Biology subjects

Singh, B.

Publications and source records attributed to Singh, B..

At least 91 records · Page 5Linked to original sources

bioRxiv2020

IFN signaling and neutrophil degranulation transcriptional signatures are induced during SARS-CoV-2 infection

The novel virus SARS-CoV-2 has infected more than 14 million people worldwide resulting in the Coronavirus disease 2019 (COVID-19). Limited information on the underlying immune mechanisms that drive disease or protection during COVID-19 severely hamper development of therapeutics and vaccines. Thus, the establishment of relevant animal models that mimic the pathobiology of the disease is urgent. Rhesus macaques infected with SARS-CoV-2 exhibit disease pathobiology similar to human COVID-19, thus serving as a relevant animal model. In the current study, we have characterized the transcriptional signatures induced in the lungs of juvenile and old rhesus macaques following SARS-CoV-2 infection. We show that genes associated with Interferon (IFN) signaling, neutrophil degranulation and innate immune pathways are significantly induced in macaque infected lungs, while pathways associated with collagen formation are downregulated. In COVID-19, increasing age is a significant risk factor for poor prognosis and increased mortality. We demonstrate that Type I IFN and Notch signaling pathways are significantly upregulated in lungs of juvenile infected macaques when compared with old infected macaques. These results are corroborated with increased peripheral neutrophil counts and neutrophil lymphocyte ratio in older individuals with COVID-19 disease. In contrast, pathways involving VEGF are downregulated in lungs of old infected macaques. Using samples from humans with SARS-CoV-2 infection and COVID-19, we validate a subset of our findings. Finally, neutrophil degranulation, innate immune system and IFN gamma signaling pathways are upregulated in both tuberculosis and COVID-19, two pulmonary diseases where neutrophils are associated with increased severity. Together, our transcriptomic studies have delineated disease pathways to improve our understanding of the immunopathogenesis of COVID-19 to facilitate the design of new therapeutics for COVID-19.

immunology↗

bioRxiv2020

Evidence for the plant recruitment of beneficial microbes to suppress soil-borne pathogen

O_LIEmerging experimental framework suggests that plants under biotic stress may actively seek help from soil microbes, but empirical evidence underlying such a cry for help strategy is limited. C_LIO_LIWe used integrated microbial community profiling, pathogen and plant transcriptive gene quantification and culture-based methods to systematically investigate a three-way interaction between the wheat plant, wheat-associated microbiomes and Fusarium pseudograminearum (Fp). C_LIO_LIA clear enrichment of a dominant bacterium, Stenotrophomonas rhizophila (SR80), was observed in both the rhizosphere and root endosphere of Fp-infected wheat. SR80 reached 3.7x107 cells g-1 in the rhizosphere and accounted for up to 11.4% of the microbes in the root endosphere. Its abundance had a positive linear correlation with the pathogen load at base stems and expression of multiple defense genes in top leaves. Upon re-introduction in soils, SR80 enhanced plant growth, both the below- and above-ground, and induced strong disease resistance by priming plant defense in the aboveground plant parts, but only when the pathogen was present C_LIO_LITogether, the bacterium SR80 seems to have acted as an early warning system for plant defense. This work provides novel evidence for the potential protection of plants against pathogens by an enriched beneficial microbe via modulation of the plant immune system. C_LI

microbiology↗

bioRxiv2020

SARS-CoV2 genome analysis of Indian isolates and molecular modelling of D614G mutated spike protein with TMPRSS2 depicted its enhanced interaction and virus infectivity

COVID-19 that emerged as a global pandemic is caused by SARS-CoV-2 virus. The virus genome analysis during disease spread reveals about its evolution and transmission. We did whole genome sequencing of 225 clinical strains from the state of Odisha in eastern India using ARTIC protocol-based amplicon sequencing. Phylogenetic analysis identified the presence of all five reported clades 19A, 19B, 20A, 20B and 20C in the population. The analyses revealed two major routes for the introduction of the disease in India i.e. Europe and South-east Asia followed by local transmission. Interestingly, 19B clade was found to be much more prevalent in our sequenced genomes (17%) as compared to other genomes reported so far from India. The haplogroup analysis for clades showed evolution of 19A and 19B in parallel whereas the 20B and 20C appeared to evolve from 20A. Majority of the 19A and 19B clades were present in cases that migrated from Gujarat state in India suggesting it to be one of the major initial points of disease transmission in India during month of March and April. We found that with the time 20A and 20B clades evolved drastically that originated from central Europe. At the same time, it has been observed that 20A and 20B clades depicted selection of four common mutations i.e. 241 C>T (5UTR), P323L in RdRP, F942F in NSP3 and D614G in the spike protein. We found an increase in the concordance of G614 mutation evolution with the viral load in clinical samples as evident from decreased Ct value of spike and Orf1ab gene in qPCR. Molecular modelling and docking analysis identified that D614G mutation enhanced interaction of spike with TMPRSS2 protease, which could impact the shedding of S1 domain and infectivity of the virus in host cells.

