome engineering methods. we engineered the pig genome at 42 alleles using CRISPR-Cas9 and transposon and produced PERVKO{middle dot}3KO{middle dot}9TG pigs which carry PERV inactivation, xeno-antigen KO and 9 effective human transgenes. The engineered pigs exhibit normal physiology, fertility, and germline transmission of the edited alleles. In vitro assays demonstrated that these pigs gain significant resistance to human humoral and cell mediated damage, and coagulation dysregulations, similar to that of allotransplantation. Successful creation of PERVKO{middle dot}3KO{middle dot}9TG pigs represents a significant step forward towards safe and effective porcine xenotransplantation, which also represents a synthetic biology accomplishment of engineering novel functions in a living organism. One Sentence SummaryExtensive genome engineering is applied to modify pigs for safe and immune compatible organs for human transplantation

synthetic biology↗

bioRxiv2019

Annexin A2 depletion exacerbates the intracerebral microhemorrhage induced by acute rickettsia and Ebola virus infections

Intracerebral microhemorrhages (CMHs) are small foci of hemorrhages in the cerebrum. Acute infections induced by some intracellular pathogens, including rickettsia, can result in CMHs. Annexin a2 (ANXA2) has been documented to play a functional role during intracellular bacterial adhesion. Here we report that ANXA2-knockout (KO) mice are more susceptible to CMHs in response to rickettsia and Ebola virus infections, suggesting an essential role of ANXA2 in protecting vascular integrity during these intracellular pathogen infections. Proteomic analysis via mass spectrometry of whole brain lysates and brain-derived endosomes from ANXA2-KO and wild-type (WT) mice post-infection with R. australis revealed that a variety of significant proteins were differentially expressed, and the follow-up function enrichment analysis had identified several relevant cell-cell junction functions. Immunohistology study confirmed that both infected WT and infected ANXA2-KO mice were subjected to adherens junctional protein (VE-cadherin) damages. However, key blood-brain barrier (BBB) components, tight junctional proteins ZO-1 and occludin, were disorganized in the brains from R. australis-infected ANXA2-KO mice, but not those of infected WT mice. Similar ANXA2-KO dependent CMHs and fragments of ZO-1 and occludin were also observed in Ebola virus-infected ANXA2-KO mice, but not found in infected WT mice. Overall, our study revealed a novel role of ANXA2 in the formation of CMHs during R. australis and Ebola virus infections; and the underlying mechanism is relevant to the role of ANXA2-regulated tight junctions and its role in stabilizing the BBB in these deadly infections. Author SummaryTraditionally, spontaneous intracerebral microhemorrhages (CMHs) were defined as small foci of intracerebral hemorrhages. Such atraumatic CMHs are due to the rupture of a weak blood vessel wall. Infections complicating cerebrovascular accidents have been extensively investigated. However, the role of CMHs complicating infections, in particularly acute systemic infections, has been poorly explored. Population-based retrospective cohort studies suggest there are potentially more undiscovered cases of CMHs accompanying acute systemic infections. Given both the lack of an animal model and cellular/molecular pathophysiology of CMHs following acute systemic infections, there is an urgent need to increase our comprehensive understanding of acute infection-induced CMHs. Overall, our study revealed a novel role of annexin a2 (ANXA2) in the formation of CMHs during R. australis and Ebola virus infections; and the underlying mechanism is relevant to the role of ANXA2-regulated endothelial tight junctions and its role in stabilizing the blood-brain barrier in these deadly infections.

pathology↗

bioRxiv2019

Mapping Human Pluripotent Stem Cell Derived Erythroid Differentiation by Single-Cell Transcriptome Analysis

There is an imbalance between the supply and demand of functional red blood cells (RBCs) in clinical applications. This imbalance can be addressed by regenerating RBCs using several in vitro methods. Induced pluripotent stem cells (iPSCs) can handle the low supply of cord blood and the ethical issues in embryonic stem cell research and provide a promising strategy to eliminate immune rejection. However, no complete single-cell level differentiation pathway exists for the iPSC-derived RBC differentiation system. In this study, we used iPSC line BC1 to establish a RBCs regeneration system. The 10x genomics single-cell transcriptome platform was used to map the cell lineage and differentiation trajectories on day 14 of the regeneration system. We observed that iPSCs differentiation was not synchronized during embryoid body (EB) culture. The cells (day 14) mainly consisted of mesodermal and various blood cells, similar to the yolk sac hematopoiesis. We identified six cell classifications and characterized the regulatory transcription factors (TFs) networks and cell-cell contacts underlying the system. iPSCs undergo two transformations during the differentiation trajectory, accompanied by the dynamic expression of cell adhesion molecules and estrogen-responsive genes. We identified different stages of erythroid cells such as burst-forming unit erythroid (BFU-E) and orthochromatic erythroblasts (ortho-E) and found that the regulation of TFs (e.g., TFDP1 and FOXO3) is erythroid-stage specific. Immune erythroid cells were identified in our system. This study provides systematic theoretical guidance for optimizing the iPSCs-derived RBCs differentiation system, and this system is a useful model for simulating in vivo hematopoietic development and differentiation.

