Biology subjects

Patel, D.

Publications and source records attributed to Patel, D..

At least 91 records · Page 5Linked to original sources

bioRxiv2021

Structure-function Specialisation of the Interfascicular Matrix in the Human Achilles Tendon

ObjectiveTendon consists of highly aligned collagen-rich fascicles surrounded by interfascicular matrix (IFM). Some tendons act as energy stores to improve locomotion efficiency; these tendons are prone to debilitating injuries, the incidence of which increases with ageing. In equine tendons, energy storage is achieved primarily through specialisation of the IFM. However, no studies have investigated IFM structure-function specialisation in human tendons. Here, we compare the positional anterior tibialis and energy storing Achilles tendons, testing the hypothesis that the Achilles IFM has specialised composition and mechanical properties, which are lost with ageing. MethodsWe used a multidisciplinary combination of mechanical testing, immunolocalisation and proteomics to investigate structure-function specialisations in functionally distinct human tendons and how these are altered with ageing. ResultsThe IFM in the energy storing Achilles tendon is more elastic and fatigue resistant than the IFM in the positional anterior tibialis tendon, with a trend towards decreased fatigue resistance with age in the Achilles IFM. With ageing, alterations occur predominantly to the proteome of the Achilles IFM. ConclusionThe Achilles tendon IFM is specialised for energy storage, and changes to its proteome with ageing are likely responsible for the observed trends towards decreased fatigue resistance. Knowledge of key energy storing specialisations and their changes with ageing offers insight towards developing effective treatments for tendinopathy. Key messagesO_ST_ABSWhat is already known about this subject?C_ST_ABSO_LIEnergy storing tendons in animals and humans are particularly prone to tendinopathy and the incidence increases with increasing age. C_LIO_LIPrevious work in some animal models has shown that the specialisation of tendon properties for energy storage is achieved primarily through adaptation of the interfascicular matrix, with specialisation lost in ageing. However, the structural specialisations that provide the human Achilles tendon with its energy storing ability, and how these are affected by ageing, remain to be established. C_LI What does this study add?O_LIWe demonstrate that the interfascicular matrix in the human Achilles tendon is specialised for energy storing, with increased elastic recoil and fatigue resistance, and that these specialisations are partially lost with ageing, likely due to alterations to the proteome of the interfascicular matrix. C_LI How might this impact on clinical practice or future developments?O_LIShort term, the specialist IFM mechanics we have demonstrated can be detected with new developments in ultrasound functional imaging, offering improved opportunities for contextual tendinopathy diagnostics. Personalised rehabilitation programmes can now be explored and designed specifically to target IFM mechanics. C_LIO_LILonger term, the knowledge of key specialisations in injury prone energy storing tendons and how they are affected by ageing, offers crucial insight towards developing cell or tissue engineering treatments targeted at restoring tendon structure and function post-injury, specifically targeted at the IFM. C_LI

bioengineering↗

bioRxiv2021

High Light and High Temperature Reduce Photosynthesis via Different Mechanisms in the C4 Model Setaria viridis

C4 plants frequently experience damaging high light (HL) and high temperature (HT) conditions in native environments, which reduce growth and yield. However, the mechanisms underlying these stress responses in C4 plants have been under-explored, especially the coordination between mesophyll (M) and bundle sheath (BS) cells. We investigated how the C4 model plant Setaria viridis responded to a four-hour HL or HT treatment at the photosynthetic, transcriptomic, and ultrastructural levels. Although we observed a comparable reduction of photosynthetic efficiency in HL- or HT-treated leaves, detailed analysis of multi-level responses revealed important differences in key pathways and M/BS specificity responding to HL and HT. We provide a systematic analysis of HL and HT responses in S. viridis, reveal different acclimation strategies to these two stresses in C4 plants, discover unique light/temperature responses in C4 plants in comparison to C3 plants, and identify potential targets to improve abiotic stress tolerance in C4 crops.

plant biology↗

bioRxiv2020

In-Silico analysis reveals lower transcription efficiency of C241T variant of SARS-CoV-2 with host replication factors MADP1 and hnRNP-1