genomics↗

bioRxiv2020

AE-LGBM: Sequence-Based Novel Approach To Detect Interacting Protein Pairs via Ensemble of Autoencoder and LightGBM.

Protein-protein interactions (PPIs) are a vital phenomenon for every biological process. Prediction of PPI can be very helpful in the probing of protein functions which can further help in the development of new and powerful therapy designs for disease prevention. A lot of experimental studies have been done previously to study PPIs. However, lab-based experimental studies of PPI prediction are resource-extensive and time-consuming. In recent years, several high throughput, computational approaches to predict PPI have been developed but they could be fallible in terms of accuracy and false-positive rate. To overcome these shortcomings, we propose a novel approach AE-LGBM to predict the PPI more accurately. This method is based on the LightGBM classifier and utilizes the Autoencoder, which is an artificial neural network, to efficiently produce lower-dimensional, discriminative, and noise-free features. We incorporate conjoint triad (CT) features along with Composition-Transition-Distribution (CTD) features into the model and obtained promising results. The ten-fold cross-validation results indicate that the prediction accuracies obtained for Human and Yeast datasets are 98.7% and 95.4% respectively. This method was further evaluated on other datasets and has achieved excellent accuracies of 100%, 100%, 99.9%, 99.2% on E.coli, M.musculus, C.elegans, and H.sapiens respectively. We also executed AE-LGBM over three important PPI networks namely, single-core network (CD9), the multiple-core network (The Ras/Raf/MEK/ERK pathway), and the cross-connection network (Wnt Network). The method was successful in predicting the pathway with an impressive accuracy of 100%, 100%, and 98.9% respectively. These figures are significantly higher than previous methods that are based on state-of-the-art models and models including LightGBM or Autoencoder, proving AE-LGBM to be highly versatile, efficient, and robust.Competing Interest StatementThe authors have declared no competing interest.View Full Text

bioinformatics↗

bioRxiv2020

De novo genome assembly and transcriptome analysis for the drought and salt resistant Solanum sitiens

Solanum sitiens is a self-incompatible wild relative of tomato, characterised by salt and drought resistance traits, with the potential to contribute to crop improvement in cultivated tomato. This species has a distinct morphology, classification and ecotype compared to other stress resistant wild tomato relatives such as S. pennellii and S. chilense. Therefore, the availability of a high-quality reference genome for S. sitiens will facilitate the genetic and molecular understanding of salt and drought resistance. Here, we present a de novo genome and transcriptome assembly for S. sitiens (Accession LA1974). A hybrid assembly strategy was followed using Illumina short reads ([~]159X coverage) and PacBio long reads ([~]44X coverage), generating a total of [~]262 Gbp of DNA sequence; in addition, [~]2,670 Gbp of BioNano data was obtained. A reference genome of 1,245 Mbp, arranged in 1,481 scaffolds with a N50 of 1,826 Mbp was generated. Genome completeness was estimated at 95% using the Benchmarking Universal Single-Copy Orthologs (BUSCO) and the K-mer Analysis Tool (KAT); this is within the range of current high-quality reference genomes for other tomato wild relatives. Additionally, we identified three large inversions compared to S. lycopersicum, containing several drought resistance related genes, such as beta-amylase 1 and YUCCA7. In addition, [~]63 Gbp of RNA-Seq were generated to support the prediction of 31,164 genes from the assembly, and perform a de novo transcriptome. Some of the protein clusters unique to S. sitiens were associated with genes involved in drought and salt resistance, including GLO1 and FQR1. This first reference genome for S. sitiens will provide a valuable resource to progress QTL studies to the gene level, and will assist molecular breeding to improve crop production in water-limited environments.

bioinformatics↗

bioRxiv2020

SARS-CoV-2 infection leads to acute infection with dynamic cellular and inflammatory flux in the lung that varies across nonhuman primate species

