Indirect Binding Research Articles

Unbiased proteomic profiling reveals the IP3R modulator AHCYL1/IRBIT as a novel interactor of microtubule-associated protein tau

Microtubule-associated protein tau is a naturally unfolded protein that can modulate a vast array of physiological processes through direct or indirect binding with molecular partners. Aberrant tau homeostasis has been implicated in the pathogenesis of several neurodegenerative disorders, including Alzheimer’s disease. In this study, we performed an unbiased high-content protein profiling assay by incubating recombinant human tau on microarrays containing thousands of human polypeptides. Among the putative tau-binding partners, we identify SAH hydrolase–like protein 1/inositol 1,4,5-trisphosphate receptor (IP3R)–binding protein (AHCYL1/IRBIT), a member of the SAH hydrolase family and a previously described modulator of IP3R activity. Using coimmunoprecipitation assays, we show that endogenous as well as overexpressed tau can physically interact with AHCYL1/IRBIT in brain tissues and cultured cells. Proximity ligation assay experiments demonstrate that tau overexpression may modify the close localization of AHCYL1/IRBIT to IP3R at the endoplasmic reticulum. Together, our experimental evidence indicates that tau interacts with AHCYL1/IRBIT and potentially modulates AHCYL1/IRBIT function.

Molecular and structural insights of β-boswellic acid and glycyrrhizic acid as potent SARS-CoV-2 Envelope protein inhibitors

BackgroundOver million people have been infected with SARS-CoV-2 virus worldwide, with around 3% reported deaths till date. A few conventional antiviral treatments have been tried to mitigate the coronavirus. However, many alternative therapeutics are being evaluated worldwide. In the present study, we investigated traditional Indian medicinal compounds antiviral potencies as an effective drug for targeting SARS-CoV-2E. SARS-CoV-2 E protein plays a key role in coronavirus life cycle and is an interesting target for the development of anti-SARS-CoV-2 E drugs. MethodsMolecular docking studies of medicinal compounds possessing wide range of pharmacological and antiviral activities against enveloped viruses were evaluated with the computer-aided drug design screening software; PyRx. Twelve medicinal compounds isolated from plants were screened and visualized on Biovia Discovery-Studio. Moreover, SARS-CoV-2 E protein's secondary structural insights were deciphered using Swiss Model and ProFunc web server. ResultsGlycyrrhizic acid, triterpene glycoside isolated from plants of Glycyrrhiza (licorice) showed interactions with envelope protein at chain A: Arg 61, chain B: Phe 23, chain B: Tyr 57, and chain C: Val 25. β- boswellic acid, an ayurvedic herb (pentacyclic terpenoid are produced by Boswellia) represented direct interactions and indirect binding with chain C. Their pharmacological aspects and drug-likeness properties were deduced by DruLiTo. Toxicological assessment, along with their ADME profiling, was validated using vNNADMET. The findings showed that ligands, β-boswellic acid, and glycyrrhizic acid possessed the best bindings, with the target having binding affinity (-9.1 kcal/mol) amongst compounds tested against SARS-CoV-2 E. In-vitro studies reveals the promising effect as potent SARS-CoV-2 E inhibitors. Functionality loss and structural disruptions with ∼90% were observed by UV-spectra and fluorescent based analyses. ConclusionThe study demonstrated that β-boswellic acid, and glycyrrhizic acid are strong SARS-CoV-2 E protein inhibitors. In addition, the work linked GA antiviral activity to its effect on SARS-CoV- 2 E protein that can pave the way for designing antiviral therapeutics.

Thermodynamics-based modeling reveals regulatory effects of indirect transcription factor-DNA binding in Drosophila

Sequence-to-expression relationship is one of the major research topics in the field of regulatory genomics. Prediction of epigenomic states such as DNA accessibility or transcription factor(TF)-DNA binding has been studied extensively, but predicting a gene's expression in different cell types based on its regulatory enhancer sequences remains largely unsolved. Here, we present a thermodynamics-based sequence-to-expression model that combines DNA accessibility, TF concentrations and enhancer sequence information to classify enhancers into their spatio-temporal activity classes in Drosophila mesoderm.