Novel severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) has claimed more than 1.5 million lives worldwide and counting. As per the GISAID database, the genomics of SARS-CoV2 is extensively studied with more than 500 genome submissions per day. Out of several hotspot mutations within the SARS-CoV-2 genome, researchers have focused a lot on missense variants but the least work is done on the UTRs. One of the most frequent 5 UTR variants in the SARS-CoV-2 genome is the C241T with a global frequency of more than 0.9. In the present study, the effect of the C241T mutation has been studied with respect to change in RNA structure and its interaction with the host replication factors MADP1 Zinc finger CCHC-type and RNA-binding motif 1 (hnRNP1). The results obtained from molecular docking and molecular dynamics simulation indicated weaker interaction of C241T mutant stem loops with host transcription factor MADP1 indicating reduced replication efficiency. The results are also correlated with increased recovery rates and decreased death rates of global SARS-CoV-2 cases.

bioinformatics↗

bioRxiv2020

CGGBP1-dependent CTCF-binding sites restrict ectopic transcription from SV40 early promoter

Binding sites of the chromatin regulator protein CTCF function as important landmarks in the human genome. The recently characterized CTCF-binding sites at LINE-1 repeats depend on another repeat-regulatory protein CGGBP1. These CGGBP1-dependent CTCF-binding sites serve as potential barrier elements for epigenetic marks such as H3K9me3. Such CTCF-binding sites are associated with asymmetric H3K9me3 levels as well as RNA levels in their flanks. The functions of these CGGBP1-dependent CTCF-binding sites remain un-known. By performing targeted studies on candidate CGGBP1-dependent CTCF-binding sites cloned in an SV40 promoter-enhancer episomal system we show that these regions act as inhibitors of ectopic transcription from the SV40 promoter. CGGBP1-dependent CTCF-binding sites that recapitulate their genomic function of loss of CTCF binding upon CGGBP1 depletion and H3K9me3 asymmetry in immediate flanks are also the ones that show the strongest inhibition of ectopic transcription. By performing a series of strand-specific reverse transcription PCRs we demonstrate that this ectopic transcription results in the synthesis of RNA from the SV40 promoter in a direction opposite to the downstream reporter gene in a strand-specific manner. The unleashing of the bidirectionality of the SV40 promoter activity and a breach of the transcription barrier seems to depend on depletion of CGGBP1 and loss of CTCF binding proximal to the SV40 promoter. RNA-sequencing reveals that CGGBP1-regulated CTCF-binding sites act as barriers to transcription at multiple locations genomewide. These findings suggest a role of CGGBP1-dependent binding sites in restricting ectopic transcription.

molecular biology↗

bioRxiv2020

Homeodomain protein 1 is an essential regulator of gene expression during sexual differentiation of malaria parasites

Transmission of Plasmodium falciparum and other malaria parasites requires their differentiation from asexual blood stages into gametocytes, the non-replicative sexual stage necessary for transmission to the mosquito vector. This transition involves changes in gene expression and chromatin reorganization that result in the activation and silencing of stage-specific genes. However, the genomes of malaria parasites have been noted for their dearth of transcriptional and chromatin regulators and the molecular mediators of these changes remain largely unknown. We recently identified HomeoDomain Protein 1 (HDP1) as a DNA-binding protein that enhances the expression of key genes that are critical for early sexual differentiation. The discovery of a homeodomain-like DNA-binding protein marks a new class of transcriptional regulator in malaria parasites outside of the better-characterized ApiAP2 family. In this study, we show that HDP1 facilitates the necessary upregulation of inner membrane complex components during early gametocytogenesis that gives P. falciparum gametocytes they characteristic shape and is required for gametocyte maturation and parasite transmission.

microbiology↗

bioRxiv2020

Implementing and assessing an alchemical method for calculating protein-protein binding free energy