There are no known cures or vaccines for COVID-19, the defining pandemic of this era. Animal models are essential to fast track new interventions and nonhuman primate (NHP) models of other infectious diseases have proven extremely valuable. Here we compare SARS-CoV-2 infection in three species of experimentally infected NHPs (rhesus macaques, baboons, and marmosets). During the first 3 days, macaques developed clinical signatures of viral infection and systemic inflammation, coupled with early evidence of viral replication and mild-to-moderate interstitial and alveolar pneumonitis, as well as extra-pulmonary pathologies. Cone-beam CT scans showed evidence of moderate pneumonia, which progressed over 3 days. Longitudinal studies showed that while both young and old macaques developed early signs of COVID-19, both groups recovered within a two-week period. Recovery was characterized by low-levels of viral persistence in the lung, suggesting mechanisms by which individuals with compromised immune systems may be susceptible to prolonged and progressive COVID-19. The lung compartment contained a complex early inflammatory milieu with an influx of innate and adaptive immune cells, particularly interstitial macrophages, neutrophils and plasmacytoid dendritic cells, and a prominent Type I-interferon response. While macaques developed moderate disease, baboons exhibited prolonged shedding of virus and extensive pathology following infection; and marmosets demonstrated a milder form of infection. These results showcase in critical detail, the robust early cellular immune responses to SARS-CoV-2 infection, which are not sterilizing and likely impact development of antibody responses. Thus, various NHP genera recapitulate heterogeneous progression of COVID-19. Rhesus macaques and baboons develop different, quantifiable disease attributes making them immediately available essential models to test new vaccines and therapies.

immunology↗

bioRxiv2020

Transcriptional regulator PAX4 links Receptor Tyrosine Kinases and cytoskeleton stability in Alzheimer's disease and type 2 diabetes

Alzheimers Disease (AD) and Type 2 Diabetes (T2D) share a common hallmark of insulin resistance. Besides Insulin Receptor (IR), two non-canonical RTKs, ALK and RYK, exhibit significant and consistent functional downregulation in post-mortem AD and T2D tissues. Incidentally, both have Grb2 as a common downstream adapter and NOX4 as a common ROS producing factor. Here we show that Grb2 and NOX4 play critical roles in reducing the severity of both the diseases. The study demonstrates that the abundance of Grb2 in degenerative conditions, in conjunction with NOX4, reverse cytoskeletal degradation by counterbalancing the network of small GTPases. PAX4, a transcription factor for both Grb2 and NOX4, emerges as the key link between the common pathways of AD and T2D. Both ALK and RYK downregulation elevate the PAX4 level by reducing its suppressor ARX via Wnt/{beta}-Catenin signaling pathway. For the first time, this study brings together RTKs other than Insulin Receptor (IR), their common transcription factor PAX4 and both AD and T2D pathologies on a common regulatory platform.

neuroscience↗

bioRxiv2020

Identification of miRNAs and their corresponding mRNA targets from chickpea root infected with M. ciceri and functional characterization of candidate miRNAs by overexpression in chickpea roots

O_LILegumes developed symbiotic associations with rhizobia to meet its nitrogen requirement. The nitrogen fixation takes place in root nodules which involves bacterial colonization, organogenesis and nitrogen fixation. C_LIO_LIOne microRNA and four parallel analysis of RNA ends (PARE) libraries were sequenced to unravel the miRNA mediated regulation of symbiosis in chickpea. C_LIO_LISequencing of microRNA library identified a set of 91 miRNAs comprising of 84 conserved and 7 novel miRNAs. Additionally, PARE library analysis revealed 564 genes being targeted by 85 miRNAs. C_LIO_LIPhylogenetic analysis of the precursor sequences of the 91 miRNAs clearly indicated a clustering of two distinct miRNAs in the same clade representing a close ancestral precursor. C_LIO_LIFurther, biogenesis of miRNAs was predicted using the miRNAs identified from different legume genomes. C_LIO_LIThe miRNA reads from the nodule library were also mapped onto bacterial genomes from which bacterial small RNA were predicted. C_LIO_LIThe antagonistic expression of some of the miRNA-target pairs was investigated and the negative co-related expression profiling proved the validity of the libraries and the miRNA-target pairs. Four miRNAs were selected based on the antagonistic expression profiling and were ectopically expressed in chickpea roots by hairy root transformation. C_LIO_LIThe overexpression lines showed significant change in nodule numbers. The target of miR171f (NRK), miR394 (HP) and miR1509 (AK) are novel ones being reported for the first time. This analysis opens a wide arena for investigation of the novel miRNAs and target pairs, polycistronic miRNAs and the bacterial derived smRNAs predicted in this study. C_LI

plant biology↗

bioRxiv2019

Non-Genetic Reprogramming of Monocytes via Microparticle Phagocytosis for Sustained Modulation of Macrophage Phenotype

