Biology subjects

Tiwari, R.

Publications and source records attributed to Tiwari, R..

33 records · Page 2Linked to original sources

bioRxiv2024

Oxytocin facilitates social behavior of female rats via selective modulation of interneurons in the medial prefrontal cortex

The hypothalamic neuropeptide oxytocin is best known for its prosocial behavioral effects. However, the precise anatomical and cellular targets for oxytocin in the cortex during social behavior remain elusive. Here we show that oxytocin neurons project directly to the medial prefrontal cortex where evoked axonal oxytocin release facilitates social behaviors in adult female rats. In conjunction, we report that local oxytocin receptor-expressing (OTR+) cells are predominantly interneurons, whose activation promotes social interaction. Notably, this prosocial effect persists even under physiological challenge (hunger), pointing to a dedicated prosocial circuit capable of overriding primary survival drives. We further demonstrate that activation of these OTR+ interneurons inhibits principal cells specifically projecting to the basolateral amygdala, thus providing a putative mechanism of selective oxytocin action in this sociability-promoting cortical network.

neuroscience↗

bioRxiv2024

Multi-Model Machine Learning for Automated Identification of Rice Diseases Using Leaf Image Data

PurposeRice is grown almost everywhere in the world but is notably prevalent in Asian nations where it serves as the main food source for nearly half of the worlds population. Yet, enduring agricultural problems like various rice diseases have been a problem for farmers and planting specialists for ages. A fast, efficient, less expensive, and reliable approach to detecting rice diseases is urgently required in agricultural information since severe rice infections could result in no harvest of grains. Automated disease monitoring of rice plants using leaf images is critical for transitioning from labor-intensive, experience-based decision-making to an automated, data-driven strategy in agricultural production. In the modern era, Artificial Intelligence (AI) is being widely investigated in various areas of the medical and plant sciences to assess and diagnose the types of diseases. MethodsThis work proposes a hybrid deep-machine learning system for the automated detection of rice plant diseases using a leaf image dataset. Benchmarked MobileNetV2 architecture is employed to extract the deep features from the input images. Obtained features are fed as input to various machine learning classifiers with different kernel functions using a 10-fold validation strategy. ResultsThe developed hybrid system attained the highest classification accuracy of 98.6%, specificity of 98.85%, and sensitivity of 97.25% using a medium neural network. The results demonstrate that the established system is computationally faster and more efficient. The proposed system is ready for testing with more databases. ConclusionsThe suggested technology accurately diagnoses various rice plant illnesses, reducing manual labor and allowing farmers to receive prompt treatment. Future research topics include incorporating cloud-based monitoring for leaf image capture in non-connected farms, as well as building mobile IoT platforms for continuous screening.

plant biology↗

bioRxiv2023

Chronically disrupted sleep induces senescence in the visceral adipose tissue of C57BL/6J mice

The role of sleep in systemic aging remains poorly understood, despite sleeps essential function in preserving overall health and the prevalence of reduced sleep quality in modern society. Although reduced sleep correlates with an elevated risk of age-related diseases in humans, the mechanisms underlying this are unclear. In this study, we established a link between sleep and aging by demonstrating that disrupting sleep in C57BL/6 mice drives cellular senescence in the visceral adipose tissue. Sleep disruption also led to increased oxidative stress and DNA damage, both recognized triggers for senescence induction. Cellular senescence is implicated in numerous age-related conditions which are associated with insufficient sleep, such as cardiovascular disease, type 2 diabetes, and chronic inflammation. Our findings identify an accumulation of senescent cells in the adipose tissue, which serves as a potential target through which disturbed sleep accelerates the aging process and elevates the risk of age-related diseases.

molecular biology↗

bioRxiv2023

Post-ischemic inactivation of HIF prolyl hydroxylases in endothelium promotes maladaptive kidney repair by inducing glycolysis