Multiplex Human Papillomavirus L1L2 virus-like particle antibody binding assay

A variety of in vitro techniques are available to estimate the level of antibodies present in human serum samples. Such tests are highly specific and are used to determine prior exposure to a pathogen or to estimate the magnitude, breadth and durability of individual and population level vaccine immunity. Multiplex (or multi-analyte) platforms are increasingly being used to evaluate immune responses against multiple antigens at the same time, usually at reduced per sample cost and a more efficient use of available samples. Consequently, multiplex serology is an essential component of a wide range of public health programmes. Human papillomavirus (HPV) serology is limited to a small number of academic, public health and vaccine manufacturer laboratories globally. Such platforms include indirect binding to the major (L1) capsid protein virus-like particles (VLP), monoclonal antibody competition against L1 VLP and indirect binding to L1 and L2 (minor capsid protein) VLP on multiplex (Luminex®, Meso Scale Discovery®) and standard (ELISA) platforms. The methodology described here utilizes a common multi-analyte platform and L1L2-based VLP expressed in house, which allows the simultaneous detection and quantification of antibody responses against nine vaccine-relevant HPV genotypes.

Overexpression of fucosyltransferase 8 reverses the inhibitory effect of high-dose dexamethasone on osteogenic response of MC3T3-E1 preosteoblasts.

BackgroundCore fucosylation catalyzed by FUT8 is essential for TGF-β binding to TGF-β receptors.MethodsIndirect TGF-β1 binding assay was used to evaluate the ability of TGF-β1 to bind to TGFBRs, Alizarin red and alkaline phosphatase staining were used to detect osteogenic differentiation and mineralization ability , western blot and quantitative RT-PCR were used to measure the differential expression of osteogenesis-related proteins and genes. Plasmid-mediated gain-of-function study. The scale of core fucosylation modification was detected by Lectin-blot and LCA laser confocal.ResultsOur results showed that compared with vehicle treatment, high-dose (10−6 and 10−5 M) dexamethasone significantly inhibited cell proliferation, osteogenic differentiation, and FUT8 mRNA expression while promoting mRNA expression of adipogenesis-related genes in MC3T3-E1 cells, suggesting that downregulation of FUT8 is involved in the inhibitory effect of high-dose dexamethasone on osteogenesis. Overexpression of FUT8 significantly promoted osteogenic differentiation and activated TGF-β/Smad signaling in MC3T3-E1 cells in the presence of high-dose dexamethasone, suggesting that FUT8 reverses the inhibitory effect of high-dose dexamethasone on osteogenesis. In addition, lectin fluorescent staining and blotting showed that overexpression of FUT8 significantly reversed the inhibitory effects of high-dose dexamethasone on core fucosylation of TGFBR1 and TGFBR2. Furthermore, indirect TGF-β1 binding assay showed that overexpression of FUT8 remarkably promoted TGF-β1 binding to TGFBRs in MC3T3-E1 cells in the presence of high-dose dexamethasone.ConclusionsTaken together, these results suggest that overexpression of FUT8 facilitates counteracting the inhibitory effect of dexamethasone on TGF-β signaling and osteogenesis.

Lab on fiber nano-cavity integrated with charge responsive microgels for biosensing

The development of photonic-based point of care (POC) devices requires the use of compact integrated biosensing platforms enabling reliable detection performances in small volumes. The sensing system should be designed to overcome limitations of benchtop technologies in terms of portability and the easy-of-use. The Lab-on-Fiber (LOF) technology is continuously trying to develop miniaturized, lightweight, and light-coupled optrodes with unprecedented detection performances associated with portable, plug-and-play and user-friendly features. The sensing platforms, yet successfully developed in planar configurations, are being reimagined and suitably scaled down to be directly integrated onto optical fibers. In the wake of this ‘chip-to-fiber translation’ vision, we have realized a micro-sized optical cavity made of smart materials based on synthetic microgels and integrated on the top of an optical fiber. As a specific case study, the optrode has been here tested within a ‘general purpose’ detection assay for biochemical applications. Specifically, we detected nanomolar concentrations of homopolyarginines and positively charged linear polymers that are frequently exploited as surplus in indirect binding assays. The high degree of miniaturization of our probe enables a significant improvement in the minimum detectable concentration (at least 1 nM) compared to those achieved with similar devices having planar configurations. The probe sensitivity and overall performance are strongly influenced by the molecular weight and number of positive charges of the polymer. Overall, our findings represent a step forward in establishing LOF optrodes as sensing units for optical POC, further enlarging their applicability for the detection of a broad range of relevant biomarkers.