Monocyte-derived macrophages orchestrate tissue regeneration by homing to sites of injury, phagocytosing pathological debris, and stimulating other cell types to repair the tissue. Accordingly, monocytes have been investigated as a translational and potent source for cell therapy, but their utility has been hampered by their rapid acquisition of a pro-inflammatory phenotype in response to the inflammatory injury microenvironment. To overcome this problem, we designed a cell therapy strategy where we collect and exogenously reprogram monocytes by intracellularly loading the cells with biodegradable microparticles containing an anti-inflammatory drug in order to modulate and maintain an anti-inflammatory phenotype over time. To test this concept, poly(lactic-co-glycolic) acid microparticles were loaded with the anti-inflammatory drug dexamethasone (Dex) and administered to primary human monocytes for four hours to facilitate phagocytic uptake. After removal of non-phagocytosed microparticles, microparticle-loaded monocytes differentiated into macrophages and stored the microparticles intracellularly for several weeks in vitro, releasing drug into the extracellular environment over time. Cells loaded with intracellular Dex microparticles showed decreased expression and secretion of inflammatory factors even in the presence of pro-inflammatory stimuli up to 7 days after microparticle uptake compared to untreated cells or cells loaded with blank microparticles. This study represents a new strategy for long-term maintenance of anti-inflammatory macrophage phenotype using a translational monocyte-based cell therapy strategy without the use of genetic modification. Because of the ubiquitous nature of monocyte-derived macrophage involvement in pathology and regeneration, this strategy holds potential as a treatment for a vast number of diseases and disorders.

bioengineering↗

bioRxiv2019

Global analysis of Two Component System (TCS) member of chickpea and other legume genomes implicates its role in enhanced nodulation

Nodule organogenesis is governed primarily by phytohormone cytokinin. We observed the significant nodulation in chickpea at particular cytokinin concentration (2.5x10-7) which indicated the importance of cytokinin in nodule development. Cytokinin signaling is mediated through the Two Component System (TCS) which comprises of sensor histidine kinases (HKs), histidine phosphotransfer proteins (HPs), and response regulators (RRs). Therefore, we analyzed the interconnection of cytokinin with TCS molecules during root nodule development through global analysis of TCS candidates in legumes with special consideration to cytokinin receptor and Type-B RR member. We have conducted an in depth global analysis of TCS family members in chickpea and other legumes, Medicago and pigeon pea. Higher number of TCS genes were found in Medicago (96), followed by pigeonpea (75) and chickpea (67). A good correlation between TCS members with their corresponding total number of genes were observed in all three-legume species. Collinearity analysis of TCS revealed phylogenetically closer proximity of Cicer to Medicago followed by Glycine than Cajanus. Comprehensive analysis of 3-dimensional structure, genomic organisation and domain arrangement showed a conservation of TCS members within species. In depth investigation showed that HKs were mainly conserved among TCS members in legumes and non-legumes while divergence occurred at level of RRs. Further, Type-B RRs were functionally most diversified in RRs based on phylogeny, syntenic and transcript analysis. Few numbers of segmentally duplicated pair of TCS showed difference in their transcriptional regulation suggesting the functional evolution. For functional characterization the cre1 mutants of (Medicago) were complemented with chickpea cytokinin responsive HKs and nodulation deficient phenotype of mutants were restored. A synchronous cytokinin-induced expression of chickpea cytokinin receptor HKs and CaNIN provides strong relation of cytokinin signaling during nodulation. Furthermore, interesting potential candidate CaRR13 was selected to deduce the underlying molecular mechanism of nodulation, chickpea in specific and legumes in general.

