Biology subjects

Rao, P.

Publications and source records attributed to Rao, P..

11 recordsLinked to original sources

bioRxiv2022

A novel transcriptional signature identifies T-cell infiltration in high-risk paediatric cancer

Molecular profiling of the tumour immune microenvironment (TIME) has enabled the rational choice of immunotherapies in some adult cancers. In contrast, the TIME of paediatric cancers is relatively unexplored. We speculated that a more refined appreciation of the TIME in childhood cancers, rather than a reliance on commonly used biomarkers such as tumour mutation burden (TMB), neoantigen load and PD-L1 expression, is an essential prerequisite for improved immunotherapies in childhood solid cancers. We combined immunohistochemistry (IHC) and molecular profiling to develop an alternative, expression-based signature associated with CD8+ T-cell infiltration of the TIME in high-risk paediatric tumours. Using this novel 15-gene immune signature, Immune Paediatric Signature Score (IPASS), we estimate up to 31% of high-risk cancers harbour infiltrating T-cells. Our data provides new insights into the variable immune-suppressive mechanisms dampening responses in paediatric solid cancers. Effective immune-based interventions in high-risk paediatric cancer will require individualised analysis of the TIME.

cancer biology↗

bioRxiv2022

Purification and biochemical analysis of native AMPA receptors from three different mammalian species

The majority of fast, excitatory synaptic transmission in the central nervous system (CNS) is mediated by -amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs), which are glutamate-activated ion channels integral to synaptic plasticity, motor coordination, learning, and memory. AMPARs are multiprotein assemblies comprised of a tetrameric receptor core that co-assembles with a broad range of peripheral auxiliary proteins which shape subcellular localization and signaling properties of the resulting complexes. Structure determination of AMPARs has traditionally relied on recombinant expression systems; however, these methods are not well suited to elucidate the diverse array of AMPAR assemblies that are differentially expressed in mammalian brains. While recent studies of native receptor complexes have advanced our understanding of endogenous assemblies, receptors thus far have only been isolated from rodent brain tissue. Here, we employed an immunoaffinity purification strategy to isolate native AMPARs from the brains of three different mammals - pigs, sheep, and cows. Compared to rodents; pigs, sheep, and cows are from a unique clade of mammals - ungulates, that share a closer genomic identity with humans. We determined the molecular size, overall yield, and purity of native AMPARs isolated from these three mammals, thereby demonstrating that structural determination and biochemical analysis is possible from a clade of mammals evolutionarily distinct from rodents.

biochemistry↗

bioRxiv2022

Sucrose metabolism and candidate genes during catkin fibers development in poplar

Poplar is an important tree species for ecological protection, wood production, bioenergy and urban greening; it has been widely planted worldwide. However, the catkin fibers produced by female poplars can cause environmental pollution and safety hazards during spring. This study focused on Populus tomentosa, and revealed the sucrose metabolism regulatory mechanism of catkin fibers development from morphological, physiological and molecular aspects. Paraffin section suggested that poplar catkin fibers were not seed hairs and produced from the epidermal cells of funicle and placenta. Sucrose degradation via invertase and sucrose synthase played the dominant role during poplar catkin fibers development. The expression patterns revealed that sucrose metabolism-related genes played important roles during catkin fibers development. Y1H analysis indicated that there was a potential interaction between sucrose synthase 2 (PtoSUS2)/vacuolar invertase 3 (PtoVIN3) and MYB/ bHLH transcription factors in poplar. Finally, the two key genes, PtoSUS2 and PtoVIN3, had roles in Arabidopsis trichome density, indicating that sucrose metabolism is important in poplar catkin fibers development. This study is not only helpful for clarifying the mechanism of sucrose regulation during trichome development in perennial woody plants, but also establishes a foundation to solve poplar catkin fibers pollution through genetic engineering methods. HighlightSucrose degradation via invertase and sucrose synthase plays an important role in poplar catkin fibers development, and PtoSUS2 and PtoVIN3 are potential promising targets to solve poplar catkin fibers pollution.

molecular biology↗

bioRxiv2022

Fish oil Nano-emulsion Kills Macrophage: Ferroptosis and Autophagy Triggered by Catalase-catalysed Superoxide Eruption

Fish oil is increasingly utilized in the form of nano-emulsion as nutrient and function fortifier. The nano-emulsions high polyunsaturated fatty acids content and electron donors at oil/water interface could be a potential site of redox reaction, if a previously unrecognised trigger was provided. Here we report that a vigorous superoxide production occurred in fish oil nano-emulsion in the presence of mammalian catalase in both acellular and cellular systems. Furthermore, the resulting superoxide increased cytosolic ROS and membrane lipid peroxidation of murine macrophage, and eventually caused fatal oxidative damage, which involves autophagy and ferroptosis but not apoptosis. The cell death was significantly inhibited by a catalase-specific inhibitor. The ferroptosis was independent of protease Caspase-3 activation or glutathione peroxidase suppression. Our findings discover a hidden risk factor of the widely acclaimed fish oil emulsion and suggest a novel mechanism of cellular damage by dietary lipids on mucosal layer of the alimentary tract.