Protein-protein binding is fundamental to most biological processes. It is important to be able to use computation to accurately estimate the change in protein-protein binding free energy due to mutations in order to answer biological questions that would be experimentally challenging, laborious or time consuming. Although non-rigorous free energy methods are faster, rigorous alchemical molecular dynamics-based methods are considerably more accurate and are becoming more feasible with the advancement of computer hardware and molecular simulation software. Even with sufficient computational resources, there are still major challenges to using alchemical free energy methods for protein-protein complexes, such as generating hybrid structures and topologies, maintaining a neutral net charge of the system when there is a charge-changing mutation, and setting up the simulation. In the current study, we have used the pmx package to generate hybrid structures and topologies, and a double-system/single-box approach to maintain the net charge of the system. To test the approach, we predicted relative binding affinities for two protein-protein complexes using a non-equilibrium alchemical method based on the Crooks fluctuation theorem and compared the results with experimental values. The method correctly identified stabilizing from destabilizing mutations for a small protein-protein complex, but was not as successful to the larger, more challenging antibody complex. In addition, the correlation between predicted and experimental relative binding affinities was high for smaller complex, and low for the other larger complex.

biophysics↗

bioRxiv2020

Converting Microwave Ovens into Plasma-Generating Decontamination Units for N-95 Respirators

We show how a common microwave oven, a coat-hanger and a coffee cup can be used to decontaminate N-95 respirators in 30 seconds. Tulane virus in the artificial saliva was reduced by >3 log and Geobacillus stearothermophilus spores were reduced by >6 log. Respirators maintained filtration and fit even after 10 cycles. Filtration and fit tests performed by the CDC confirmed there was no damage to the respirators. Spectroscopy of the plasma reveals that OH and C-containing radicals as the most prevalent active species.

bioengineering↗

bioRxiv2020

Short-wavelength-sensitive 2 (Sws2) visual photopigment models combined with atomistic molecular simulations to predict spectral peaks of absorbance

For many species, vision is one of the most important sensory modalities for mediating essential tasks that include navigation, predation and foraging, predator avoidance, and numerous social behaviors. The vertebrate visual process begins when photons of the light interact with rod and cone photoreceptors that are present in the neural retina. Vertebrate visual photopigments are housed within these photoreceptor cells and are sensitive to a wide range of wavelengths that peak within the light spectrum, the latter of which is a function of the type of chromophore used and how it interacts with specific amino acid residues found within the opsin protein sequence. Minor differences in the amino acid sequences of the opsins are known to lead to large differences in the spectral peak of absorbance (i.e. the {lambda}max value). In our prior studies, we developed a new approach that combined homology modeling and molecular dynamics simulations to gather structural information associated with chromophore conformation, then used it to generate statistical models for the accurate prediction of {lambda}max values for photopigments derived from Rh1 and Rh2 amino acid sequences. In the present study, we test our novel approach to predict the {lambda}max of phylogenetically distant Sws2 cone opsins. To build a model that can predict the {lambda}max using our approach presented in our prior studies, we selected a spectrally-diverse set of 11 teleost Sws2 photopigments for which both amino acid sequence information and experimentally measured {lambda}max values are known. The final first-order regression model, consisting of three terms associated with chromophore conformation, was sufficient to predict the {lambda}max of Sws2 photopigments with high accuracy. This study further highlights the breadth of our approach in reliably predicting {lambda}max values of Sws2 cone photopigments, evolutionary-more distant from template bovine RH1, and provided mechanistic insights into the role of known spectral tuning sites. Author SummaryIn vertebrates, color vision depends on the complement of cone visual photopigments that have different spectral peaks of absorbance ({lambda}max) within the cone population. Together, the type of chromophore and the amino acid sequence of the opsin protein directly affect the {lambda}max value. To understand this relationship further at a structural level, we previously developed a new molecular modeling approach to study Rh1 and Rh2 opsin classes by combining homology modeling, molecular dynamics simulations to extract structural parameters of chromophore conformations and statistical modeling. Here, we used this novel modeling approach to accurately predict the {lambda}max values for teleost Sws2 photopigments. Such a genome-to-phenome approach for predicting visual pigment function will be of great interest to evolutionary biologists, vision scientists, and molecular modelers, to better understand the diversity and mechanisms of sensory function. Moreover, it will pave the way for novel strategies to forward engineer visual pigments suitable for optogenetics applications.

biophysics↗

bioRxiv2020

Does plant root architecture respond to potassium nutrition under water stress? A case from rice seedling root system architectural and growth responses.