Current models of apolipoprotein A-I lipidation by adenosine triphosphate binding cassette transporter A1.

The primary cardioprotective function of high-density lipoprotein (HDL) is to remove excess cellular free cholesterol (FC) from peripheral tissues and deliver it to the liver. Here, we summarize recent research that examines apolipoprotein A-I (apoA-I) lipidation models by adenosine triphosphate binding cassette transporter A1 (ABCA1) and discuss its relevance in atherosclerotic cardiovascular disease (ASCVD). The first step in HDL formation involves the interaction between apoA-I and ABCA1, where ABCA1 mediates the removal of FC and phospholipids from lipid-laden macrophages to form discoidal nascent HDL (nHDL). However, there are currently no clear-cut systematic models that characterize HDL formation. A number of recent studies have investigated the importance of apoA-I C- and N-terminal domains required for optimal cholesterol efflux and nHDL production. Furthermore, functional ABCA1 is required for direct or indirect binding to apoA-I where ABCA1 dimer-monomer interconversion facilitates apoA-I lipidation from plasma membrane microdomains. Microparticles are also another lipid source for apoA-I solubilization into nHDL. ApoA-I and ABCA1 are key factors in macrophage-mediated cholesterol efflux and nHDL production. Understanding of the key steps in HDL formation may unlock the therapeutic potential of HDL and improve clinical management of ASCVD.

Experimental study of a 3D printed permanent implantable porous Ta-coated bone plate for fracture fixation

Metal plates have always been the gold standard in the clinic for internal fracture fixation due to their high strength advantages. However, high elastic modulus can cause stress shielding and lead to bone embrittlement. This study used an electron beam melting method to prepare personalized porous Ti6Al4V (pTi) bone plates. Then, chemical vapor deposition (CVD) technology coats tantalum (Ta) metal on the pTi bone plates. The prepared porous Ta-coated bone plate has an elastic modulus similar to cortical bone, and no stress shielding occurred. In vitro experiments showed that compared with pTi plates, Ta coating significantly enhances the attachment and proliferation of cells on the surface of the scaffold. To better evaluate the function of the Ta-coated bone plate, animal experiments were conducted using a coat tibia fracture model. Our results showed that the Ta-coated bone plate could effectively fix the fracture. Both imaging and histological analysis showed that the Ta-coated bone plate had prominent indirect binding of callus formation. Histological results showed that new bone grew at the interface and formed good osseointegration with the host bone. Therefore, this study provides an alternative to bio-functional Ta-coated bone plates with improved osseointegration and osteogenic functions for orthopaedic applications.

Mechanisms of Binding Specificity among bHLH Transcription Factors.

The transcriptome of every cell is orchestrated by the complex network of interaction between transcription factors (TFs) and their binding sites on DNA. Disruption of this network can result in many forms of organism malfunction but also can be the substrate of positive natural selection. However, understanding the specific determinants of each of these individual TF-DNA interactions is a challenging task as it requires integrating the multiple possible mechanisms by which a given TF ends up interacting with a specific genomic region. These mechanisms include DNA motif preferences, which can be determined by nucleotide sequence but also by DNA’s shape; post-translational modifications of the TF, such as phosphorylation; and dimerization partners and co-factors, which can mediate multiple forms of direct or indirect cooperative binding. Binding can also be affected by epigenetic modifications of putative target regions, including DNA methylation and nucleosome occupancy. In this review, we describe how all these mechanisms have a role and crosstalk in one specific family of TFs, the basic helix-loop-helix (bHLH), with a very conserved DNA binding domain and a similar DNA preferred motif, the E-box. Here, we compile and discuss a rich catalog of strategies used by bHLH to acquire TF-specific genome-wide landscapes of binding sites.