cell biology↗

bioRxiv2022

RelCoVax(R), a two antigen subunit protein vaccine candidate against SARS-CoV-2 induces strong immune responses in mice

The COVID-19 pandemic has spurred an unprecedented movement to develop safe and effective vaccines against the SARS-CoV-2 virus to immunize the global population. The first set of vaccine candidates that received emergency use authorization targeted the spike (S) glycoprotein of the SARS-CoV-2 virus that enables virus entry into cells via the receptor binding domain (RBD). Recently, multiple variants of SARS-CoV-2 have emerged with mutations in S protein and the ability to evade neutralizing antibodies in vaccinated individuals. We have developed a dual RBD and nucleocapsid (N) subunit protein vaccine candidate named RelCoVax(R) through heterologous expression in mammalian cells (RBD) and E. coli (N). The RelCoVax(R) formulation containing a combination of aluminum hydroxide (alum) and a synthetic CpG oligonucleotide as adjuvants elicited high antibody titers against RBD and N proteins in mice after a prime and boost dose regimen administered 2 weeks apart. The vaccine also stimulated cellular immune responses with a potential Th1 bias as evidenced by increased IFN-{gamma} release by splenocytes from immunized mice upon antigen exposure particularly N protein. Finally, the serum of mice immunized with RelCoVax(R) demonstrated the ability to neutralize two different SARS-CoV-2 viral strains in vitro including the Delta strain that has become dominant in many regions of the world and can evade vaccine induced neutralizing antibodies. These results warrant further evaluation of RelCoVax(R) through advanced studies and contribute towards enhancing our understanding of multicomponent subunit vaccine candidates against SARS-CoV-2.

immunology↗

bioRxiv2021

Delta variant with P681R critical mutation revealed by ultra-large atomic-scale ab initio simulation: Implications for the fundamentals of biomolecular interactions

SARS-CoV-2 Delta variant is emerging as a globally dominant strain. Its rapid spread and high infection rate are attributed to a mutation in the spike protein of SARS-CoV-2 allowing the virus to invade human cells much faster and with increased efficiency. Particularly, an especially dangerous mutation P681R close to the furin cleavage site has been identified as responsible for increasing the infection rate. Together with the earlier reported mutation D614G in the same domain, it offers an excellent instance to investigate the nature of mutations and how they affect the interatomic interactions in the spike protein. Here, using ultra large-scale ab initio computational modeling, we study the P681R and D614G mutations in the SD2-FP domain including the effect of double mutation and compare the results with the wild type. We have recently developed a method of calculating the amino acid-amino acid bond pairs (AABP) to quantitatively characterize the details of the interatomic interactions, enabling us to explain the nature of mutation at the atomic resolution. Our most significant find is that the mutations reduce the AABP value, implying a reduced bonding cohesion between interacting residues and increasing the flexibility of these amino acids to cause the damage. The possibility of using this unique mutation quantifiers in a machine learning protocol could lead to the prediction of emerging mutations.

biochemistry↗

bioRxiv2021

Polygenic enrichment distinguishes disease associations of individual cells in single-cell RNA-seq data

Gene expression at the individual cell-level resolution, as quantified by single-cell RNA-sequencing (scRNA-seq), can provide unique insights into the pathology and cellular origin of diseases and complex traits. Here, we introduce single-cell Disease Relevance Score (scDRS), an approach that links scRNA-seq with polygenic risk of disease at individual cell resolution without the need for annotation of individual cells to cell types; scDRS identifies individual cells that show excess expression levels for genes in a disease-specific gene set constructed from GWAS data. We determined via simulations that scDRS is well-calibrated and powerful in identifying individual cells associated to disease. We applied scDRS to GWAS data from 74 diseases and complex traits (average N =346K) in conjunction with 16 scRNA-seq data sets spanning 1.3 million cells from 31 tissues and organs. At the cell type level, scDRS broadly recapitulated known links between classical cell types and disease, and also produced novel biologically plausible findings. At the individual cell level, scDRS identified subpopulations of disease-associated cells that are not captured by existing cell type labels, including subpopulations of CD4+ T cells associated with inflammatory bowel disease, partially characterized by their effector-like states; subpopulations of hippocampal CA1 pyramidal neurons associated with schizophrenia, partially characterized by their spatial location at the proximal part of the hippocampal CA1 region; and subpopulations of hepatocytes associated with triglyceride levels, partially characterized by their higher ploidy levels. At the gene level, we determined that genes whose expression across individual cells was correlated with the scDRS score (thus reflecting co-expression with GWAS disease genes) were strongly enriched for gold-standard drug target and Mendelian disease genes.

bioinformatics↗

bioRxiv2021

Integrated Omics Analysis Reveals Sirtuin Signaling is Central to Hepatic Response to a High Fructose Diet