The root is the sensing organ for potassium (K) and water availability. We evaluated whether K availability influences root architecture and contributes to drought tolerance under moisture stress. Rice seedling growth was severely affected by low K availability under water stress, and the substantial reductions in root projected area, maximum width, and width to depth ratio were observed. High K availability helps maintain root top and bottom angles and reduces root steepness under mild water stress, but over K nutrition does not ensure higher seedling growth. Under severe water stress, the steepness was more regulated by water than K availability.

plant biology↗

bioRxiv2020

Linkage mapping and QTL analysis of flowering time using ddRAD sequencing with genotype error correction in Brassica napus

Brassica napus is an important oilseed crop cultivated worldwide. During domestication and breeding of B. napus, flowering time has been a target of selection because of its substantial impact on yield. Here we use double digest restriction-site associated DNA sequencing (ddRAD) to investigate the genetic basis of flowering in B. napus. An F2 mapping population was derived from a cross between an early-flowering spring type and a late-flowering winter type. Flowering time in the mapping population differed by up to 25 days between individuals. High genotype error rates persisted after initial quality controls, as suggested by a genotype discordance of ∼12% between biological sequencing replicates. After genotype error correction, a linkage map spanning 3,605.70 cM and compromising 14,630 single nucleotide polymorphisms (SNPs) was constructed. A quantitative trail locus (QTL) on chromosome C2 was detected in the vicinity of flowering time genes including FT and FLC. These findings demonstrate the effectiveness of the ddRAD approach to sample the B. napus genome. Our results also suggest that ddRAD genotype error rates can be higher than expected in F2 populations. Quality filtering and genotype correction and imputation can substantially reduce these error rates and allow effective linkage mapping and QTL analysis.Competing Interest StatementBASF supported this work and employs SR.View Full Text

plant biology↗

bioRxiv2020

CGGBP1-regulated cytosine methylation at CTCF-binding motifs resists stochasticity

The human CGGBP1 is implicated in a variety of cellular functions. It regulates genomic integrity, cell cycle, gene expression and cellular response to growth signals. Evidence suggests that these functions of CGGBP1 manifest through binding to GC-rich regions in the genome and regulation of interspersed repeats. Recent works show that CGGBP1 is needed for cytosine methylation homeostasis and genome-wide occupancy patterns of the epigenome regulator protein CTCF. It has remained unknown if cytosine methylation regulation and CTCF occupancy regulation by CGGBP1 are independent or interdependent processes. By sequencing immunoprecipitated methylated DNA, we have found that some transcription factor-binding sites resist stochastic changes in cytosine methylation. Of these, we have analyzed the CTCF-binding sites thoroughly and show that cytosine methylation regulation at CTCF-binding DNA sequence motifs by CGGBP1 is deterministic. These CTCF-binding sites are positioned at locations where the spread of cytosine methylation in cis depends on the levels of CGGBP1. Our findings suggest that CTCF occupancy and functions are determined by CGGBP1-regulated cytosine methylation patterns.

genomics↗

bioRxiv2020

Single exposure to social defeat or immobilization stress fails to induce lasting alterations in general anxiety and hippocampal neurogenesis in Wistar and wild-type Groningen rats

Suppression of hippocampal neurogenesis is a readout for stress-induced alterations in neuroplasticity. In this study, we hypothesized that a single episode of severe social or non-social stress would differentially suppress neurogenesis in the dentate gyrus (DG) 10 days later in two rat strains. We anticipated that the suppression following social stress would be less severe in wildtype Groningen (WTG) rats, a rat strain considered relatively resilient to social stressors. Male Wistar and WTG were subjected to either social defeat or to immobilization stress. Behavioral response to social defeat and acute corticosterone response to both stressors was measured as well as anxiety behavior 10 days later on the elevated plus maze. Subsequently, brains were collected following cardiac aldehyde perfusion. The behavioral freezing response to defeat was much stronger in Wistar rats as compared to WTG rats. Acute corticosteroid response was similar in both strains although Wistar rats more rapidly resumed baseline values. There was no significant effect of both stressors on hippocampal DG cell proliferation and differentiation as well as on anxiety behavior. However, a striking strain difference appeared in anxiety behavior and both markers of neurogenesis. The WTG strain exhibiting much lower anxiety as well as reduced rate of hippocampal neurogenesis under all treatments. The results in this study suggest that both short-lasting acute stressors failed to induce lasting anxiety or decreased neurogenesis in the DG. Future studies could explore if and how rate of hippocampal neurogenesis is related with behavioral coping with stress.

