Biology subjects

Tiwari, B.

Publications and source records attributed to Tiwari, B..

10 recordsLinked to original sources

bioRxiv2026

LINE-1 transposon derepression and epigenetic remodeling in Retinoblastoma

Retinoblastoma (RB), the most common pediatric intraocular malignancy, is initiated by biallelic inactivation of the RB1 tumor suppressor gene. Although LINE-1 (L1) retrotransposon activation has been linked to genomic instability and tumor evolution in several adult cancers, its contribution to pediatric malignancies remains poorly understood. Here, we investigated L1 expression in retinoblastoma samples from an Indian pediatric cohort by integrating transcriptomic, locus-specific, and immunohistochemistry. Transcriptomic profiling revealed increased L1 expression in RB tumors compared with control retinal tissues, which was validated by ORF1p immunostaining in tumor sections. Locus-specific analysis identified a subset of transcriptionally active L1 loci, suggesting selective activation of young L1 elements. We also observed altered expression of genes associated with proliferation, chromatin regulation, and oncogenic signaling, including MYCN, MDM2, E2F3, SOX4 and CREBBP. Our findings identify previously underexplored L1 activation as a molecular feature associated with retinoblastoma and suggest that disruption of transposon silencing accompanies the RB transcriptional landscape. Validation in larger patient cohorts will be important; however, this study provides a foundation for exploring L1 activity as a potential molecular marker and future therapeutic vulnerability in retinoblastoma. HighlightsO_LITranscriptomic profiling reveals increased LINE-1 expression in retinoblastoma. C_LIO_LIORF1p protein is robustly expressed in retinoblastoma. C_LIO_LIA subset of LINE-1 loci exhibits transcriptional activation, suggesting locus-specific derepression. C_LI In BriefRetinoblastoma is initiated by RB1 loss, but the contribution of transposable elements to its molecular landscape remains poorly understood. Here, we show that retinoblastoma is associated with LINE-1 promoter hypomethylation, increased LINE-1 transcript expression, ORF1p accumulation, and preferential activation of selected LINE-1 loci. These changes occur together with altered expression of oncogenic signalling pathways and host factors involved in transposable element repression, identifying dysregulated LINE-1 control as a molecular feature of this paediatric cancer. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=115 SRC="FIGDIR/small/740688v1_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@197e89org.highwire.dtl.DTLVardef@1c26c04org.highwire.dtl.DTLVardef@14180a4org.highwire.dtl.DTLVardef@16493bb_HPS_FORMAT_FIGEXP M_FIG C_FIG Retinoblastoma exhibits epigenetic activation of LINE-1 characterized by promoter hypomethylation, increased expression of young LINE-1 elements, and ORF1p accumulation. This is accompanied by coordinated transcriptomic remodeling and disruption of host mechanisms that normally restrain transposable elements, linking LINE-1 activation to the molecular landscape of retinoblastoma.

cancer biology↗

bioRxiv2026

Recurrent LINE 1 exonization drives transcriptome remodelling in NSCLC

The only autonomously active transposable elements in the human genome are Long interspersed nuclear element-1 (LINE-1) elements. These elements are known to play an important role in changing the transcriptome. LINE-1 sequences affect gene regulation during post-transcription processing, along with their established role in retrotransposition. Exonization is one mechanism where the LINE-1 integrated genome undergoes alternative splicing to produce new isoforms of transcripts. Our work mainly highlights the effect of LINE-1 associated exonization, focusing on the formation of isoforms of transcripts. Using Non-small cell lung cancer (NSCLC) as a model, we conducted a detailed transcriptome study that combines splice junction profiling with gene expression data. Our results show that LINE-1 sequences are often included as exons in host transcripts, leading to the formation of new exons and their various isoforms. The events are validated by solid splice junction evidence that proves the reliability and reproducibility. In particular, it was observed that repetitive analyses revealed certain LINE-1 exonization events that were consistent. The finding indicates that LINE-1 act as recurrent sources of splice ready sequences. Though exonizations do not necessarily affect the total expression levels of genes, our study reveals that they certainly contribute to transcript diversity. The diversity of isoforms generated potentially contributes to the effects of gene function. This study is limited to NSCLC, but it is likely that the exonizations events play a crucial role in the altering RNA diversity in cancers. Therefore the study elucidates new insights into how transposable elements modify gene structure and function during cancer development.

genomics↗

bioRxiv2026

LINE-1 ORF2p endonuclease activity links DNA damage signalling to enhancer associated chromatin remodeling