developmental biology↗

bioRxiv2019

A Synthetic Biology Approach to Sequential Stripe Patterning and Somitogenesis

Reaction-diffusion (RD) based clock and wavefront model has long been proposed as the mechanism underlying biological pattern formation of repeated and segmented structures including somitogenesis. However, systematic molecular level understanding of the mechanism remains elusive, largely due to the lack of suitable experimental systems to probe RD quantitatively in vivo. Here we design a synthetic gene circuit that couples gene expression regulation (reaction) with quorum sensing (diffusion) to guide bacterial cells self-organizing into stripe patterns at both microscopic and colony scales. An experimentally verified mathematical model confirms that these periodic spatial structures are emerged from the integration of oscillatory gene expression as the molecular clock and the outward expanding diffusions as the propagating wavefront. Furthermore, our paired model-experiment data illustrate that the RD-based patterning is sensitive to initial conditions and can be modulated by external inducers to generate diverse patterns, including multiple-stripe pattern, target-like pattern and ring patterns with reversed fluorescence. Powered by our synthetic biology setup, we also test different topologies of gene networks and show that network motifs enabling robust oscillations are foundations of sequential stripe pattern formation. These results verified close connections between gene network topology and resulting RD driven pattern formation, offering an engineering approach to help understand biological development.

synthetic biology↗

bioRxiv2019

Aquila_stLFR: assembly based variant calling package for stLFR and hybrid assembly for linked-reads

MotivationIdentifying structural variants (SVs) is of critical importance in health and disease, however, detecting them remains a scientific and computing challenge. Several linked-read sequencing technologies, including 10X linked-read, TELL-Seq, and single tube long fragment read (stLFR), have been recently developed as cost-effective approaches to reconstruct multi-megabase haplotypes (phase blocks) from sequence data of a single sample. These technologies provide an optimal sequencing platform to characterize SVs, though few computational algorithms can utilize them. Thus, we developed Aquila_stLFR, an approach that resolves SVs through haplotype-based assembly of stLFR linked-reads. ResultsAquila_stLFR first partitions LFRs into two haplotype-specific blocks, by taking advantage of the potential phasing ability of the linked-read itself. Each haplotype is then assembled independently, to achieve a complete diploid assembly to finally reconstruct the genome-wide SVs. We benchmarked Aquila_stLFR on a well-studied sample, NA24385, and showed Aquila_stLFR can detect medium to large size (50bp - 10kb) deletions with a high sensitivity and insertions with a high specificity. AvailabilitySource code and documentation are available on https://github.com/maiziex/Aquila_stLFR. Contactmaizie.zhou@vanderbilt.edu Supplementary informationSupplementary data are available at Bioinformatics online.

bioinformatics↗

bioRxiv2019

Acute characterization of tissue and functional deficits in a clinically translatable pig model of ischemic stroke

The acute stroke phase is a critical time frame used to evaluate stroke severity, therapeutic options, and prognosis while also serving as a major target for the development of diagnostics. To better understand stroke pathophysiology and to enhance the development of treatments, our group developed a translational pig ischemic stroke model. In this study, the evolution of acute ischemic stroke tissue damage, immune response, and functional deficits were further characterized in the pig model. Stroke was induced by middle cerebral artery occlusion in Landrace pigs. At 24 hours post-stroke, magnetic resonance imaging revealed a decrease in ipsilateral diffusivity and an increase in hemispheric swelling and intracranial hemorrhage resulting in notable midline shift. Stroke negatively impacted white matter integrity leading to decreased fractional anisotropy. Similar to acute clinical patients, stroked pigs showed a reduction in circulating lymphocytes and a surge in neutrophils and band cells. Functional responses corresponded with structural changes with reduced exploration in open field testing and impairments in spatiotemporal gait parameters. This novel, acute ischemia characterization provides important insights into tissue and functional level changes in a pig model that can be used to identify treatment targets and future testing of therapeutics and diagnostics.

neuroscience↗

bioRxiv2019

K+ uptake and root-to-shoot allocation in Arabidopsis require coordination of nitrate transporter1/peptide transporter family member NPF6.3/NRT1.1