Ischemic acute kidney injury (AKI) is common in hospitalized patients and increases the risk for chronic kidney disease (CKD). Impaired endothelial cell (EC) functions are thought to contribute in AKI to CKD transition, but the underlying mechanisms remain unclear. Here, we identify a critical role for endothelial oxygen sensing prolyl hydroxylase domain (PHD) enzymes 1-3 in regulating post-ischemic kidney repair. In renal endothelium, we observed compartment-specific differences in the expression of the three PHD isoforms in both mice and humans. We found that post-ischemic concurrent inactivation of endothelial PHD1, PHD2, and PHD3 but not PHD2 alone promoted maladaptive kidney repair characterized by exacerbated tissue injury, fibrosis, and inflammation. Single-cell RNA-seq analysis of the post-ischemic endothelial PHD1, PHD2 and PHD3 deficient (PHDTiEC) kidney revealed an endothelial glycolytic transcriptional signature, also observed in human kidneys with severe AKI. This metabolic program was coupled to upregulation of the SLC16A3 gene encoding the lactate exporter monocarboxylate transporter 4 (MCT4). Strikingly, treatment with the MCT4 inhibitor syrosingopine restored adaptive kidney repair in PHDTiEC mice. Mechanistically, MCT4 inhibition suppressed pro-inflammatory EC activation reducing monocyte-endothelial cell interaction. Our findings suggest avenues for halting AKI to CKD transition based on selectively targeting the endothelial hypoxia-driven glycolysis/MCT4 axis.

molecular biology↗

bioRxiv2023

Simultaneous recording of ultrasonic vocalizations and sniffing from socially interacting individual rats using a miniature microphone

Vocalizations are pivotal in mammalian communication, especially in humans. Rodents accordingly rely on ultrasonic vocalizations (USVs) that reflect their internal state as a primary channel during social interactions. However, attributing vocalizations to specific individuals remains challenging, impeding internal state assessment. Rats emit 50 kHz USVs to indicate positive states and intensify sniffing during alertness and social interactions. Here, we present a method involving a miniature microphone attached to the rat nasal cavity that allows to capture both male and female individual rat vocalizations and sniffing patterns during social interactions. We found that while the emission of 50 kHz USVs increases during close interactions, these signals lack specific behavioral associations. Moreover, a previously unreported low-frequency vocalization type marking rat social interactions was uncovered. Finally, different dynamics of sniffing and vocalization activities point to distinct underlying internal states. Thus, our method facilitates the exploration of internal states concurrent with social behaviors. MotivationHere, we sought to solve the problem of identifying the emitter of a specific vocalization during free social interaction of animal models. The use of a miniature microphone carried by the subject and connected to its nasal cavity solved this challenge and allowed for assignment of any vocalization recorded during a social interaction to a specific individual. Moreover, the miniature microphone enabled simultaneous recording of sniffing and vocalization activities of an individual, thus facilitating recognition of the internal states underlying these traits.

animal behavior and cognition↗

bioRxiv2023

CD38 regulates ovarian function and fecundity via NAD+ metabolism

Mammalian female reproductive lifespan is typically significantly shorter than life expectancy and is associated with a decrease in ovarian NAD+ levels. However, the mechanisms underlying this loss of ovarian NAD+ are unclear. Here, we show that CD38, a NAD+ consuming enzyme, is expressed in the ovarian extrafollicular space, primarily in immune cells, and its levels increase with reproductive age. Reproductively young mice lacking CD38 exhibit larger primordial follicle pools, elevated ovarian NAD+ levels, and increased fecundity relative to wild type controls. This larger ovarian reserve results from a prolonged window of follicle formation during early development. However, the beneficial effect of CD38 loss on reproductive function is not maintained at advanced age. Our results demonstrate a novel role of CD38 in regulating ovarian NAD+ metabolism and establishing the ovarian reserve, a critical process that dictates a females reproductive lifespan.

developmental biology↗

bioRxiv2023

Development of a novel epigenetic clock resistant to changes in immune cell composition

Epigenetic clocks are age predictors that use machine-learning models trained on DNA CpG methylation values to predict chronological or biological age. Increases in predicted epigenetic age relative to chronological age (epigenetic age acceleration) are connected to aging-associated pathologies, and changes in epigenetic age are linked to canonical aging hallmarks. However, epigenetic clocks rely on training data from bulk tissues whose cellular composition changes with age. We found that human naive CD8+ T cells, which decrease during aging, exhibit an epigenetic age 15-20 years younger than effector memory CD8+ T cells from the same individual. Importantly, homogenous naive T cells isolated from individuals of different ages show a progressive increase in epigenetic age, indicating that current epigenetic clocks measure two independent variables, aging and immune cell composition. To isolate the age-associated cell intrinsic changes, we created a new clock, the IntrinClock, that did not change among 10 immune cell types tested. IntrinClock showed a robust predicted epigenetic age increase in a model of replicative senescence in vitro and age reversal during OSKM-mediated reprogramming.