Aberrant expression of Long Interspersed Element-1 (LINE-1/L1) retrotransposons is increasingly linked to genomic instability in cancer, particularly in the context of compromised p53 function. The endonuclease (EN) domain of the LINE-1 ORF2 protein (ORF2p) generates DNA strand breaks, yet its role in shaping downstream chromatin and transcriptional responses remains poorly defined. Here, we investigate the impact of ORF2p-EN activity on DNA damage response signalling and epigenomic remodelling in A375 melanoma cells using a L1 expression system. ORF2p-EN led to activation of ATM dependent DNA damage signalling, evidenced by increased {gamma}H2AX accumulation. The EN induced DNA damage showed enrichment of yH2AX at fragile hotspot regions, consistent with localized DNA damage. We also observed increase in levels of the DNA repair factor XRCC5, suggesting a possible engagement of non-homologous end joining pathway. EN-induced DNA damage correlated with increased P300 activity and enhanced H3K27ac enrichment at regulatory regions, linking DNA damage signalling with chromatin acetylation. Pharmacological inhibition of ATM attenuated both {gamma}H2AX accumulation and H3K27ac levels, supporting a model in which chromatin remodelling occurring downstream of DNA damage signalling. Collectively, our findings position ORF2p endonuclease activity as an initiating source of DNA damage that is followed by chromatin remodelling and transcriptional reprogramming, with p53 acting as a critical modulator of this axis. These results provide mechanistic insight into how LINE-1 activation may contribute to oncogenic gene expression programs in cancer.

cancer biology↗

bioRxiv2025

A choanoflagellate cGLR-STING pathway reveals evolutionary links between bacterial and animal immunity

Animal innate immunity evolved from ancient pathways in bacterial anti-phage defense. How bacterial immune components were first acquired and adapted within eukaryotic cells remains poorly understood. Here we identify a complete cGLR-STING signaling axis in choanoflagellates, the closest living relatives of animals, that exhibits a mosaic of features from both bacterial and animal immunity. Comparative genomics reveals choanoflagellate cGLR and STING genes organized in operon-like arrangements reminiscent of bacterial defense loci. Reconstitution of choanoflagellate cGLR-STING signaling in vitro demonstrates that activation occurs through the conserved nucleotide immune signal 2'3'-cGAMP. Structural analysis of a choanoflagellate STING-2'3'-cGAMP complex explains how retention of bacterial-like features in early eukaryotic proteins shapes ligand specificity and receptor activation. We analyze cGLR and STING evolution in unicellular eukaryotes and identify further STING homologs in choanoflagellates and fungi that support additional independent acquisition events. Our results reveal molecular fossils that bridge bacterial and animal immunity and illuminate early eukaryotic immune system evolution.

immunology↗

bioRxiv2024

p53 restores the silencing of activated L1 Transposons

Long Interspersed Nuclear Element 1 (LINE1/L1) retrotransposons, comprising around 17% of the human genome, typically remain quiescent in healthy somatic cells but become activated in various cancer types. Our recent investigation reveals that p53 silences L1 transposons in human somatic cells, potentially constituting a tumor suppressive pathway. In this study, we demonstrate that p53 silences both L1mRNA-gDNA (cis L1 R-loops) and L1mRNA-cDNA hybrids (trans L1 R-loops) formed during retrotransposition. The activation of L1 transposons by HDAC inhibitors (HDACi) led to accumulation of these cis and trans L1 R-loops in p53-/- cells, which were mitigated by treatment with a reverse transcriptase inhibitor. Furthermore, p53 established re-silencing of hyperactivated L1 transposons induced by HDACi. The p53-mediated restoration of silencing was accompanied by recruiting histone repressive marks specifically H3K9me3 and H3K27me3 and inhibiting the deposition of H3K4me3 and H3K9ac marks at the L1 promoter. This study elucidates a novel role of p53 in regulating the formation of RNA-DNA hybrids, a pivotal intermediate component of retrotransposition, and initiating the suppression of hyperactivated L1 elements. These findings underscore the significance of p53 in preserving genome stability through the regulation of L1-derived R-loops. In BriefThe role of L1 transposon derived L1mRNA-cDNA hybrids; an intermediate product formed during retrotransposition, in DNA damage and inflammation is not clear. Paul et al. reveals that p53 prevents L1cDNA derived RNA-DNA hybrids to control DNA damage and activation of inflammatory genes. The findings also elucidate the role of p53 in initiating the repression of hyperactivated transposons by facilitating the recruitment of epigenetic repressive marks and preventing the deposition of activating marks at L1-5UTR. HighlightsO_LIp53 loss facilitates accumulation of both cis (L1mRNA-gDNA) and trans (L1mRNA-cDNA) forms of L1 R-loops. C_LIO_LIThe youngest, actively retrotransposing full-length L1s contribute to the formation of trans (L1mRNA-cDNA) R-loops. C_LIO_LIp53 aids immediate L1 re-silencing by restoring deposition of epigenetic repressive and inhibition of activating marks. C_LIO_LIReverse transcriptase inhibitor prevents L1 mediated DNA damage. C_LI Subject Categories: L1/LINE1, p53, Retrotransposons, RNA-DNA hybrids, Cis R loops, Trans R-loop, L1/LINE1 Graphical O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=123 SRC="FIGDIR/small/589154v3_ufig1.gif" ALT="Figure 1"> View larger version (37K): org.highwire.dtl.DTLVardef@1e37111org.highwire.dtl.DTLVardef@11430c8org.highwire.dtl.DTLVardef@8e9c0corg.highwire.dtl.DTLVardef@a6e98d_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2023

