Biology subjects

Balaji, A.

Publications and source records attributed to Balaji, A..

14 recordsLinked to original sources

bioRxiv2026

Food acquisition patterns of the three-striped palm squirrel are influenced by interactions: variation in foraging response across an urban-rural gradient

Urbanisation significantly alters the habitat and resources accessible to urban animals. Synanthropic animals adapt by exploiting available resources and modulating abundance across the urban habitat matrix. Urban arboreal mammals, such as palm squirrels, encountering altered foraging resources in green spaces and across the matrix, develop strategies to maximise their fitness, including modulating food selection, spatial microhabitat use, and social interactions. We examined whether squirrel abundance affects foraging patterns and whether social interactions at the feeding site regulated individuals' exploratory tendencies. We conducted cafeteria experiments with F. palmarum in rural and urban areas around Tirupati, India and tested for differences in responses to novelty, food selection, and dominance interactions among individuals. We observed a preferential bias towards processed food and approach tendencies were stronger in urban individuals. Urban regions marked with a higher density of squirrels, were associated with higher competitive interactions showing transient social despots of dominance, leading to increased exploration by non-dominant individuals. On the contrary, a lower density of squirrels in rural areas, leads to reduced dominance interactions and subsequent lack of exploration. The potential exploration and consumption of novel resources feeds into the urban loop which enhances population in urban areas, whereas the opposite effect is observed in rural areas. This study highlights how urban commensal organisms, such as squirrels, adopt behaviours to maximise resource utilisation which is modulated by social interactions and can potentially determine the population dynamics in changing landscapes.

ecology↗

bioRxiv2026

Organization of Myosin H in the Apical Complex of Toxoplasma Gondii Revealed by 3D Single-Molecule Super-Resolution Microscopy

Toxoplasma gondii is a single-celled eukaryotic parasite with prolific invasion capability. The parasite uses an apical complex comprised of proteinaceous structures and secretory organelles to efficiently enter host cells. As a result, the apical complex remains a vital structure of interest, with many studies dedicated to understanding its protein organization. One such protein is the motor Myosin H (MyoH), which is indispensable for parasite motility and host cell invasion. Given the small size of the complex, roughly a diffraction-limited volume in the visible, high-resolution techniques are required to make precise determinations of protein organization. In this work, we use 3D single-molecule localization microscopy in both traditionally fixed and gel-expanded parasites to localize the indispensable motor Myosin H within the apical complex. Labeling of the N- and C-terminus of MyoH in fixed parasites resolved the orientation of the motor protein in the apical complex, showing the motor head radially exterior to the tail. Two-color imaging of MyoH with tubulin in fixed parasites allowed for localization of the MyoH termini relative to the conoid, a barrel of tubulin-based fibers in the apical complex and showed the MyoH tail toward the interior face of the conoid and the head at the conoid exterior. Gel expansion showed improved labeling density for both tubulin and MyoH but altered MyoH localization, highlighting the nuanced effects of gel expansion on protein organization. Statement of SignificanceThis work employs 3D single-molecule super-resolution microscopy to provide quantitative physical analysis of the spatial organization of a vital myosin motor, MyoH, in the model apicomplexan parasite Toxoplasma gondii. While previous studies have provided high-resolution views of the parasites invasion machinery, MyoH has remained elusive at the nanoscale. We resolved differences in radial organization between the N- and C-termini of the motor, thus determining the orientation of the protein in the apical space. Two-color imaging revealed the organization of the motor in the greater context of the parasites invasion complex. 3D single-molecule imaging in gel-expanded samples revealed an increase in labeling efficiency but perturbed localization of only the MyoH C-terminus, highlighting the nuanced effects of gel expansion on protein organization.

biophysics↗

bioRxiv2026

The role of two GLYCOALKALOID METABOLISM genes in α-tomatine biosynthesis and basal defense in tomato

Steroidal glycoalkaloids and saponins are plant cholesterol-based steroid metabolites with antimicrobial activities and potential pharmacological value. The saponin uttroside B from black nightshade (Solanum nigrum) plays an important role in defense against herbivorous insect and exhibits anti-hepatocellular carcinoma activity. The tomato (S. lycopersicum) glycoalkaloid -tomatine has been studied because of its antinutritional effects, however, its role in protecting plants from fungal pathogens remains understudied. The biosynthetic pathway of -tomatine involves multiple clustered genes designated as glycoalkaloid metabolism (GAME) genes. In this study, we generated single knockout mutants of SlGAME4 and SlGAME2 by CRISPR/Cas9-based genome editing. The SlGAME4 mutants did not accumulate glycoalkaloids but instead redirected resources towards steroidal saponin (uttroside B) synthesis. SlGAME2 mutants contained unaltered -tomatine contents indicating that the SlGAME2 gene, previously reported to catalyze the transfer of xylose to {beta}1-tomatine, is not involved in -tomatine biosynthesis. Infection assays with four fungal tomato pathogens demonstrated that the SlGAME4 mutant plants were slightly more susceptible to Botrytis cinerea, but equally susceptible to the other three fungi. Up-regulation of -tomatine-responsive genes in B. cinerea was observed during infection on SlGAME4 mutant tomato, as well as on S. nigrum suggesting that uttroside B induces a fungal transcriptional response similar to -tomatine. Furthermore, we observed that tolerance mechanisms to plant saponins mediated by glycosyl hydrolases and a glycosyltransferase contribute to virulence of B. cinerea on SlGAME4 mutant plants and S. nigrum. This indicates that also uttroside B contributes to defense against fungal pathogens and can be detoxified by B. cinerea.

