Biology subjects

Behera, A.

Publications and source records attributed to Behera, A..

12 recordsLinked to original sources

bioRxiv2026

AMPK Signalling Rewires WWP1 mediated RUNX1 Ubiquitination in CML Progression

Despite the remarkable success of BCR:ABL1-targeted tyrosine kinase inhibitors, disease progression and therapeutic resistance remain major challenges in chronic myeloid leukemia (CML), highlighting the need to identify additional molecular mechanisms that sustain leukemic survival. In this study, we identify the HECT-type E3 ubiquitin ligase WWP1 as a previously unrecognized regulator of RUNX1 and define a signaling network linking ubiquitination to AMP-activated protein kinase (AMPK)-dependent phosphorylation in CML. We demonstrate that WWP1 directly associates with the PPxY motif of RUNX1 and catalyzes its K63-linked polyubiquitination, thereby enhancing protein stability and promoting leukemic cell growth. Disruption of WWP1, either by genetic silencing or pharmacological inhibition, diminished RUNX1 ubiquitination, impaired proliferation, induced G2/M cell-cycle arrest, and triggered apoptosis. Mechanistic investigations further revealed that WWP1 is a direct substrate of AMPK. Phosphorylation of WWP1 at Thr692 by AMPK promoted its lysosomal degradation, thereby limiting WWP1-dependent ubiquitination of RUNX1. In parallel, AMPK-mediated phosphorylation of RUNX1 reduced its leukemogenic activity, collectively shifting the signaling balance toward growth suppression. Activation of AMPK with metformin reproduced these molecular effects in leukemia cell lines, primary CML patient-derived cells, and xenograft tumors, supporting the physiological relevance of this pathway. Together, our findings establish an AMPK-WWP1-RUNX1 signaling axis that integrates phosphorylation and non-proteolytic ubiquitination to regulate RUNX1 function and CML progression. These results identify WWP1 as a potential therapeutic vulnerability and provide a mechanistic rationale for targeting the AMPK-WWP1 pathway in CML.

cancer biology↗

bioRxiv2026

Amaranthus viridis-derived phytopriming reprograms redox homeostasis and limits arsenic accumulation in rice

Arsenic (As) poses a major threat to rice productivity and food safety due to its high bioaccumulation potential and subsequent entry into the human food chain. In rice, As impairs seed germination, disrupts morpho-anatomical development, and induces oxidative stress. This study evaluates seed priming with an aqueous extract of the agricultural weed Amaranthus viridis (AvE) as a sustainable strategy to alleviate As-induced phytotoxicity. AvE priming significantly improved germination (71-75%) and morpho-physiological performance under As stress. It reduced oxidative stress markers, including H2O2 (21-38%), malondialdehyde (13-26%), and proline (18.9-44.7%), while increasing antioxidant metabolites, polyphenols and glutathione by up to 2.34-fold and 41%, respectively. Microscopy confirmed restoration of cellular integrity and anatomical organisation in primed seedlings. ICP-OES analysis showed that AvE priming reduced root As uptake by up to 39%, root-to-shoot translocation by up to 58%, and grain As accumulation by up to 95% compared with unprimed plants. qRT-PCR revealed modulation of genes involved in As homeostasis, indicating coordinated physiological and transcriptional responses. Importantly, improved agronomic performance further demonstrated the translational potential of this approach. This study provides the first evidence that A. viridis extract is a cost-effective, sustainable biostimulant for producing low-As rice in contaminated regions.

plant biology↗

bioRxiv2026

Phage-assisted continuous evolution of enzymes for noncanonical tyrosine biosynthesis

Genetic code expansion introduces new-to-nature chemical moieties into ribosomally synthesized proteins. In practice, the scope of functional groups that can be accessed using this method is often limited by noncanonical amino acid (ncAA) availability. Producing ncAAs directly in cells can circumvent poor ncAA uptake or commercial unavailability, but limited enzymes suitable for this application exist. In vitro evolution campaigns have been remarkably successful in yielding synthetically useful "ncAA synthases." However, these enzymes are optimized for preparative-scale synthesis and their activities often do not translate well to cellular biosynthesis. Thus, expanding strategies to engineer enzymes specifically for ncAA production within cells will benefit further implementation of genetic code expansion. Here, we use phage-assisted noncontinuous and continuous evolution to evolve enzymes for improved synthesis of non-canonical tyrosine derivatives in E. coli. Using simple serial passaging, we uncovered mutations that doubled the production of an expensive ncAA, 3-methoxytyrosine, by tyrosine phenol lyase, and furthermore evolved variants that enable 3-iodotyrosine biosynthesis, a transformation the parent enzyme is unable to catalyze. Additionally, we evolved a recently reported tyrosine synthase for improved production of 3-halogenated tyrosines, identifying variants that exhibit high activity even at low substrate concentrations owing to a [~]8-fold reduction in KM. Our results demonstrate that phage assisted evolution can be used to rapidly improve the activity of enzymes for ncAA production in cells.