K+ and NO3- are the major forms of potassium and nitrogen that are absorbed by the roots of most terrestrial plants. In this study, we observed that the close relationship between NO3- and K+ homeostasis was mediated by nitrate transporter1 (NRT1.1) in Arabidopsis. The nrt1.1 mutants lacking NRT1.1 function showed disturbed K+ uptake and root-to-shoot allocation, especially under K+-limited conditions, and had a yellow-shoot sensitive phenotype on K+-limited medium. The K+ uptake and root-to-shoot allocation of these mutants were partially rescued by expressing NRT1.1 in the root epidermis-cortex and central vasculature by using Sultr1;2 and PHO1 promoters, respectively. Furthermore, two-way analysis of variance based on the K+ content in nrt1.1-1/akt1, nrt1.1-1/hak5-3, nrt1.1-1/kup7, and nrt1.1-1/skor-2 double mutants and their corresponding single mutants and wild-type plants revealed physiological interactions between NRT1.1 and K+ channels located in the root epidermis-cortex and central vasculature. Taken together, these data suggest that the expression of NRT1.1 in the root epidermis-cortex coordinates with K+ uptake channels to improve K+ uptake, whereas its expression in the root central vasculature coordinates with the channels loading K+ into the xylem to facilitate K+ allocation from the roots to the shoot.

plant biology↗

bioRxiv2019

Adaptive laboratory evolution of Escherichia coli under acid stress

The ability of Escherichia coli to tolerate acid stress is important for its survival and colonization in the human digestive tract. Here, we performed adaptive laboratory evolution of the laboratory strain E. coli K-12 MG1655 at pH 5.5 in glucose minimal medium. By 800 generations, six independent populations under evolution reached 18.0% higher growth rates than their starting strain at pH 5.5, while maintaining comparable growth rates to the starting strain at pH 7. We characterized the evolved strains to find that: (1) whole genome sequencing of isolated clones from each evolved population revealed mutations in rpoC appearing in 5 of 6 sequenced clones; (2) gene expression profiles revealed different strategies to mitigate acid stress, that are related to amino acid metabolism and energy production and conversion. Thus, a combination of adaptive laboratory evolution, genome resequencing, and expression profiling reveals, on a genome-scale, the strategies that E. coli deploys to mitigate acid stress.

microbiology↗

bioRxiv2019

A Clostridia-rich microbiota contributes to increased excretion of bile acids in diarrhea-predominant irritable bowel syndrome

ObjectiveAn excess of fecal bile acids (BAs) is thought to be one of the mechanisms for diarrhea-predominant irritable bowel syndrome (IBS-D). However, the factors causing excessive BA excretion remains unclear. Given the importance of gut microbiota in BA metabolism, we hypothesized that gut dysbiosis might contribute to excessive BA excretion in IBS-D.\n\nDesignMetabolomic and metagenomic analyses were performed of specimens from 290 IBS-D patients and 89 healthy volunteers. By transplanting human microbiota and manipulating specific microbiome species in mice, the effects of microbiota on host BA metabolism were assessed at metabolic, genetic and protein levels. Effects of individual and mixed BAs on enterohepatic feedback pathways were also tested in vitro and in vivo.\n\nResultsTotal fecal BAs were excessively excreted in 24.5% of IBS-D patients. Their fecal metagenomes showed increased abundances of Clostridia and BA-transforming genes (hdhA and bais). The increases of Clostridia bacteria (e.g. C. scindens) were positively associated with the levels of fecal BAs and serum 7-hydroxy-4-cholesten-3-one (C4), while being negatively correlated with serum fibroblast growth factor 19 (FGF19). Both Clostridia-rich human microbiota and C. scindens enhanced levels of serum C4 and hepatic conjugated BAs in mice recipients and reduced ileal FGF19 expression. Inhibition of Clostridium species by vancomycin yielded opposite findings. Clostridia-derived BAs (e.g. conjugated and free ursodeoxycholic acid) significantly suppressed intestinal FGF19 expression.\n\nConclusionThe Clostridia-rich microbiota contributes to excessive BA excretion in IBS-D patients. This study provided the basis for more precise clinical diagnosis and management for IBS-D.

microbiology↗

bioRxiv2019

Microarray screening of differentially expressed genes after up-regulating miR-205 or down-regulating miR-141 in cervical cancer cells