SARS-CoV-2 antibody persistence in COVID-19 convalescent plasma donors: Dependency on assay format and applicability to serosurveillance.

BackgroundAntibody response duration following severe acute respiratory syndrome coronavirus 2 infection tends to be variable and depends on severity of disease and method of detection.Study design and methodsCOVID‐19 convalescent plasma from 18 donors was collected longitudinally for a maximum of 63–129 days following resolution of symptoms. All the samples were initially screened by the Ortho total Ig test to confirm positivity and subsequently tested with seven additional direct sandwich or indirect binding assays (Ortho, Roche, Abbott, Broad Institute) directed against a variety of antigen targets (S1, receptor binding domain, and nucleocapsid [NC]), along with two neutralization assays (Broad Institute live virus PRNT and Vitalant Research Institute [VRI] Pseudovirus reporter viral particle neutralization [RVPN]).ResultsThe direct detection assays (Ortho total Ig total and Roche total Ig) showed increasing levels of antibodies over the time period, in contrast to the indirect IgG assays that showed a decline. Neutralization assays also demonstrated declining responses; the VRI RVPN pseudovirus had a greater rate of decline than the Broad PRNT live virus assay.DiscussionThese data show that in addition to variable individual responses and associations with disease severity, the detection assay chosen contributes to the heterogeneous results in antibody stability over time. Depending on the scope of the research, one assay may be preferable over another. For serosurveillance studies, direct, double Ag‐sandwich assays appear to be the best choice due to their stability; in particular, algorithms that include both S1‐ and NC‐based assays can help reduce the rate of false‐positivity and discriminate between natural infection and vaccine‐derived seroreactivity.

Effects of sheared chromatin length on ChIP-seq quality and sensitivity.

Chromatin immunoprecipitation followed by massively parallel, high throughput sequencing (ChIP-seq) is the method of choice for genome-wide identification of DNA segments bound by specific transcription factors or in chromatin with particular histone modifications. However, the quality of ChIP-seq datasets varies widely, with a substantial fraction being of intermediate to poor quality. Thus, it is important to discern and control the factors that contribute to variation in ChIP-seq. In this study, we focused on sonication, a user-controlled variable, to produce sheared chromatin. We systematically varied the amount of shearing of fixed chromatin from a mouse erythroid cell line, carefully measuring the distribution of resultant fragment lengths prior to ChIP-seq. This systematic study was complemented with a retrospective analysis of additional experiments. We found that the level of sonication had a pronounced impact on the quality of ChIP-seq signals. Over-sonication consistently reduced quality, while the impact of under-sonication differed among transcription factors, with no impact on sites bound by CTCF but frequently leading to the loss of sites occupied by TAL1 or bound by POL2. The bound sites not observed in low-quality datasets were inferred to be a mix of both direct and indirect binding. We leveraged these findings to produce a set of CTCF ChIP-seq datasets in rare, primary hematopoietic progenitor cells. Our observation that the amount of chromatin sonication is a key variable in success of ChIP-seq experiments indicates that monitoring the level of sonication can improve ChIP-seq quality and reproducibility and facilitate ChIP-seq in rare cell types.