synthetic biology↗

bioRxiv2025

Comparative study of Dog and Human Lipid Droplet-associated protein (PLIN2)

Intracellular lipid droplets (LDs) are vital organelles that store energy, manage oxidative stress, and coordinate cellular signaling. A key protein family associated with LDs is the perilipins, with perilipin 2 (PLIN2) being prominent. This study focuses on a comparative analysis of canine and human PLIN2 to understand their structural and functional similarities. The genomic data for PLIN2 in both species were curated, revealing location, transcripts, exons, and introns. Sequence analysis demonstrated 85% identity between dog and human PLIN2, with conserved residues essential for structural and functional roles. Amino acid composition analysis revealed similarities in charged residues and in stabilizing and destabilizing residues. Despite their structural similarities (72% common secondary structure elements), a 3D comparison revealed significant differences in conformation. Physicochemical properties, including stability indices, were found to be distinct, possibly influencing their interactions. The study highlights the conservation of PLIN2s functional regions, shedding light on its role in protecting LDs from degradation across species.

bioinformatics↗

bioRxiv2025

Inhibiting ezrin triggers formin-mediated actin remodelling reducing cellular mechano-protection

Ezrin plays a crucial role in linking the cortical actin cytoskeleton to the plasma membrane in animal cells. Inactivated by small molecule ezrin inhibitors (EzrInh), ezrin is known to regulate cellular functions like adhesion and motility. However, whether ezrin plays any significant role in protecting cells against mechanical stresses is less explored. We use hypo-osmotic shock (HS) to challenge cells and study the mechano-protective role of ezrin. Inhibition of ezrin phosphorylation led to increased cell rupture rates on hypo-shock, with a greater impact at later time points ([~] 10 min). The time scales match with that of enhanced tension contrast at edges at the basal plane. We also observed a lowered relative retraction rate at protrusions created by hypo-shock and a less contractile mid-plane cortex - both indicate the role of a weakening of the cortex by EzrInh in compromising membrane integrity on hypo-shock. Concomitantly, EzrInh also increased stress fibres (SFs), traction forces and cell spreading clearly indicative of additional cellular pre-stress. Utilizing formin inhibitor (SMIFH2), we showed that formin-mediated actin polymerization was critical for the SF response to EzrInh while it also prevented the subsequent EzrInh-treatment from enhancing cell rupture rates. Together EzrInh reduced mechano-protection in cells by tilting of balance of actins organization from the cortex to SF enhancing formin-mediated SF assembly. HighlightsO_LIHypo-osmotically shocked CHO cells rupture more on ezrin inhibition C_LIO_LIEzrin inhibition weakens contractility of cortical actin but enhances stress fibre numbers at the base C_LIO_LIHypo-osmotic shock creates basal protrusions that are also retracted. C_LIO_LIRetractive potential in cells is weakened by ezrin inhibition C_LIO_LIFormin inhibitor prevents net increase in stress fibres by inhibition of ezrin C_LIO_LIOn pre-treatment with formin inhibitor, subsequent inhibition of ezrin doesnot render cells more vulnerable to hypo-osmotic shock C_LI

biophysics↗

bioRxiv2024

U2AF1 S34F enhances tumorigenic potential of lung cells by exhibiting synergy with KRAS mutation and altering response to environmental stress