plant biology↗

bioRxiv2025

Efficient Double Helix Detection with Steerable Filters

We present an efficient detection scheme for localization of Double Helix point-spread functions for 3D single-molecule localization microscopy or tracking. Using steerable filters, we extract both 2D position and lobe orientation (axial position) estimates using just 7 convolutions, orders of magnitude less than used in deep-learning-based approaches. We pair this detection with a fitter and implement both as a plug-in for the open source PYthon Microscopy Environment (PYME), which features percentile-based background subtraction, signal-to-noise-based detection thresholding, and performant parallel analysis. Our complete localization analysis pipeline achieves state-of-the-art performance with minimal user input.

biophysics↗

bioRxiv2025

A Super-Resolution Spatial Atlas of SARS-CoV-2 Infection in Human Cells

Coronaviral replication depends on double-membrane vesicles (DMVs), yet where polyprotein processing occurs and how replication organelles mature remain unresolved. We built a multi{-}color super-resolution atlas of SARS-CoV-2 RNA, non-structural and structural proteins in infected human cells, combining 3D single-molecule localization with radial and angular pair-correlation analysis. The atlas uncovers fundamental spatial principles. First, nsp5 (3CLpro) localizes within the DMV lumen near pores, with nsp7-nsp16 positioned interior to nsp4, supporting a protease-dependent maturation model in which nsp5 action permits membrane closure and subsequently completes intravesicular polyprotein processing to activate replication complexes. Second, we identify dsRNA connectors bridging DMVs and non-canonical, nsp3/4-lacking dsRNA granules decorated with replicase components, consistent with a condensate-mediated route for trafficking replication intermediates. Third, mapping structural proteins reveals M- and S-positive virion assembly intermediates along the secretory route, while premature intracellular S1 shedding captures the baseline instability of the original virus isolate prior to evolutionary adaptation. Finally, the antiviral nirmatrelvir induces multilayered bodies of uncleaved polyproteins (nsp4-5-10-16) that persist after washout and precede rapid rebound, suggesting a drug-induced reservoir state. Beyond mapping the viral architecture, this atlas resolves the spatial context of proteolysis and organelle remodeling, providing a framework for pan-coronaviral mechanisms and antiviral design.

microbiology↗

bioRxiv2025

Mice lacking the endocannabinoid-synthesizing enzyme NAPE-PLD exhibit sex-dependent dysregulations in responsiveness to oxycodone and a natural reward

The endogenous opioid and endogenous cannabinoid (endocannabinoid) systems are highly interconnected in the context of drug reward. Bioactive lipids known as N-acylethanolamines (NAEs), and, specifically, anandamide (AEA), influence several unwanted side effects of opioids, including dependence and tolerance. AEA undergoes degradation by the enzyme fatty-acid amide hydrolase (FAAH), whereas the biosynthesis of AEA in vivo is catalyzed by the enzyme N-acyl phosphatidylethanolamine phospholipase-D (NAPE-PLD). AEA and FAAH are implicated in opioid reward, but the impact of genetic deletion of NAPE-PLD on responsiveness to opioids remains unknown. Here we explored the role of NAPE-PLD in behavioral sensitivity to the opioid analgesic oxycodone. We evaluated NAPE-PLD knockout (KO) and wild type (WT) mice of both sexes in preclinical assays that assess either opioid-induced psychomotor responses or voluntary oral consumption of oxycodone. In our studies, genetic deletion of NAPE-PLD produced a shift in sexually dimorphic responses to oxycodone. Psychomotor response to oxycodone was reduced in female NAPE-PLD KO mice but not in males. Female NAPE-PLD KO mice consumed more oral oxycodone that female WT mice, while no genotypic differences in consumption were observed in males. Oxycodone consumption also increased the number of striatal {Delta}FosB positive cells in female WT mice, but not in male WT mice or NAPE-PLD KO mice of either sex. Additionally, NAPE-PLD KO mice of both sexes consumed more sucrose than WT mice. Together, these findings suggest that NAPE-PLD may regulate responses to opioids in a sexually dimorphic manner as the impact of genetic deletion of NAPE-PLD was greater in females than males.