Ultrasound-mediated drug diffusion, uptake, and cytotoxicity in glioblastoma 3D tumour sphere model

A myriad of biological effects can be stimulated by the ultrasound (US) for the treatment of cancer. The objective of our research was to investigate the effect of ultrasound alone and in combination with the chemotherapeutic drugs such as doxorubicin (DOX) and temozolomide (TMZ) on human glioblastoma (GBM) and the human epidermoid carcinoma cancer 2D and 3D cell cultures. Results indicated that the US 96-probe device could induce tumour sphere cytotoxicity in a dosage- and time-dependent manner, with multiple treatments augmenting this cytotoxic effect. With enhanced cytotoxicity, US decreased tumour sphere growth metabolic activity, disrupted spheroid integrity, and heightened the occurrence of DNA double strand breaks, resulting in damage to tumour spheres and an inability to rebuild tumour spheres after multiple US treatments. The combination of US and TMZ / DOX enhanced the efficiency of treatment for GBM and epidermoid carcinoma by enhancing induced cytotoxicity in 3D tumour spheres compared to 2D monolayer cells, and also by increasing the incubation time, which is the most crucial way to differentiate between the effectiveness of drug treatment with and without US. In conclusion, our data demonstrate that US enhances drug diffusion, uptake, and cytotoxicity using 3D spheroid models when compared with 2D cultures. It also demonstrates the significance of 3D cell culture models in drug delivery and discovery research. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=114 SRC="FIGDIR/small/564145v1_ufig1.gif" ALT="Figure 1"> View larger version (34K): org.highwire.dtl.DTLVardef@182e90org.highwire.dtl.DTLVardef@1992240org.highwire.dtl.DTLVardef@97a8fborg.highwire.dtl.DTLVardef@d71e05_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2021

Synergistic Cytotoxicity Between Cold Atmospheric Plasma and Pyrazolopyrimidinones Against Glioblastoma Cells

Pyrazolopyrimidinone is a fused nitrogen-containing heterocyclic system, which acts as a core scaffold in many pharmaceutically relevant compounds. Pyrazolopyrimidinones have been demonstrated to be efficient in treating several diseases, including cystic fibrosis, obesity, viral infection and cancer. We have tested the synergistic anti-cancer effects of 15 pyrazolopyrimidinones, synthesised in a two-step process, combined with cold atmospheric plasma (CAP), a novel innovation generating reactive species with other unique chemical and physical effects. We identify two pyrazolopyrimidinones that act as prodrugs and display enhanced reactive-species dependent cytotoxicity when used in combination with cold atmospheric plasma. Synergistic activation was evident for both direct CAP treatment on prodrug loaded tumour cells and indirect CAP treatment of prodrug in media prior to adding to tumour cells. Our results demonstrate the potential of CAP combined with pyrazolopyrimidinones as a programmable cytotoxic therapy against cancer. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=113 SRC="FIGDIR/small/429831v1_ufig1.gif" ALT="Figure 1"> View larger version (47K): org.highwire.dtl.DTLVardef@56c8aeorg.highwire.dtl.DTLVardef@1913ea9org.highwire.dtl.DTLVardef@a8f522org.highwire.dtl.DTLVardef@75a529_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2021

Pin Electrode Reactor: A novel cold atmospheric plasma device and its potential in glioblastoma treatment

Glioblastoma multiforme (GBM) is the most common and biologically aggressive brain tumour. The current standard therapy for GBM consists in surgical resection, followed by radiotherapy and chemotherapy. Yet, the treatment is limited due to the area for the surgical resection and for the inability of some drugs to cross the brain blood barrier, leading to a general prognostic of no more than a year. Cold atmospheric plasma (CAP) is a new approach in the treatment of this challenging disease. CAP interaction with cells is dependent on physical and chemical factors, with different plasma discharges, cell type, and culture conditions leading to different CAP activity. Considering the plasma self-adaptation that different plasma discharge modes can undergo, which leads to different interaction plasma/cells, the characterization of a new device is essential. In this study we analysed the effect of a novel large pin-to-plate non-thermal atmospheric plasma on U-251 MG cells under different conditions. The analysis of reactive oxygen and nitrogen species (RONS) on plasma, media and cells were also assessed. We were able to demonstrate that the pin-to-plate device is cytotoxic to GBM cells in a dose, time and ROS dependent manner. The measurements of RONS on plasma/media also give us an insight on the chemical effect of this novelty device, and the possibility to better understand the use of this device as a promising GBM therapy.