Although U2AF1S34F is a recurrent splicing factor mutation in lung adenocarcinoma (ADC), U2AF1S34F alone is insufficient for producing tumors in previous models. Because lung ADCs with U2AF1S34F frequently have co-occurring KRAS mutations and smoking histories, we hypothesized that tumor-forming potential arises from U2AF1S34F interacting with oncogenic KRAS and environmental stress. To elucidate the effect of U2AF1S34F co-occurring with a second mutation, we generated human bronchial epithelial cells (HBEC3kt) with co-occurring U2AF1S34F and KRASG12V. Transcriptome analysis revealed that co-occurring U2AF1S34F and KRASG12V differentially impacts inflammatory, cell cycle, and KRAS pathways. Subsequent phenotyping found associated suppressed cytokine production, increased proliferation, anchorage-independent growth, and tumors in mouse xenografts. Additionally, HBEC3kts harboring U2AF1S34F alter splicing in stress granule protein genes, show increased stress granule formation, and have increased viability in cigarette smoke concentrate. Our results suggest that U2AF1S34F may potentiate transformation by granting precancerous cells survival advantage in environmental stress, permitting accumulation of additional mutations, like in KRAS, which synergize with U2AF1S34F to transform the cell.

cancer biology↗

bioRxiv2024

Light and temperature-dependent developmental role of Auxin Binding Protein 1 (ABP1) in Arabidopsis thaliana

Auxin Binding Protein 1 (ABP1) is a small glycoprotein of about 22 kDa that has long been debated as the auxin receptor, and has been put into question for its unclear functions. Despite its conservancy during land plant evolution, its precise role in plant development is still elusive. Historically, it has been implicated in various rapid responses such as membrane polarization, calcium fluxes, TMK1-based cell-surface signalling, auxin canalization, etc. A relatively recent observation questioning the role of ABP1 in plant development led us to explore its probable functions if any. In the current study, we reinvestigated the plausible function of ABP1 using its CRISPR-based loss-of-function mutants, namely abp1-C1 and abp1-C2. Here we show that, ABP1 acts as a positive regulator for primary root elongation under red and secondary root elongation under blue light in seedlings at 22 {degrees}C. Under red light at 18 {degrees}C, it has a negative effect on hypocotyl growth inhibition. Furthermore, it is involved in flowering time control at 18 {degrees}C irrespective of the photoperiod. We show that the transcript levels of Phytochrome B (phyB) and GIGANTEA (GI) are altered in the mutants of ABP1 under red light and low temperature (18 {degrees}C) regimes. Further, ABP1 show a pronounced role in tolerance to dehydration induced due to low temperature (18 {degrees}C), which correlates with an increase in endogenous abscisic acid (ABA), salicylic acid (SA), and a decrease in jasmonic acid (JA) content in leaves. The functional roles of ABP1 under red light, low temperature and dehydration tolerance in Arabidopsis thaliana once again frames it to be an important regulator under adverse and varied conditions that the plant can experience, and thus opened up new avenues for further studies.

plant biology↗

bioRxiv2023

Connexin 43 mediated collective cell migration is independent of Golgi orientation

Cell migration is vital for multiple physiological functions and is involved in the metastatic dissemination of tumour cells in various cancers. For effective directional migration, cells often reorient their secretory traffic towards the leading edge by reorienting the Golgi apparatus. However, conflicting results on the positioning of the Golgi relative to the migrating cell raise questions about its regulation. Herein, we address the role of gap junction protein Connexin 43, which connects cells allowing the direct exchange of molecules and also controls the distribution of microtubules. We utilized HeLa WT (wild-type cells lacking Connexin 43) and HeLa 43 cells (stably expressing Connexin 43). We found that functional Connexin 43 channels affected Golgi morphology and reduced the ability of reorientation of Golgi (towards the leading edge) during cell migration. Although Connexin 43 also reduced migration velocity, the front displayed enhanced coherence in movement, implying an augmented collective nature of migration compared to HeLa WT cells lacking the cell-cell connectivity evidenced in HeLa 43. The increase in vimentin and basal actin in HeLa 43 shows that Connexin 43 expression alters the cytoskeleton. Non-invasive measurement of basal membrane height fluctuations revealed that HeLa 43 had a lower membrane tension and a more uniform intra- and inter-cellular distribution than HeLa WT, similar to their coherence in migration. We, therefore, propose that the reduced Golgi reorientation in HeLa 43 might be linked to lesser dependence on directed secretory trafficking, which is believed to be required to buffer the increasing tension caused by migration.