genomics↗

bioRxiv2022

Multi-level monitoring of EME1-MUS81 in CPT induced nucleolar rDNA repair

We identified EME1, the regulatory subunit of the structure-specific endonuclease complex EME1-MUS81, as substrate for the sumoylated UBC9 and demonstrated synergistic functions in promoting Camptothecin (CPT)-induced nucleolar ribosomal DNA (rDNA) repair (Nagamalleswari et al, co-submitted). Sumoylation of EME1 appears complex involving mono- and poly-sumoylation. Hence, we addressed here whether these modifications differentially regulate EME1 functions by analyzing EME1-variant expressing cell lines including mono-sumo(1)ylation and poly-sumo(2)ylation mimetic fusions. We complemented our analysis with the regulated endogenous EME1 interactome and our observation that Trichostatin A (TSA) induced EME1 and UBC9 sumoylation. Our findings are substantiated by identifying several regulatory proteins and by detecting CPT-induced endogenous di- and poly-sumoylated EME1 in different cell fractions. Together, our data suggest that Histone H4 acetylation, two mono-sumoylation events, poly-sumoylation, ISG15 and ubiquitination sequentially and in mutual dependence tightly control the intracellular localization, recruitment to DNA lesions, enzymatic activity, withdrawal from DNA lesions and the stability of EME1.

molecular biology↗

bioRxiv2022

Chemical inhibition of prolyl hydroxylases impairs angiogenic competence of human vascular endothelium through metabolic reprogramming

Endothelial cell (EC) metabolism has emerged as a driver of angiogenesis. While hypoxia inactivates prolyl-4 hydroxylase domain containing proteins 1-3 (PHD1-3) and stabilizes hypoxia inducible factors (HIFs) stimulating angiogenesis, the effects of PHDs on EC functions remain unclear. Here, we investigated the impact of PHD inhibition by dimethyloxalylglycine (DMOG) on angiogenic competence and metabolism of human vascular ECs. PHD inhibition reduced EC proliferation, migration, and tube formation capacities. Furthermore, transcriptomic and metabolomic analyses revealed an unfavorable metabolic reprogramming for angiogenesis following treatment with DMOG. Despite the induction of glycolytic genes and high levels of lactate, multiple genes encoding sub-units of mitochondrial complex I were suppressed with concurrent decline in nicotinamide adenine dinucleotide (NAD+) levels. Importantly, defective EC migration due to DMOG could be partially restored by augmenting NAD+ levels. Combined, our data provide metabolic insights into the mechanism by which chemical PHD inhibition impairs angiogenic competence of human vascular ECs.

molecular biology↗

bioRxiv2022

Human Elp3/Kat9 is a mitochondrial tRNA modifying enzyme

Post-translational modifications, such as lysine acetylation, regulate the activity of diverse proteins across many cellular compartments. Protein deacetylation in mitochondria is catalyzed by the enzymatic activity of the NAD+-dependent deacetylase sirtuin 3 (SIRT3), however it remains unclear whether corresponding mitochondrial acetyltransferases exist. We used a bioinformatics approach to search for mitochondrial proteins with an acetyltransferase catalytic domain, and identified a novel splice variant of ELP3 (mt-ELP3) of the elongator complex, which localizes to the mitochondrial matrix in mammalian cells. Unexpectedly, mt-ELP3 does not mediate mitochondrial protein acetylation but instead induces a post-transcriptional modification of mitochondrial-transfer RNAs (mt-tRNAs). Overexpression of mt-ELP3 leads to the protection of mt-tRNAs against the tRNA-specific RNase angiogenin, increases mitochondrial translation, and furthermore increases expression of OXPHOS complexes. This study thus identifies mt-ELP3 as a non-canonical mt-tRNA modifying enzyme.

molecular biology↗

bioRxiv2019

The Five-Primer Challenge: An inquiry-based laboratory module for synthetic biology