neuroscience↗

bioRxiv2024

Pre-fusion AAA+ remodeling of target-SNARE protein complexes enables synaptic transmission

Synaptic vesicles containing neurotransmitters fuse with the plasma membrane upon the arrival of an action potential at the active zone. Multiple proteins organize trans-SNARE complex assembly and priming, leading to fusion. One target membrane SNARE, syntaxin, forms nanodomains at the active zone, and another, SNAP-25, enters non-fusogenic complexes with it. Here, we reveal mechanistic details of AAA+ protein NSF (N-ethylmaleimide sensitive factor) and SNAP (soluble NSF attachment protein) action before fusion. We show that syntaxin clusters are conserved, that NSF colocalizes with them, and characterize SNARE populations that may exist within or near them using cryo-EM. Supercomplexes of NSF, -SNAP, and either a syntaxin tetramer or one of two binary complexes of syntaxin--SNAP-25 reveal atomic details of SNARE processing and show how sequential ATP hydrolysis drives disassembly. These results suggest a functional role for syntaxin clusters as reservoirs and a corresponding role for NSF in syntaxin liberation and SNARE protein quality control preceding fusion.

neuroscience↗

bioRxiv2024

Engineered Nanotopographies Induce Transient Openings in the Nuclear Membrane

Materials with engineered nano-scale surface topographies, such as nanopillars, nanoneedles, and nanowires, mimic natural structures like viral spike proteins, enabling them to bypass biological barriers like the plasma membrane. These properties have led to applications in nanoelectronics for intracellular sensing and drug delivery platforms, some of which are already in clinical trials. Here, we present evidence that nanotopographic materials can induce transient openings in the nuclear membranes of various cell types without penetrating the cells, breaching the nucleo-cytoplasmic barrier and allowing uncontrolled molecular exchange across the nuclear membrane. These openings, induced by nanoscale curvature, are temporary and repaired through ESCRT-mediated mechanisms. Our findings suggest a potential for nano topographic materials for direct nuclear sensing and delivery, holding promise for improving the delivery, efficiency, and safety of therapeutic agents to the nucleus.

bioengineering↗

bioRxiv2023

JAMS - A framework for the taxonomic and functional exploration of microbiological genomic data

Shotgun microbiome sequencing analysis presents several challenges to accurately and consistently depict sample composition and functional potential. Here we present a two-part framework - JAMS (Just a Microbiology System) - whereby with raw fastq files and metadata as input, meaningful analysis within a sample and between a sample can be performed with ease for either shotgun or 16S sequences. JAMS is the first package to provide seamless deconvolution of functions into their taxonomic contributors. We validated our JAMS framework on two human gut shotgun metagenome test datasets against the popular tool MetaPhlAn 4. We further demonstrate the application of the JAMS package, particularly the plotting functions, on a mouse shotgun dataset.

bioinformatics↗

bioRxiv2023

SeqScreen-Nano: a computational platform for rapid, in-field characterization of previously unseen pathogens.

The COVID-19 pandemic forever underscored the need for biosurveillance platforms capable of rapid detection of previously unseen pathogens. Oxford Nanopore Technology (ONT) couples long-read sequencing with in-field capability, opening the door to real-time, in-field biosurveillance. Though a promising technology, streaming assignment of accurate functional and taxonomic labels with nanopore reads remains challenging given: (i) individual reads can span multiple genes, (ii) individual reads may contain truncated genes, and pseudogenes, (iii) the error rate of the ONT platform that may introduce frameshifts and missense errors, and (iv) the computational costs of read-by-read analysis may exceed that of in-field computational equipment. Altogether, these challenges highlight a need for novel computational approaches. To this end, we describe SeqSeqscreen-Nano, a novel and portable computational platform for the characterization of novel pathogens. Based on results from simulated and synthetic microbial communities, SeqScreen-Nano can identify Open Reading Frames (ORFs) across the length of raw ONT reads and then use the predicted ORFs for accurate functional characterization and taxonomic classification. SeqScreen-Nano can run efficiently in a memory-constrained environment (less than 32GB of RAM), allowing it to be utilized in resource-limited settings. SeqScreen-Nano can also process reads directly from the ONT MinlON sequencing device, enabling rapid, in-field characterization of previously unseen pathogens. SeqScreen-Nano (v4.0) is available on GitLab at: https://gitlab.com/treangenlab/seqscreen

bioinformatics↗

bioRxiv2021

SeqScreen: Accurate and Sensitive Functional Screening of Pathogenic Sequences via Ensemble Learning