biophysics↗

bioRxiv2021

Deformation of membrane vesicles due to chiral surface proteins

Chiral, rod-like molecules can self-assemble into cylindrical membrane tubules and helical ribbons. They have been successfully modeled using the theory of chiral nematics. Models have also predicted the role of chiral lipids in forming nanometer-sized membrane buds in the cell. However, in most theoretical studies, the membrane shapes are considered fixed (cylinder, sphere, saddle, etc.), and their optimum radius of curvatures are found variationally by minimizing the energy of the composite system consisting of membrane and chiral nematics. Numerical simulations have only recently started to consider membrane deformation and chiral orientation simultaneously. Here we examine how deformable, closed membrane vesicles and chiral nematic rods mutually influence each others shape and orientation, respectively, using Monte-Carlo (MC) simulation on a closed triangulated surface. For this, we adopt a discrete form of chiral interaction between rods, originally proposed by Van der Meer et al. (1976) for off-lattice simulations. In our simulation, both conical and short cylindrical tubules emerge, depending on the strength of the chiral interaction and the intrinsic chirality of the molecules. We show that the Helfrich-Prost term, which couple nematic tilt with local membrane curvature in continuum models, can account for most of the observations in the simulation. At higher chirality, our theory also predicts chiral tweed phase on cones, with varying bandwidths. PACS numbers: PACS : 87.16.-b, 87.15.Aa, 81.40.Jj, 87.15.Rn

biophysics↗

bioRxiv2021

Temperature-dependent Self assembly of biofilaments during red blood cell sickling

Molecular self-assembly plays vital role in various biological functions. However, when aberrant molecules self-assemble to form large aggregates, it can give rise to various diseases. For example, the sickle cell disease and Alzheimers disease are caused by self-assembled hemoglobin fibers and amyloid plaques, respectively. Here we study the assembly kinetics of such fibers using kinetic Monte-Carlo simulation. We focus on the initial lag time of these highly stochastic processes, during which self-assembly is very slow. The lag time distributions turn out to be similar for two very different regimes of polymerization, namely, a) when polymerization is slow and depolymerization is fast, and b) the opposite case, when polymerization is fast and depolymerization is slow. Using temperature dependent on- and off-rates for hemoglobin fiber growth, reported in recent in-vitro experiments, we show that the mean lag time can exhibit non-monotonic behaviour with respect to change of temperature.

biophysics↗

bioRxiv2019

Confined filaments in soft vesicles - case of sickle red blood cells

A semi-rigid filament confined in a soft vesicle of similar size can mutually deform each other. An important example from biological context is Hemoglobin-S (HbS) fibers which polymerize inside red blood cell (RBC). The fibers deform the healthy RBC into sickle-like shape causing difficulty in blood flow through capillaries. Using an area difference elasticity (ADE) model for RBC and a worm-like chain model for the HbS fibers, confined within RBC, we study the shape deformations at equilibrium. We also consider multiple filaments and find that confinement can generate multipolar RBC shapes and can also promote helical filament conformations. The same model, in different parameter regime, reproduces tubulation for phospholipid vesicles, as seen in experiments, when microtubules are confined in the vesicle. We conclude that with a decrease in the surface area to volume ratio, and membrane rigidity, the vesicle prefers tubulation over sickling. Our simulations can access various non-axisymmetric shapes, which have been observed experimentally, both in the context of sickle RBC and phospholipid vesicles, but have so far remained beyond the scope of variational methods.

biophysics↗

bioRxiv2017

Mycobacterium tuberculosis LprE enhances bacterial persistence by inhibiting cathelicidin and autophagy in macrophages

Mycobacterium tuberculosis(Mtb) lipoproteins are known to facilitate bacterial survival by manipulating the host immune responses. Here, we have characterized a novel Mtb lipoprotein LprE(LprEMtb), and demonstrated its role in mycobacterial survival. LprEMtb acts by down-regulating the expression of cathelicidin, Cyp27B1, VDR and p38-MAPK via TLR-2 signaling pathway. Deletion of lprEMtb resulted in induction of cathelicidin and decreased survival in the host. Interestingly, LprEMtb was also found to inhibit autophagy mechanism to dampen host immune response. Episomal expression of LprEMtb in non-pathogenic Mycobacterium smegmatis(Msm) increased bacillary persistence by down-regulating the expression of cathelicidin and autophagy, while deletion of LprEMtb orthologue in Msm, had no effect on cathelicidin and autophagy expression. Moreover, LprEMtb blocked phago-lysosome fusion by suppressing the expression of EEA1, Rab7 and LAMP-1 endosomal markers by down-regulating IL-12 and IL-22 cytokines. Our results indicate that LprEMtb plays an important role in mycobacterial pathogenesis in the context of innate immunity.

physiology↗