New technologies in DNA synthesis and assembly give genetic engineers complete freedom in genetic design, where virtually any plasmid DNA sequence can be created efficiently and economically. Learning how to design, construct, and test new DNA sequences is a critical skill for researchers in molecular biology and biotechnology. Here we present a student-centered, inquiry-based module in which students learn how to control bacterial gene expression by appplying various DNA assembly techniques. The central activity in this learning module is termed the Five-Primer Challenge. Each student is allowed to order up to five 60-mer oligonucleotide primers to then modify a GFP expression plasmid with the goal of increasing GFP expression as much as possible. This module was developed and implemented at the 2016 Cold Spring Harbor Laboratory Synthetic Biology Course, and was effective at engaging students in critical thinking and in promoting student learning.

scientific communication and education↗

bioRxiv2019

Machine learnt image processing to predict weight and size of rice kernels

Accurate measurement of rice kernel sizes after milling is critical to design, develop and optimize rice milling operations. The size and mass of the individual rice kernels are important parameters typically associated with rice quality attributes, particularly head rice yield. In this study, we propose a novel methodology that combines image processing and machine learning (ML) ensemble to accurately measure the size and mass of several rice kernels simultaneously. We have developed an image processing algorithm with the help of recursive method to identify the individual rice kernels from an image and estimate the size of the kernels based on the pixels a kernel occupies. The number of pixels representing a rice kernel has been used as its digital fingerprint in order to predict its size and mass. We have employed a number of popular machine learning models to build a stacked ensemble model (SEM), which can predict the mass of the individual rice kernels based on the features derived from the pixels of the individual kernels in the image. The prediction accuracy and robustness of our image processing and SEM are quantified using uncertainty quantification. The uncertainty quantification showed 3.6%, 2.5%, and 2.4% for mean errors in estimating the kernel length of small-grain (Calhikari-202), medium-grain (Jupiter), and long-grain (CL153) rice, respectively. Similarly, mean errors associated with predicting the 1000 grain weight are 4.1%, 2.9%, and 4.3% for Calhikari-202, Jupiter, and CL153, respectively. Use of the developed algorithm in rice imaging analyzers could facilitate head rice yield quantifications and promote quicker rice quality appraisals.

bioengineering↗

bioRxiv2019

Size and mass prediction of almond kernels using machine learning image processing

After harvesting almond crop, accurate measurement of almond kernel sizes is a significant specification to plan, develop and enhance almond processing operations. The size and mass of the individual almond kernels are vital parameters usually associated with almond quality, particularly head almond yield. In this study, we propose a novel methodology that combines image processing and machine-learning ensemble that accurately measures the size and mass of whole raw almond kernels (classification - Nonpareil) simultaneously. We have developed an image-processing algorithm using recursive method to identify the individual almond kernels from an image and estimate the size of the kernels based on the occupied pixels by a kernel. The number of pixels representing an almond kernel was used as its digital fingerprint to predict its size and mass. Various popular machine learning (ML) models were implemented to build a stacked ensemble model (SEM), predicting the mass of the individual almond kernels based on the features derived from the pixels of the individual kernels in the image. The prediction accuracy and robustness of image processing and SEM were analyzed using uncertainty quantification. The mean error in estimating the average length of 1000 almond kernel was 3.12%. Similarly, mean errors associated with predicting the 1000 kernel mass were 0.63%. The developed algorithm in almond imaging in this study can be used to facilitate a rapid almond yield and quality appraisals.

bioengineering↗

bioRxiv2019

Androgen deprivation upregulates SPINK1 expression and potentiates cellular plasticity in prostate cancer