cancer biology↗

bioRxiv2019

Low dose Cold Atmospheric Plasma induces membrane oxidation, stimulates endocytosis and enhances uptake of nanomaterials in Glioblastoma multiforme cells

Cold atmospheric plasma (CAP) has demonstrated synergistic cytotoxic effects with nanoparticles, especially promoting the uptake and accumulation of nanoparticles inside cells. However, the mechanisms driving the effects need to be explored. In this study, we investigate the enhanced uptake of theranostic nanomaterials by CAP. Numerical modelling of the uptake of gold nanoparticle into U373MG Glioblastoma multiforme (GBM) cells predicts that CAP may introduce a new uptake route. We demonstrate that cell membrane repair pathways play the main role in this stimulated new uptake route, following non-toxic doses of dielectric barrier discharge CAP (30 s, 75 kV). CAP treatment induces cellular membrane damage, mainly via lipid peroxidation as a result of reactive oxygen species (ROS) generation. Membranes rich in peroxidated lipids are then trafficked into cells via membrane repairing endocytosis. We confirm that the enhanced uptake of nanomaterials is clathrin-dependent using chemical inhibitors and silencing of gene expression. Therefore, CAP-stimulated membrane repair increases endocytosis and accelerates the uptake of gold nanoparticles into U373MG cells after CAP treatment. Our data demonstrate the utility of CAP to model membrane oxidative damage in cells and characterise a previously unreported mechanism of membrane repair to trigger nanomaterial uptake which will be useful for developing more efficient deliveries of nanoparticles and pharmaceuticals into cancer cells for tumour therapy and diagnosis. This mechanism of RONS-induced endocytosis will also be of relevance to other cancer therapies that induce an increase in extracellular RONS.\n\n\n\nO_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=104 SRC=\"FIGDIR/small/805192v1_ufig1.gif\" ALT=\"Figure 1\">\nView larger version (43K):\norg.highwire.dtl.DTLVardef@1487fd4org.highwire.dtl.DTLVardef@c05eeforg.highwire.dtl.DTLVardef@198c75aorg.highwire.dtl.DTLVardef@1481096_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2019

Impact of small RNAs in retrograde signalling pathways in Arabidopsis thaliana

Chloroplast perturbations activate retrograde signalling pathways causing dynamic changes of gene expression. Besides transcriptional control of gene expression different classes of small non-coding RNAs (sRNAs) act in gene expression control, but comprehensive analyses regarding their role in retrograde signalling is lacking. We performed sRNA profiling in response to norflurazon (NF) that provokes retrograde signals in A. thaliana wild type and the two retrograde signalling mutants gun1 and gun5. The RNA samples were also used for mRNA and long non-coding RNA (lncRNA) profiling to link altered sRNA levels to changes of their cognate target RNAs. We identified 122 sRNAs from all known sRNA classes that were responsive to NF in wild type. Strikingly, 140 and 213 sRNAs were found to be differentially regulated in both mutants indicating a retrograde control of these sRNAs. Concomitant with the changes in sRNA expression we detected about 1500 differentially expressed mRNAs in the NF treated wild type and around 900 and 1400 mRNAs that were differentially regulated in the gun1 and gun5 mutant with a high proportion (~30%) of genes encoding plastid proteins. Furthermore, around 20% of predicted miRNA targets code for plastid localised proteins. The analyses of sRNA-target pairs identified pairs with an anticorrelated expression as well pairs showing other expressional relations pointing to a role of sRNAs in balancing transcriptional changes upon retrograde signals. Based on the comprehensive changes in sRNA expression we assume a considerable impact of sRNAs in retrograde-dependent transcriptional changes to adjust plastidic and nuclear gene expression. Significance statementPerturbations of plastid functions trigger retrograde signalling to adjust plastidic and nuclear gene expression, however, the role of small non-coding RNAs acting as regulators in these pathways is not well understood. We analysed small non-coding RNA expression in response to retrograde signals in A. thaliana wild type and two retrograde signalling mutants and identified members of all known small non-coding RNA classes pointing to a functional role of these RNA classes in retrograde pathways.

plant biology↗