neuroscience↗

bioRxiv2020

HDAC inhibitor Givinostat targets DNA-binding of human CGGBP1

The antineoplastic agent Givinostat inhibits histone deacetylases. We present here our finding that the DNA-binding of human CGGBP1 is also inhibited by Givinostat. CGGBP1, a DNA-binding protein, is required for cancer cell proliferation. In our quest to exploit the potential anti-proliferative effects of CGGBP1 inhibition, we have developed a simple screening assay to identify chemical inhibitors of DNA-protein interactions. We have applied this screen for human CGGBP1 on a library of 1685 compounds and found that Givinostat is a direct inhibitor of CGGBP1-DNA interaction. The mechanism of action of Givinostat should thus extend beyond HDACs to include the inhibition of the myriad functions of CGGBP1 that depend on its binding to the DNA.

cancer biology↗

bioRxiv2019

Optimizing a human fecal assay that elicits bacterial swarming

A distinct property of many bacteria is swarming: swift movement across a surface through flagella propulsion. Early research indicates that bacterial swarming can be a protective host response to intestinal inflammation. Central to the further study of bacterial swarming in human health is an effective and replicable assay for swarming that can accommodate complex material, such as fecal matter. To date, nearly all swarming assays described in the literature are specific for bacteria grown in culture, most often Pseudomonas. In this paper, we describe a protocol for discerning swarming of bacteria from frozen human fecal samples. Moreover, we tested 4 variables that may influence the effectiveness of the assay: the method by which frozen samples were thawed, the concentration of agar used in the Lysogenic broth (LB) agar plate, the volume of LB agar poured in the plate, and the volume of sample inoculated. We found that while the type of thaw and volume of LB agar had little to no effect on swarming, greater concentrations of agar were negatively correlated with swarming and greater volumes of the sample were positively correlated with swarming.

microbiology↗

bioRxiv2019

Spectroscopy and Machine Learning Based Rapid Point-of-Care Assessment of Core Needle Cancer Biopsies

Solid tumor needle biopsies are essential to confirm malignancy and assess for actionable characteristics or genetic alterations to guide treatment selection. Ensuring that sufficient and suitable material is acquired for tumor profiling, while minimizing patient risk, remains a critical unmet need. Here, we evaluated the performance characteristics of transmission optical spectroscopy for rapid identification of malignant tissue in core needle biopsies (CNB). Human kidney biopsy specimens (545 CNB from 102 patients, 5583 spectra for analysis) were analyzed directly on core biopsy needles with a custom-built optical spectroscopy instrument. Machine learning classifiers were trained to differentiate malignant from normal tissue spectra. Classifiers were compared using receiver operating characteristics analysis and sensitivity and specificity were calculated relative to a histopathologic gold standard. The best performing algorithm was the random forest (sensitivity 96% and 93%, specificity 90% and 93% at the level of individual spectra and full CNB, respectively). Ex-vivo spectroscopy paired with machine learning paves the way towards rapid and accurate characterization of CNB at the time of tissue acquisition and improving tumor biopsy quality.

bioinformatics↗

bioRxiv2019

Transcription factor NF-κB in a basal metazoan, the sponge, has conserved and unique sequences, activities, and regulation