The Serine Peptidase Inhibitor, Kazal type 1 (SPINK1) overexpression represents ~10-25% of the prostate cancer (PCa) cases associated with shorter recurrence-free survival and poor prognosis. Nonetheless, androgen-deprivation therapy (ADT) remains the mainstay treatment for locally advanced and metastatic PCa patients. However, majority of these individuals eventually progress to castration-resistant stage, and a subset of these patients develop ADT-induced neuroendocrine PCa. Despite adverse effects of ADT, possible role of androgen signaling in SPINK1-mediated prostate oncogenesis remains unexplored. Here, we show that androgen receptor (AR) and its corepressor, the RE1-silencing transcription factor (REST), occupy SPINK1 promoter and functions as a direct transcriptional repressor of SPINK1, thus blocking AR signaling via ADT relieves its repression, leading to SPINK1 upregulation. In agreement, an inverse association between SPINK1 levels and AR expression was observed across multiple PCa cohorts, and in neuroendocrine differentiated cells. While, lineage reprogramming factor SOX2 in turn binds to SPINK1 promoter leading to its transactivation in androgen-deprived conditions with concomitant increase in neuroendocrine markers. Additionally, we also confirm the role of SPINK1 in epithelial-mesenchymal transition, drug resistance, stemness and cellular plasticity. Moreover, we show that Casein Kinase 1 inhibitor stabilizes the REST levels, which in cooperation with AR, conjures transcriptional repression of SPINK1 expression, and impedes SPINK1-mediated oncogenesis. Collectively, our findings provide a plausible explanation to the paradoxical clinical outcomes of ADT, possibly due to increased SPINK1 levels. This study highlights the need to take a well-informed decision prior to ADT and develop alternative therapeutic strategies for castrate-resistant PCa patients.

cancer biology↗

bioRxiv2018

Polycomb Group protein EZH2-mediated transcriptional repression of microRNA-338/-421 drives SPINK1-positive prostate cancer

PurposeSerine Peptidase Inhibitor, Kazal type-1 (SPINK1) overexpression defines the second most recurrent and aggressive prostate cancer (PCa) subtype. However, the underlying molecular mechanism and pathobiology of SPINK1 in PCa remains largely unknown.\n\nExperimental DesignMicroRNA-prediction tools were employed to examine the SPINK1-3UTR for miRNAs binding. Luciferase reporter assays were performed to confirm the SPINK1-3UTR binding of shortlisted miR-338-5p/miR-421. Further, miR-338-5p/-421 overexpressing cancer cells (SPINK1-positive) were evaluated for oncogenic properties using cell-based functional assays and mice xenograft model. Global gene expression profiling was performed to unravel the biological pathways altered by miR-338-5p/-421. Immunohistochemistry and RNA in-situ hybridization was carried-out on PCa patients tissue microarray for SPINK1 and EZH2 expression respectively. Chromatin immunoprecipitation assay was performed to examine EZH2 occupancy on the miR-338-5p/-421 regulatory regions. Bisulfite sequencing and methylated DNA-immunoprecipitation was performed on PCa cell lines and patients specimens.\n\nResultsWe established a critical role of miRNA-338-5p/-421 in post-transcriptional regulation of SPINK1. Ectopic expression of miRNA-338-5p/-421 in SPINK1-positive PCa cells abrogate oncogenic properties including cell-cycle progression, stemness and drug resistance, and show reduced tumor burden and distant metastases in mice model. Importantly, we show SPINK1-positive PCa patients exhibit increased EZH2 expression, suggesting its role in miRNA-338-5p/-421 epigenetic silencing. Furthermore, presence of CpG dinucleotide DNA methylation marks on the regulatory regions of miR-338-5p/-421 in SPINK1-positive PCa cells and patients specimens confirms epigenetic silencing.\n\nConclusionOur findings revealed that miRNA-338-5p/-421 are epigenetically silenced in SPINK1-positive PCa, while restoring the expression of these miRNAs using epigenetic drugs or synthetic mimics could abrogate SPINK1-mediated oncogenesis.\n\nTRANSLATIONAL IMPACTWe establish a regulatory model involving the functional interplay between SPINK1, miRNA-338-5p/miRNA-421 and EZH2, thereby, revealing hitherto unknown mechanism of SPINK1 up-regulation in SPINK1-positive subtype. Our findings provide a strong rationale for the development of potential therapeutic strategies for SPINK1-positive malignancies. We demonstrate that restoring miRNA-338-5p/miRNA-421 expression using epigenetic drugs including DNMTs inhibitors in combination with HDACs or HKMTs inhibitors or miRNA synthetic mimics in SPINK1-positive prostate cancer abrogate SPINK1-mediated oncogenicity. The major findings of this study will not only advance the prostate cancer field, but will also be valuable for treatment and disease management of other SPINK1-positive malignancies.

cancer biology↗