The COVID-19 pandemic has emphasized the importance of detecting known and emerging pathogens from clinical and environmental samples. However, robust characterization of pathogenic sequences remains an open challenge. To this end, we developed SeqScreen, which can accurately characterize short nucleotide sequences using taxonomic and functional labels, and a customized set of curated Functions of Sequences of Concern (FunSoCs) specific to microbial pathogenesis. We show our ensemble machine learning model can label protein-coding sequences with FunSoCs with high recall and precision. SeqScreen is a step towards a novel paradigm of functionally informed pathogen characterization and is available for download at: www.gitlab.com/treangenlab/seqscreen

bioinformatics↗

bioRxiv2020

Building a virtual summer research experience in cancer for high school and early undergraduate students: lessons from the COVID-19 pandemic

BackgroundThe COVID-19 pandemic posed a unique challenge for summer research programs in 2020, particularly for programs aimed at hands-on experience for younger trainees. The Indiana University Melvin and Bren Simon Comprehensive Cancer Center supports two pipeline programs, which traditionally immerse high school juniors, seniors, and early undergraduate students from underrepresented populations in science in hands-on projects in cancer biology labs. However, due to social distancing policies during the pandemic and reduction of research operations, these students were not physically allowed on campus. Thus, the authors set out to strategically pivot to a wholly virtual curriculum and evaluate the Virtual Summer Research Experience in Cancer outcomes. MethodsThe virtual program included four components: 1. a core science and professional development curriculum led by high school teachers and senior undergraduates; 2. faculty-delivered didactic sessions on cancer science; 3. mentored, virtual research projects with research faculty; and 4. online networking events to encourage vertical mentoring. Outcomes data were measured using an 11-item Research Preparation scale, daily electronic feedback, and structured evaluation and feedback via Zoom weekly. ResultsOutcome data suggested high self-reported satisfaction with the virtual program. Outcome data also revealed the importance of coordination between multiple entities for seamless program implementation. This includes the active recruitment and participation of high school teachers and further investment in information technology capabilities of institutions. ConclusionsFindings reveal a path to educate and train high school and early undergraduate students in cancer research when hands-on, in-person training is not feasible. Virtual research experiences are not only useful to engage students during public health crises but can provide an avenue for cancer centers to expand their cancer education footprints to remotely located schools and universities with limited resources to provide such experiences to their students.

scientific communication and education↗

bioRxiv2020

Biochemical Validation of a Fourth Guanidine Riboswitch Class in Bacteria

An intriguing consequence of ongoing riboswitch discovery efforts is the occasional identification of metabolic or toxicity response pathways for unusual ligands. Recently, we reported the experimental validation of three distinct bacterial riboswitch classes that regulate gene expression in response to the selective binding of a guanidinium ion. These riboswitch classes, called guanidine-I, -II and -III, regulate numerous genes whose protein products include previously misannotated guanidine exporters and enzymes that degrade guanidine via an initial carboxylation reaction. Guanidine is now recognized as the primal substrate of many multidrug efflux pumps that are important for bacterial resistance to certain antibiotics. Guanidine carboxylase enzymes had long been annotated as urea carboxylase enzymes but are now understood to participate in guanidine degradation. Herein we report the existence of a fourth riboswitch class for this ligand, called "guanidine-IV". Members of this class use a novel aptamer to selectively bind guanidine and use an unusual expression platform arrangement that is predicted to activate gene expression when ligand is present. The wide distribution of this abundant riboswitch class, coupled with the striking diversity of other guanidine-sensing RNAs, demonstrates that many bacterial species maintain sophisticated sensory and genetic mechanisms to avoid guanidine toxicity. This finding further highlights the mystery regarding the natural source of this nitrogen-rich chemical moiety.

biochemistry↗

bioRxiv2020

KOMB: Taxonomy-oblivious Characterization of Metagenome Dynamics via K-core Decomposition

Characterizing metagenomic samples via kmer-based, database-dependent taxonomic classification methods has provided crucial insight into underlying host-associated microbiome dynamics. However, novel approaches are needed that are able to track microbial community dynamics within metagenomes to elucidate genome flux in response to perturbations and disease states. Here we describe KOMB, a novel approach for tracking homologous regions within microbiomes. KOMB utilizes K-core graph decomposition on metagenome assembly graphs to identify repetitive and homologous regions to varying degrees of resolution. K-core performs a hierarchical decomposition which partitions the graph into shells containing nodes having degree at least K, called K-shells, yielding O(V + E) complexity compared to exact betweenness centrality complexity of O(V E) found in prior related approaches. We show through rigorous validation on simulated, synthetic, and real metagenomic datasets that KOMB accurately recovers and profiles repetitive and homologous genomic regions across organisms in the sample. KOMB can also identify functionally-rich regions in Human Microbiome Project (HMP) datasets, and can be used to analyze longitudinal data and identify pivotal taxa in fecal microbiota transplantation (FMT) samples. In summary, KOMB represents a novel approach to microbiome characterization that can efficiently identify sequences of interest in metagenomes.

bioinformatics↗