Biological and biochemical functions of immunity transcription factor NF-{kappa}B in basal metazoans are largely unknown. Herein, we characterize transcription factor NF-{kappa}B from the demosponge Amphimedon queenslandica (Aq), in the phylum Porifera. Structurally and phylogenetically, the Aq-NF-{kappa}B protein is most similar to NF-{kappa}B p100 and p105 among vertebrate proteins, with an N-terminal DNA-binding/dimerization domain, a C-terminal Ankyrin (ANK) repeat domain, and a DNA binding-site profile more similar to human NF-{kappa}B proteins than Rel proteins. Aq-NF-{kappa}B also resembles the mammalian NF-{kappa}B protein p100 in that C-terminal truncation results in translocation of Aq-NF-{kappa}B to the nucleus and increases its transcriptional activation activity. Overexpression of a human or sea anemone I{kappa}B kinase (IKK) can induce C-terminal processing of Aq-NF-{kappa}B in vivo, and this processing requires C-terminal serine residues in Aq-NF-{kappa}B. Unlike human NF-{kappa}B p100, however, the C-terminal sequences of Aq-NF-{kappa}B do not effectively inhibit its DNA-binding activity when expressed in human cells. Tissue of another demosponge, a black encrusting sponge, contains NF-{kappa}B site DNA-binding activity and an NF-{kappa}B protein that appears mostly processed and in the nucleus of cells. NF-{kappa}B DNA-binding activity and processing is increased by treatment of sponge tissue with LPS. By transcriptomic analysis of A. queenslandica we identified likely homologs to many upstream NF-{kappa}B pathway components. These results present a functional characterization of the most ancient metazoan NF-{kappa}B protein to date, and show that many characteristics of mammalian NF-{kappa}B are conserved in sponge NF-{kappa}B, but the mechanism by which NF-{kappa}B functions and is regulated in the sponge may be somewhat different.

cell biology↗

bioRxiv2019

Categorical Assignment of Pulmonary Embolism is a Simple and More Accurate Indicator of Right Ventricular Dysfunction and Short Team Mortality

Several risk stratification tools are available to predict short-term mortality in patients with acute pulmonary embolism (PE). Right ventricular (RV) dysfunction, which is common to intermediate and high risk PE, is an independent predictor of mortality and may be a faster and simpler way to assess patient risk in acute care settings. We evaluated 571 patients presenting with acute PE as the primary diagnosis, stratifying them by the Pulmonary Embolism Severity Index (PESI), by the BOVA score, or categorically as low risk (no RV dysfunction by imaging), intermediate risk (RV dysfunction by imaging), or high risk PE (RV dysfunction by imaging with sustained hypotension). Using imaging data to firstly define the presence of RV dysfunction, and plasma cardiac troponin T (cTnT) and NT-proBNP as additional evidence for myocardial strain, we evaluated the PESI and BOVA scoring systems compared to categorical assignment of PE as low risk, submassive, and massive PE. Cardiac biomarkers poorly distinguished between PESI classes and BOVA stages in patients with acute PE. Cardiac TnT and NT-proBNP easily distinguished low risk from submassive PE with an area under the curve (AUC) of 0.84 (95% C.I. 0.73 - 0.95, p< 0.0001), and 0.88 (95% C.I. 0.79-0.97, p< 0.0001), respectively, and low risk from massive PE with an area under the curve (AUC) of 0.89 (95% C.I. 0.78 - 1.00, p< 0.0001), and 0.89 (95% C.I. 0.82-0.95, p< 0.0001), respectively. Predicted short-term mortality by PESI score or BOVA stage was lower than the observed mortality for submassive PE by a two-fold order of magnitude. These data suggest the presence of RV dysfunction in the context of acute PE is sufficient for the purposes of risk stratification, while more complicated risk stratification algorithms may under-estimate short-term mortality risk.

biochemistry↗

bioRxiv2019

The nascent RNA binding complex SFiNX licenses piRNA-guided heterochromatin formation

The PIWI-interacting RNA (piRNA) pathway protects animal genome integrity in part through establishing repressive heterochromatin at transposon loci. Silencing requires piRNA-guided targeting of nuclear PIWI proteins to nascent transposon transcripts, yet the subsequent molecular events are not understood. Here, we identify SFiNX (Silencing Factor interacting Nuclear eXport variant), an interdependent protein complex required for Piwi-mediated co-transcriptional silencing in Drosophila. SFiNX consists of Nxf2-Nxt1, a gonad-specific variant of the heterodimeric mRNA export receptor Nxf1-Nxt1, and the Piwi-associated protein Panoramix. SFiNX mutant flies are sterile and exhibit transposon de-repression because piRNA-loaded Piwi is unable to establish heterochromatin. Within SFiNX, Panoramix recruits the heterochromatin effectors, while the RNA binding Nxf2 protein licenses co-transcriptional silencing. Our data reveal how Nxf2 evolved from an RNA transport receptor into a co-transcriptional silencing factor. Thus, NXF-variants, which are abundant in metazoans, can have diverse molecular functions and might have been co-opted for host genome defense more broadly.

molecular biology↗