genomics↗

bioRxiv2019

Liver-Chip: Reproducing Human and Cross-Species Toxicities

Nonclinical rodent and non-rodent toxicity models used to support clinical trials of candidate drugs may produce discordant results or fail to predict complications in humans contributing to drug failures in the clinic. Here we applied microengineered Organ-on-Chip (Organ-Chip) technology to design rat, dog, and human Liver-Chips containing species-specific primary hepatocytes interfaced with liver sinusoidal endothelial cells, with or without Kupffer cells and hepatic stellate cells, cultured under physiological fluid flow. The Liver-Chips detected diverse phenotypes of liver toxicity including hepatocellular injury, steatosis, cholestasis, and fibrosis as well as species-specific toxicities when treated with tool compounds. Multi-species Liver-Chips may provide a useful platform for prediction of liver toxicity and inform human relevance of liver toxicities detected in animal studies to better determine safety and human risk.\n\nOne Sentence SummaryMicroengineered Organ-Chip technology has been used to design rat, dog and human Liver-Chips that recapitulate species-specific liver toxicities.

pharmacology and toxicology↗

bioRxiv2019

Mechanistic characterization of RASGRP1 variants identifies an hnRNP K-regulated transcriptional enhancer contributing to SLE susceptibility

Systemic lupus erythematosus (SLE) is an autoimmune disease with a strong genetic component. We recently identified a novel SLE susceptibility locus near RASGRP1, which governs the ERK/MAPK kinase cascade and B-/T-cell differentiation and development. However, precise causal RASGRP1 functional variant(s) and their mechanisms of action in SLE pathogenesis remain undefined. Our goal was to fine-map this locus, prioritize genetic variants according to likely functionality, experimentally validate the contribution of three SNPs to SLE risk, and experimentally determine their biochemical mechanisms of action. We performed a meta-analysis across six Asian and European cohorts (9,529 cases; 22,462 controls), followed by in silico bioinformatic and epigenetic analyses to prioritize potentially functional SNPs. We experimentally validated the functional significance and mechanism of action of three SNPs in cultured T-cells. Meta-analysis identified 18 genome-wide significant (p<5x10-8) SNPs, mostly concentrated in two haplotype blocks, one intronic and the other intergenic. Epigenetic fine-mapping, allelic, eQTL and imbalance analyses predicted three transcriptional regulatory regions with four SNPs (rs7170151, rs11631591-rs7173565, and rs9920715) prioritized for functional validation. Luciferase reporter assays indicated significant allele-specific enhancer activity for intronic rs7170151 and rs11631591-rs7173565 in T-lymphoid (Jurkat) cells, but not in HEK293 cells. Following up with EMSA, mass spectrometry and ChIP-qPCR, we detected allele-dependent interactions between heterogeneous nuclear ribonucleoprotein K (hnRNP-K) and rs11631591. Furthermore, inhibition of hnRNP-K in Jurkat and primary T-cells downregulated RASGRP1 and ERK/MAPK signaling. Comprehensive association, bioinformatics, and epigenetic analyses yielded putative functional variants of RASGRP1, which were experimentally validated. Notably, intronic variant (rs11631591) is located in a cell type-specific enhancer sequence, where its risk allele binds to the hnRNP-K protein and modulates RASGRP1 expression in Jurkat and primary T-cells. As risk allele dosage of rs11631591 correlates with increased RASGRP1 expression and ERK activity, we suggest that this SNP may underlie SLE risk at this locus.

genetics↗

bioRxiv2019

Development and validation of microsatellite markers from de novo transcriptome assembly of eggplant (Solanum melongena L.) and its putative progenitor S. incanum L. cultivars

An elite cultivar of eggplant, Ramnagar Giant (Solanum melongena L.) and W-4 (S. incanum L.) with contrasting horticultural traits were used as parental lines to develop a mapping population of RILs. To accelerate breeding programs and to develop large scale SSR markers to be used in QTL mapping, RNASeq libraries from different tissues of both the parental plants were deep sequenced and assembled into representation of a high quality de novo transcriptome using Illumina-based Next Generation Sequencing technology. 99.99% of high quality bases were obtained from all the tissues and deposited in TSA database at the NCBI link. Total 3, 156 and 3, 196 SNVs were detected in S. melongena and S. incanum, respectively. In S. melongena, 11, 262 SSR while in S. incanum 11, 829 SSR containing regions were identified. Based on functional annotation, 21, 914 unique genes could be identified for S. melongena, 21,706 unique genes for S. incanum and overall, 60 different transcription factors were identified in both the lines. Further, a total of 536 SSR markers were designed and screened for polymorphism of which, 157 markers produced polymorphism between the parental lines. The polymorphic SSRs shall be used for genotyping of RILs to map QTLs for various horticultural traits in eggplant and identification of candidate genes in response to biotic and abiotic stress.