Anion effect of Cl−, I−, and F− on counterions condensation within nucleic acid ion atmosphere

In the present work, molecular dynamics simulations have been carried out to study the anion effect on counterions distribution around the DNA double helix on the character of ion hydration. The simulated systems consisted of DNA fragment d(CGCGAATTCGCG) in water solution with the counterions and anions Cl−, I− or F−. Ion distribution and the characteristic binding sites of the counterions with DNA have been determined. The results show that due to the interaction with anions average number of counterions nearly DNA surface undergo changes. The amount of Na+ experienced a decline and the number of K+ experienced an increase with the change of Cl−, I−, and F− anion environment, while Na+ content remained at a high level compared with K+, anions are less present near DNA. Due to the dehydration effect, smaller Na+ counterions can squeeze through the hydration shell of DNA to the bottom of the double helix grooves. Two counterions are mainly indirect binding DNA, which prefer to bind at the phosphate backbone compared to the major groove and negligible binding occurs in the minor groove. The K+ counterions show stronger ability to cluster with anions thus more likely to be shielded from DNA electrostatic field. The character of ion size, ion-DNA interaction, and the ion hydration establishes the correlation between the counterions and anions within nucleic acid ion atmosphere.

Thyroid Hormones Interaction With Immune Response, Inflammation and Non-thyroidal Illness Syndrome.

The interdependence between thyroid hormones (THs), namely, thyroxine and triiodothyronine, and immune system is nowadays well-recognized, although not yet fully explored. Synthesis, conversion to a bioactive form, and release of THs in the circulation are events tightly supervised by the hypothalamic–pituitary–thyroid (HPT) axis. Newly synthesized THs induce leukocyte proliferation, migration, release of cytokines, and antibody production, triggering an immune response against either sterile or microbial insults. However, chronic patho-physiological alterations of the immune system, such as infection and inflammation, affect HPT axis and, as a direct consequence, THs mechanism of action. Herein, we revise the bidirectional crosstalk between THs and immune cells, required for the proper immune system feedback response among diverse circumstances. Available circulating THs do traffic in two distinct ways depending on the metabolic condition. Mechanistically, internalized THs form a stable complex with their specific receptors, which, upon direct or indirect binding to DNA, triggers a genomic response by activating transcriptional factors, such as those belonging to the Wnt/β-catenin pathway. Alternatively, THs engage integrin αvβ3 receptor on cell membrane and trigger a non-genomic response, which can also signal to the nucleus. In addition, we highlight THs-dependent inflammasome complex modulation and describe new crucial pathways involved in microRNA regulation by THs, in physiological and patho-physiological conditions, which modify the HPT axis and THs performances. Finally, we focus on the non-thyroidal illness syndrome in which the HPT axis is altered and, in turn, affects circulating levels of active THs as reported in viral infections, particularly in immunocompromised patients infected with human immunodeficiency virus.

COVID-19 and Toll-Like Receptor 4 (TLR4): SARS-CoV-2 May Bind and Activate TLR4 to Increase ACE2 Expression, Facilitating Entry and Causing Hyperinflammation.

Causes of mortality from COVID-19 include respiratory failure, heart failure, and sepsis/multiorgan failure. TLR4 is an innate immune receptor on the cell surface that recognizes pathogen-associated molecular patterns (PAMPs) including viral proteins and triggers the production of type I interferons and proinflammatory cytokines to combat infection. It is expressed on both immune cells and tissue-resident cells. ACE2, the reported entry receptor for SARS-CoV-2, is only present on ~1-2% of the cells in the lungs or has a low pulmonary expression, and recently, the spike protein has been proposed to have the strongest protein-protein interaction with TLR4. Here, we review and connect evidence for SARS-CoV-1 and SARS-CoV-2 having direct and indirect binding to TLR4, together with other viral precedents, which when combined shed light on the COVID-19 pathophysiological puzzle. We propose a model in which the SARS-CoV-2 spike glycoprotein binds TLR4 and activates TLR4 signalling to increase cell surface expression of ACE2 facilitating entry. SARS-CoV-2 also destroys the type II alveolar cells that secrete pulmonary surfactants, which normally decrease the air/tissue surface tension and block TLR4 in the lungs thus promoting ARDS and inflammation. Furthermore, SARS-CoV-2-induced myocarditis and multiple-organ injury may be due to TLR4 activation, aberrant TLR4 signalling, and hyperinflammation in COVID-19 patients. Therefore, TLR4 contributes significantly to the pathogenesis of SARS-CoV-2, and its overactivation causes a prolonged or excessive innate immune response. TLR4 appears to be a promising therapeutic target in COVID-19, and since TLR4 antagonists have been previously trialled in sepsis and in other antiviral contexts, we propose the clinical trial testing of TLR4 antagonists in the treatment of severe COVID-19. Also, ongoing clinical trials of pulmonary surfactants in COVID-19 hold promise since they also block TLR4.