Cervical cancer is one of the most common gynecological malignancies. However,studies on the expression and molecular mechanism of miR-205 and miR-141 in CC are insufficient recently. Expression profile microarray with 21329 Oligo DNA were used to detect the expression of mRNAs in miR-205 up-regulated or miR-141 down-regulated HeLa and SiHa cells and mRNAs in normal HeLa and SiHa cells. Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway were performed to assess the potential pathways of miR-205 in SiHa cell.Compared with normal HeLa cell, there were 38 differentially expressed genes (DEGs) in miR-205 up-regulated HeLa cell. Nine were up-regulation genes and 29 were down-regulation genes. There were 23 DEGs in miR-141 down-regulated HeLa cell. One was up-regulated and 22 were down-regulated. Compared with normal SiHa cell, there were 128 DEGs in miR-205 up-regulated SiHa cell. One hundred and three were up-regulation genes and 25 were down-regulation genes. There were 80 DEGs in miR-141 down-regulated SiHa cell. Forty two were up-regulation genes and 28 were down-regulation genes. For miR-205 up-regulated SiHa cell, GO outcome showed that \"ubiquitin-protein ligase activity\", \"MAP kinase phosphatase activity\", were the most enriched terms (P < 0.05). And in KEGG analysis, \"Cell cycle\" was notably enriched, and Smad4 in this pathway was up-regulated (P < 0.05). Expression profile microarray technology can effectively screen out DEGs in cervical cancer cells after up-regulating miR-205 or down-regulating miR-141. Which may enable us to understand the pathogenesis and lay an important foundation for the prevention and treatment of cervical cancer.

cancer biology↗

bioRxiv2019

A high-precision hybrid algorithm for predicting eukaryotic protein subcellular localization

MotivationSubcellular location plays an essential role in protein synthesis, transport, and secretion, thus it is an important step in understanding the mechanisms of trait-related proteins. Generally, homology methods provide reliable homology-based results with small E-values. We must resort to pattern recognition algorithms (SVM, Fisher discriminant, KNN, random forest, etc.) for proteins that do not share significant homologous domains with known proteins. However, satisfying results are seldom obtained.\n\nResultsHere, a novel hybrid method \"Basic Local Alignment Search Tool+Smith-Waterman+Needleman-Wunsch\" or BLAST+SWNW, has been obtained by integrating a loosened E-value Basic Local Alignment Search Tool (BLAST) with the Smith-Waterman (SW) and Needleman-Wunsch (NW) algorithms, and this method has been introduced to predict protein subcellular localization in eukaryotes. When tested on Dataset I and Dataset II, BLAST+SWNW showed an average accuracy of 97.18% and 99.60%, respectively, surpassing the performance of other algorithms in predicting eukaryotic protein subcellular localization.\n\nAvailability and ImplementationBLAST+SWNW is an open source collaborative initiative available in the GitHub repository (https://github.com/ZHANGDAHAN/BLAST-SWNW-for-SLP or http://202.206.64.158:80/link/72016CAC26E4298B3B7E0EAF42288935)\n\nContactzhaqi1972@163.com; zhangdahan@genetics.ac.cn\n\nSupplementary InformationSupplementary data are available at PLOS Computational Biology online.

bioinformatics↗

bioRxiv2018

Transcriptomic responses of the marine cyanobacterium Prochlorococcus to viral lysis products

Marine phytoplankton contributes to about one half of global primary production, and a significant proportion of their photosynthetically fixed organic carbon is released after viral infection as dissolved organic matter (DOM). This DOM pool is known to be consumed by heterotrophic microorganisms; however, its impact on the uninfected co-occurring phytoplankton remains largely unknown. Here, we conducted transcriptomic analyses to study the effects of viral lysis products on the unicellular cyanobacterium Prochlorococcus, which is the most abundant photosynthetic organism on Earth. While Prochlorococcus growth was not affected by viral lysis products, many tRNAs increased in abundance, which was also seen after amino acid addition, suggesting that amino acids are one of the compounds in viral lysis products that affected the expression of tRNA genes. The decreased transcript abundances of N metabolism genes also suggested that Prochlorococcus responded to organic N compounds, consistent with abundant amino acids in viral lysis products. The addition of viral lysis products to Prochlorococcus reduced the maximum photochemical efficiency of photosystem II and CO2 fixation while increased its respiration rate, consistent with differentially expressed genes related to photosynthesis and respiration. One of the highest positive fold-changes was observed for the 6S RNA, a non-coding RNA functioning as a global transcriptional regulator in bacteria. The high level of 6S RNA might be responsible for some of the observed transcriptional responses. Taken together, our results revealed the transcriptional regulation of Prochlorococcus in response to viral lysis products and suggested its metabolic potential to utilize organic N compounds.\n\nImportancePhotosynthetic microorganisms called phytoplankton are abundant in the oceans and contribute to about one half of global CO2 fixation. Phytoplankton are frequently infected by viruses and after infection their organic carbon is released into the ocean as dissolved organic matter (DOM). Marine DOM is important for the marine food web because it supports the growth of heterotrophic microorganisms. However, the impact of viral DOM on the uninfected phytoplankton is largely unknown. In this study, we conducted transcriptomic analyses and identified many differentially expressed genes when viral DOM was added to the marine cyanobacterium Prochlorococcus. One effect of viral DOM is that the carbon fixation of Prochlorococcus was reduced by ~16%, which might affect carbon cycling in the worlds oceans since Prochlorococcus is the most abundant photosynthetic organism on Earth.

microbiology↗