genomics↗

bioRxiv2019

Transcriptional Blood Signatures for Active and Amphotericin B Treated Visceral Leishmaniasis in India

Amphotericin B provides improved therapy for visceral leishmaniasis (VL) caused by Leishmania donovani, with single dose liposomal-encapsulated Ambisome providing the best cure rates. The VL elimination program aims to reduce the incidence rate in the Indian subcontinent to <1/10,000 population/year. Ability to predict which asymptomatic individuals (e.g. anti-leishmanial IgG and/or Leishmania-specific modified Quantiferon positive) will progress to clinical VL would help in monitoring disease outbreaks. Here we examined whole blood transcriptional profiles associated with asymptomatic infection, active disease, and in treated cases. Two independent microarray experiments were performed, with analysis focussed primarily on differentially expressed genes (DEGs) concordant across both experiments. No DEGs were identified for IgG or Quantiferon positive asymptomatic groups compared to negative healthy endemic controls. We therefore concentrated on comparing concordant DEGs from active cases with all healthy controls, and in examining differences in the transcriptome following different regimens of drug treatment. In these comparisons 6 major themes emerged: (i) expression of genes and enrichment of gene sets associated with erythrocyte function in active cases; (ii) strong evidence for enrichment of gene sets involved in cell cycle in comparing active cases with healthy controls; (iii) identification of IFNG encoding interferon-{gamma} as the major hub gene in concordant gene expression patterns across experiments comparing active cases with healthy controls or with treated cases; (iv) enrichment for interleukin signalling (IL-1/3/4/6/7/8) and a prominent role for CXCL10/9/11 and chemokine signalling pathways in comparing active cases with treated cases; (v) the novel identification of Aryl Hydrocarbon Receptor signalling as a significant canonical pathway when comparing active cases with healthy controls or with treated cases; and (vi) global expression profiling support for more effective cure at day 30 post-treatment with a single dose of liposomal encapsulated amphotericin B compared to multi-dose non-liposomal amphotericin B treatment over 30 days. Author SummaryVisceral leishmaniasis (VL), also known as kala-azar, is a potentially fatal disease caused by intracellular parasites of the Leishmania donovani species complex. VL is a serious public health problem in rural India, causing high morbidity and mortality, as well as major costs to local and national health budgets. Amphotericin B provides improved therapy for VL with single dose liposomal-encapsulated Ambisome, now affordable through WHO-negotiated price reductions, providing the best cure rates. The VL elimination program aims to reduce the incidence rate in the Indian subcontinent to <1/10,000 population/year. By assessing immune responses to parasites in people infected with L. donovani, but with different clinical status, we can determine the requirements for immune cell development and predict which asymptomatic individuals, for example healthy individuals with high anti-leishmanial antibody levels, will progress to clinical VL. This will help in monitoring disease outbreaks. In this study we looked at global gene expression patterns in whole blood to try to understand more about asymptomatic infection, active VL, and the progress to cure in cases treated with single or multi-dose amphotericin B. The signatures of gene expression identified aid in our understanding of disease pathogenesis and provide novel targets for therapeutic intervention in the future.

genomics↗

bioRxiv2018

The ER chaperone PfGRP170 is essential for asexual development and is linked to stress response in malaria parasites.

The vast majority of malaria mortality is attributed to one parasite species: Plasmodium falciparum. Asexual replication of the parasite within the red blood cell is responsible for the pathology of the disease. In Plasmodium, the endoplasmic reticulum (ER) is a central hub for protein folding and trafficking as well as stress response pathways. In this study, we tested the role of an uncharacterized ER protein, PfGRP170, in regulating these key functions by generating conditional mutants. Our data show that PfGRP170 localizes to the ER and is essential for asexual growth, specifically required for proper development of schizonts. PfGRP170 is essential for surviving heat shock, suggesting a critical role in cellular stress response. The data demonstrate that PfGRP170 interacts with the Plasmodium orthologue of the ER chaperone, BiP. Finally, we found that loss of PfGRP170 function leads to the activation of the Plasmodium eIF2 kinase, PK4, suggesting a specific role for this protein in this parasite stress response pathway.