T cell receptor interactions with human leukocyte antigen govern indirect peptide selectivity for the cancer testis antigen MAGE-A4

T cell-mediated immunity is governed primarily by T cell receptor (TCR) recognition of peptide-human leukocyte antigen (pHLA) complexes and is essential for immunosurveillance and disease control. This interaction is generally stabilized by interactions between the HLA surface and TCR germline-encoded complementarity-determining region (CDR) loops 1 and 2, whereas peptide selectivity is guided by direct interactions with the TCR CDR3 loops. Here, we solved the structure of a newly identified TCR in complex with a clinically relevant peptide derived from the cancer testis antigen melanoma antigen-A4 (MAGE-A4). The TCR bound pHLA in a position shifted toward the peptide's N terminus. This enabled the TCR to achieve peptide selectivity via an indirect mechanism, whereby the TCR sensed the first residue of the peptide through HLA residue Trp-167, which acted as a tunable gateway. Amino acid substitutions at peptide position 1 predicted to alter the HLA Trp-167 side-chain conformation abrogated TCR binding, indicating that this indirect binding mechanism is essential for peptide recognition. These findings extend our understanding of the molecular rules that underpin antigen recognition by TCRs and have important implications for the development of TCR-based therapies.

Tristetraprolin-RNA interaction map reveals a novel TTP-RelB regulatory network for innate immunity gene expression

Tristetraprolin (TTP) regulates inflammatory and immune responses by destabilizing target mRNAs via binding to their 3′-UTR AREs. We have recently reported that TTP preferentially up-regulates the expression level of innate immunity genes involved in the type I interferon-mediated signaling pathway and viral response in cancer cells. To elucidate the role of TTP-RNA interaction in TTP-mediated upregulation of gene expression, we performed iRIP-seq experiments to obtain the RNA interaction map consisting of direct and indirect binding sites of TTP in HeLa cells. We found substantial TTP binding signals in mRNA regions and the introns. ARE-motif AUUUA is over-represented in TTP binding peaks. Strikingly, AUUUA frequency is high both in 3′UTR and intronic regions, and the intronic peaks were more associated with TTP-regulated genes. Analysis of the over-represented motifs in TTP peaks revealed the high frequencies of UAGG and GUGUG motifs reported for hnRNPA2/B1 and CELF1 respectively in the 3′UTR and introns, and also the UGGAC motif overlapping with the m6A motif GGACU in the CDS regions. We further demonstrated that TTP binds to multiple intronic and exonic sites in the pre-mRNA/mRNA of the transcription factor RelB, correlating with the TTP-upregulated expression of RelB. TTP-up-regulated genes without a TTP binding site, but not those with, are highly enriched in innate immunity pathways and show higher tendency of harboring RelB binding sites in their promoter regions. These findings support a model in which TTP binding of RelB pre-mRNA/mRNA coordinates the RelB upregulation and activation of the innate immunity for antiviral response.

Comparative analysis of ChIP-exo peak-callers: impact of data quality, read duplication and binding subtypes

BackgroundChIP (Chromatin immunoprecipitation)-exo has emerged as an important and versatile improvement over conventional ChIP-seq as it reduces the level of noise, maps the transcription factor (TF) binding location in a very precise manner, upto single base-pair resolution, and enables binding mode prediction. Availability of numerous peak-callers for analyzing ChIP-exo reads has motivated the need to assess their performance and report which tool executes reasonably well for the task.ResultsThis study has focussed on comparing peak-callers that report direct binding events with those that report indirect binding events. The effect of strandedness of reads and duplication of data on the performance of peak-callers has been investigated. The number of peaks reported by each peak-caller is compared followed by a comparison of the annotated motifs present in the reported peaks. The significance of peaks is assessed based on the presence of a motif in top peaks. Indirect binding tools have been compared on the basis of their ability to identify annotated motifs and predict mode of protein-DNA interaction.ConclusionBy studying the output of the peak-callers investigated in this study, it is concluded that the tools that use self-learning algorithms, i.e. the tools that estimate all the essential parameters from the aligned reads, perform better than the algorithms which require formation of peak-pairs. The latest tools that account for indirect binding of TFs appear to be an upgrade over the available tools, as they are able to reveal valuable information about the mode of binding in addition to direct binding. Furthermore, the quality of ChIP-exo reads have important consequences on the output of data analysis.