BackgroundDietary high fructose (HFr) is a known metabolic disruptor contributing to development of obesity and diabetes in Western societies. Initial molecular changes from exposure to HFr on liver metabolism may be essential to understand the perturbations leading to insulin resistance and abnormalities in lipid and carbohydrate metabolism. We studied vervet monkeys (Clorocebus aethiops sabaeus) fed a HFr (n=5) or chow diet (n=5) for 6 weeks, and obtained clinical measures of liver function, blood insulin, cholesterol and triglycerides. In addition, we performed untargeted global transcriptomics, proteomics, and metabolomics analyses on liver biopsies to determine the molecular impact of a HFr diet on coordinated pathways and networks that differed by diet. ResultsWe show that integration of omics data sets improved statistical significance for some pathways and networks, and decreased significance for others, suggesting that multiple omics datasets enhance confidence in relevant pathway and network identification. Specifically, we found that sirtuin signaling and a peroxisome proliferator activated receptor alpha (PPARA) regulatory network were significantly altered in hepatic response to HFr. Integration of metabolomics and miRNAs data further strengthened our findings. ConclusionsOur integrated analysis of three types of omics data with pathway and regulatory network analysis demonstrates the usefulness of this approach for discovery of molecular networks central to a biological response. In addition, metabolites aspartic acid and docosahexaenoic acid (DHA), protein ATG3, and genes ATG7, HMGCS2 link sirtuin signaling and the PPARA network suggesting molecular mechanisms for altered hepatic gluconeogenesis from consumption of a HFr diet.

systems biology↗

bioRxiv2020

Evaluation of Safety and Immunogenicity of an Adjuvanted, TH-1 Skewed, Whole Virion Inactivated SARS-CoV-2 Vaccine - BBV152

We report the development and evaluation of safety and immunogenicity of a whole virion inactivated SARS-COV-2 vaccine (BBV152), adjuvanted with aluminium hydroxide gel (Algel), or a novel TLR7/8 agonist adsorbed Algel. We used a well-characterized SARS-CoV-2 strain and an established vero cell platform to produce large-scale GMP grade highly purified inactivated antigen, BBV152. Product development and manufacturing were carried out in a BSL-3 facility. Immunogenicity was determined at two antigen concentrations (3g and 6g), with two different adjuvants, in mice, rats, and rabbits. Our results show that BBV152 vaccine formulations generated significantly high antigen-binding and neutralizing antibody titers, at both concentrations, in all three species with excellent safety profiles. The inactivated vaccine formulation containing TLR7/8 agonist adjuvant-induced Th1 biased antibody responses with elevated IgG2a/IgG1 ratio and increased levels of SARS-CoV-2 specific IFN-{gamma}+ CD4 T lymphocyte response. Our results support further development for Phase I/II clinical trials in humans.

immunology↗

bioRxiv2020

Hybrid origin of Populus tomentosa Carr. identified through genome sequencing and phylogenomic analysis

Populus tomentosa is widely distributed and cultivated in the Northern and Central China, where it is of great economic and ecological importance. However, the origin of P. tomentosa remains controversial. Here, we used a PacBio+Hi-C+Illumina strategy to sequence and assemble its 740.2 Mb (2n) genome. The assembly accounts for greater than 92.1% of the 800-megabase genome, comprises 38 chromosomes, and contains 59,124 annotated protein-coding genes. Phylogenomic analyses elucidated dynamic genome evolution events among its closely related white poplars, and revealed that tomentosa is comprised of two subgenomes, which we deomonstrate is likely to have resulted from hybridization between Populus adenopoda as the female, and Populus alba var. pyramidalis as the male, around 3.93 Mya. We also detected structural variations and allele-indels across genome. Our study presents a high quality and well assembled genome, unveils the origin of the widely distributed and planted P. tomentosa, and provides a powerful resource for comparative plant biology, breeding, and biotechnology.

genomics↗

bioRxiv2019

JUUL and Combusted Cigarettes Comparably Impair Endothelial Function

ObjectivesJUUL and earlier generation electronic cigarettes (e-cigs) are promoted as being less hazardous than cigarettes. While JUUL Labs, in particular, claims that switching from smoking to vaping has beneficial impacts, the health effects of such products are not well understood. We investigated whether exposure to JUUL and previous generation e-cig aerosol impairs endothelial function comparably to cigarette smoke.\n\nMethodsWe exposed rats to aerosol from Virginia Tobacco flavor JUUL, an e-cig tank system using unflavored freebase nicotine e-liquid, Marlboro Red combustible tobacco cigarettes, or clean air for 10 cycles of 2 second inhalation over 5 minutes. Endothelial function (FMD) was measured pre- and post-exposure. Blood was collected 20 mins post-exposure for serum nicotine analysis.\n\nResultsAerosol/smoke from JUUL, previous generation e-cigs, and cigarettes all impaired FMD. The extent of impairment ranged from 34%-58%, although the differences between groups were insignificant. Nicotine was highest in serum from the JUUL group; for the other e-cig and cigarette groups, nicotine levels were lower and comparable to each other.\n\nConclusionsAerosol from JUUL and previous generation e-cigs impairs endothelial function in rats, comparable to impairment by cigarette smoke.

pharmacology and toxicology↗