cell biology↗

bioRxiv2018

How molecular mechanisms of resistance affect resistance evolution

Combatting antibiotic resistance will require both new antibiotics and strategies to preserve the effectiveness of existing drugs. Both approaches would benefit from predicting optimal dosing of antibiotics based on drug-target binding parameters that can be measured early in drug development and that can change when bacteria become resistant. This would avoid the currently frequently employed trial-and-error approaches and might reduce the number of antibiotic candidates that fail late in drug development.\n\nHere, we describe a computational model (COMBAT-COmputational Model of Bacterial Antibiotic Target-binding) that leverages accessible biochemical parameters to quantitatively predict antibiotic dose-response relationships. We validate our model with MICs of a range of quinolone antibiotics in clinical isolates demonstrating that antibiotic efficacy can be predicted from drug-target binding (R2 > 0.9). To further challenge our approach, we do not only predict antibiotic efficacy from biochemical parameters, but also do the reverse: estimate the magnitude of changes in drug-target binding based on antibiotic dose-response curves. We experimentally demonstrate that changes in drug-target binding can be predicted from antibiotic dose-response curves with 92-94 % accuracy by exposing bacteria overexpressing target molecules to ciprofloxacin. To test the generality of COMBAT, we apply it to a different antibiotic class, the beta-lactam ampicillin, and can again predict binding parameters from dose-response curves with 90 % accuracy. We then apply COMBAT to predict antibiotic concentrations that can select for resistance due to novel resistance mutations.\n\nOur goal here is dual: First, we address a fundamental biological question and demonstrate that drug-target binding determines bacterial response to antibiotics, although antibiotic action involves many additional effects downstream of drug-target binding. Second, we create a tool that can help accelerate drug development by predicting optimal dosing and preserve the efficacy of existing antibiotics by predicting optimal treatment for possible resistant mutants.

biochemistry↗

bioRxiv2018

Analysis of genetic control and QTL mapping of essential wheat grain quality traits in a recombinant inbred population

Wheat cultivars are genetically crossed for improving end use quality for apt traits as per need of baking industry and broad consumers preferences. The processing and baking qualities of bread wheat underlie into genetic make-up of a variety and influence by environmental factors and their interactions. WL711 and C306 derived recombinant inbred lines (RILs) population of 206 was used for phenotyping of quality related traits in three different environmental conditions. The genetic analysis of quality traits showed considerable variation for measurable quality traits with normal distribution and transgressive segregation across the years. From the 206 RIL, few RILs found to be superior to those of the parental cultivars for key quality traitsindicating their potential usefor improvement of end use quality and also suggestingprobability of finding new alleles and allelic combinations from the RIL population. A genetic linkage map including 346 markers was constructed withtotal map distance of 4526.8cM andinterval distance between adjacent markersof 12.9cM. Mapping analysis identified 38 putative QTLs for 13 quality related traits with QTLs explaining 7.9% - 16.8% phenotypic variation spanning over 14 chromosomes i.e. 1A, 1B, 1D, 2A, 2D, 3B, 3D, 4A, 4B, 4D, 5D, 6A, 7A and 7B. Major novel QTLs regions for quality traits have been identified on several chromosome in studied RIL population posing their potential role in marker assisted selection for better bread making quality after validation.

genetics↗

bioRxiv2018

Cdc23/Mcm10 Primase Generates the Lagging Strand-Specific Ribonucleotide Imprint in Fission Yeast

The developmental asymmetry of fission yeast daughter cells derives from inheriting \"older Watson\" versus \"older Crick\" DNA strand from the parental cell, strands that are complementary but not identical with each other. A novel DNA strand-specific \"imprint\", installed during DNA replication at the mating-type locus (mat1), imparts competence for cell type inter-conversion to one of the two chromosome replicas. The biochemical nature of the imprint and the mechanism of its installation are still not understood. The catalytic subunit of DNA Polymerase (Pol) has been implicated in the imprinting process. Based on its known biochemical function, Pol might install the mat1 imprint during lagging strand synthesis. The nature of the imprint is not clear: it is either a nick or a ribonucleotide insertion. Our investigations do not support a role of Pol in nicking through putative endonuclease domains but confirm its role in installing an alkali-labile moiety as the imprint. A detailed genetic and molecular analysis reveals a direct role of the Cdc23/Mcm10 primase activity in installing the imprint in cooperation with Pol and Swi1.

genetics↗