Cutaneous T-Cell Lymphoma (CTCL) Cell Line-Derived Extracellular Vesicles Contain HERV-W-Encoded Fusogenic Syncytin-1

Cutaneous T-Cell Lymphoma (CTCL) Cell Line-Derived Extracellular Vesicles Contain HERV-W-Encoded Fusogenic Syncytin-1

Modelling bacterial chemotaxis for indirectly binding attractants

In bacterial chemotaxis, chemoattractant molecules may bind either directly or indirectly with receptors within the cell periplasmic space. The indirect binding mechanism, which involves an intermediate periplasmic binding protein, has been reported to increase sensitivity to dilute attractant concentrations as well as range of response. Current mathematical models for bacterial chemotaxis at the population scale do not appear to take the periplasmic binding protein (BP) concentration or the indirect binding mechanics into account. We formulate an indirect binding extension to the existing Rivero equation for chemotactic velocity based on fundamental reversible enzyme kinetics. The formulated indirect binding expression accounts for the periplasmic BP concentration and the dissociation constants for binding between attractant and periplasmic BP, as well as between BP and chemoreceptor. We validate the indirect-binding model using capillary assay simulations of the chemotactic responses of E. coli to the indirectly-binding attractants maltose and AI-2. The predicted response agrees well with experimental data from a number of maltose capillary assay studies conducted in previous literature. The model is also able to achieve good agreement with AI-2 capillary assay data of one study out of two tested. The chemotactic response of E. coli towards AI-2 appears to be of higher complexity due to reports of variable periplasmic BP concentration as well as the low concentration of periplasmic BP relative to the total receptor concentration. Our current model is thus suitable for indirect binding chemotactic response systems with constant periplasmic BP concentration that is significantly larger than the total receptor concentration, such as the response of E. coli towards maltose. Further considerations may be taken into account to model the chemotactic response towards AI-2 with greater accuracy.

Transcription Factor Bound Regions Prediction: Word2Vec Technique with Convolutional Neural Network

Genome-wide epigenomic datasets allow us to validate the biological function of motifs and understand the regulatory mechanisms more comprehensively. How different motifs determine whether transcription factors (TFs) can bind to DNA at a specific position is a critical research question. In this project, we apply computational techniques that were used in Natural Language Processing (NLP) to predict the Transcription Factor Bound Regions (TFBRs) given motif instances. Most existing motif prediction methods using deep neural network apply base sequences with one-hot encoding as an input feature to realize TFBRs identification, contributing to low-resolution and indirect binding mechanisms. However, how the collective effect of motifs on binding sites is complicated to figure out. In our pipeline, we apply Word2Vec algorithm, with names of motifs as an input to predict TFBRs utilizing Convolutional Neural Network (CNN) to realize binary classification, based on the ENCODE dataset. In this regard, we consider different types of motifs as separate “words”, and their corresponding TFBR as the meanings of “sentences”. One “sentence” itself is merely the combination of these motifs, and all “sentences” compose of the whole “passage”. For each binding site, we do the binary classification within different cell types to show the performance of our model in different binding sites and cell types. Each “word” has a corresponding vector in high dimensions, and the distances between each vector can be figured out, so we can extract the similarity between each motif, and the explicit binding mechanism from our model. We apply Convolutional Neural Network (CNN) to extract features in the process of mapping and pooling from motif vectors extracted by Word2Vec Algorithm and gain the result of 87% accuracy at the peak.