Biology subjects

Tripathi, K.

Publications and source records attributed to Tripathi, K..

10 recordsLinked to original sources

bioRxiv2026

STING agonists in combination with epigenetic drugs potentiate ZNFX1-driven inflammatory necroptosis in TP53-mutated AML

TP53-mutated acute myeloid leukemia (AML) has dismal outcomes with current treatments and represents a critical unmet need. TP53-mutated AML is proposed to be susceptible to immunotherapeutic approaches but, to date, there is no established immunotherapy for this sub-group. Expression of stimulator of interferon genes (STING), a key innate immune driver that activates interferon (IFN) signaling, is decreased by epigenetic silencing or mutation in many cancers, including those with TP53 mutations. Here, we report that response to the next-generation synthetic STING agonist C92 is potentiated in AML cell lines and primary cells with TP53-mutated versus wild-type (WT) cells, representing a previously undescribed vulnerability of these leukemia cells to STING small molecule therapies. Moreover, combining treatment with the DNA methyltransferase inhibitor (DNMTi) decitabine (DAC), significantly increases STING activation, with marked transcriptome-wide increase in repetitive elements (REs) and upregulation of a critical set of interferon-related genes. Cell death in TP53 KO versus WT AML is specifically dependent on innate immune zinc finger NFX1-type containing 1 (ZNFX1) and Z-DNA-binding protein 1 (ZBP1) driving increased cleavage and activation of Receptor-Interacting-Serine/Threonine-Protein Kinase 3 (RIPK3) and mixed lineage kinase domain-like protein (MLKL), suggesting mechanisms of necroptosis. Finally, C92 and DAC combination significantly reduces leukemia burden in humanized AML mouse models, accompanied by increased immune responses, including cytokines and cytotoxic T lymphocytes in the leukemia microenvironment. These results support development of clinical trial strategies combining STING agonists with DNMTis for patients with TP53-mutated AML. SummaryO_LITP53-mutated AML potentiates effects of novel next-generation STING agonist C92, with unique allosteric and non-cyclic dinucleotide (non-CD) mechanism of action, inducing increased STING activation and cytokine release C_LIO_LISTING agonists and DNMTis, synergistically increase STING activation with marked transcriptome-wide increase in repetitive elements (REs) and upregulation of a critical set of interferon-related genes in TP53-mutated AML C_LIO_LISTING agonists induce necroptosis via a STING-ZNFX1-ZBP1-necroptosis axis in TP53-mutated AML. C_LIO_LIThis drug combination reduces leukemia burden, activates immune responses in AML models and supports translation for high-risk AML patients. C_LI Statement of Translational RelevanceThis pre-clinical study identifies a novel therapeutic vulnerability in (TP53)-mutated acute myeloid leukemia (AML), a poor prognosis subtype with a critical unmet need. Novel next-generation STING agonist C92, with unique allosteric and non-cyclic dinucleotide (non-CD) mechanism of action, induces increased STING activation and cytokine release, compared with WT TP53 in AML cell lines and primary cells, and has superior STING activity with respect to several STING agonists currently in clinical studies. Combining C92 treatment with the DNA methyltransferase inhibitor (DNMTi) decitabine (DAC) synergistically increases STING activation, with marked transcriptome-wide increase in repetitive elements (REs) and upregulation of a critical set of interferon-related genes, driving ZNFX1-driven inflammatory necroptotic cell death. Utilizing humanized mouse models, C92 in combination with DAC significantly reduces leukemia burden and enhances cytotoxic T-cell responses in the tumor microenvironment, supporting clinical translation for high-risk AML patients.

cancer biology↗

bioRxiv2026

QuadCleave: Enzymatic cleavage and analysis of DNA G-quadruplexes.

Prediction of G-quadruplexes (G4), noncanonical DNA secondary structures containing guanine tetrads, can have profound relevance across cancer, neurodegenerative and other genetic diseases, but require experimental validation by instrument intensive methods including CD spectroscopy, fluorescence, NMR, or crystallography. We report a novel, inexpensive, efficient and scalable 2-enzyme system for confirmation of G4 formation and copy number repeat count. First, QuadCleave, in which Mismatch Endonuclease I was used to repeatably, site-specifically and selectively cleave oligodeoxynucleotides containing exclusively G4-forming sequences. Second, HaeIII enzyme identified G4-forming sequences that can also generate GG/CC base pairs, and a sequences cleavage by both established it in C9ORF72, the most mutated gene in amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). These findings establish ubiquitously accessible experimental validation of G4 forming sequences and their copy number repeat assessment, with specificity to C9ORF72 oligodeoxynucleotides whose repeat expansion has critical clinical implications in a subpopulation of ALS and FTD patients.

molecular biology↗

bioRxiv2025

Endogenously generated Dutch-type Aβ nonfibrillar aggregates dysregulate presynaptic neurotransmission in the absence of detectable inflammation

Structured AbstractO_ST_ABSBackgroundC_ST_ABSAPPE693Q ("Dutch") transgenic mice develop aging-related learning deficits and accumulate endogenously generated nonfibrillar aggregates of A{beta} (NFA-A{beta}) and APP -carboxy terminal fragments. NFA-A{beta} correlates with synaptic loss and memory deficits more closely than does fibrillar A{beta}. MethodsWe assessed the physiological, transcriptomic, ultrastructural, histological, and metabolic changes associated with the accumulation of NFA of Dutch A{beta} in brains of APPE693Q mice. ResultsAging-related accumulation of NFA-A{beta} in APPE693Q mice was revealed by A11 immunohistochemistry and cyclic D,L--peptide-FITC microscopy. Presynaptic termini of APPE693Q mice developed physiological abnormalities in post-tetanic potentiation, synaptic fatigue, synaptic vesicle replenishment, and an aging-related reduction in mitochondrial complex I activity. Single-cell RNA sequencing showed that excitatory neurons exhibited an altered transcriptomic profile involving "protein translation" and "oxidative phosphorylation". DiscussionAccumulation of NFA-A{beta} alters neuronal metabolism but does not activate inflammation. Depletion of all forms of A{beta} may be required to eliminate A{beta} toxicity with anti-amyloid antibodies.

neuroscience↗

bioRxiv2024

ZNFX1 is a Novel Master Regulator in Epigenetically-induced Pathogen Mimicry and Inflammasome Signaling in Cancer

DNA methyltransferase and poly(ADP-ribose) polymerase inhibitors (DNMTis, PARPis) induce a stimulator of interferon (IFN) genes (STING)-dependent pathogen mimicry response (PMR) in ovarian (OC) and other cancers. We now show that combining DNMTis and PARPis upregulates expression of a little-studied nucleic-acid sensor, NFX1-type zinc finger-containing 1 protein (ZNFX1). We demonstrate that ZNFX1 is a novel master regulator for PMR induction in mitochondria, serving as a gateway for STING-dependent PMR. In patient OC databases, high ZNFX1 expression levels correlate with advanced stage disease. ZNFX1 expression alone significantly correlates with an increase in overall survival in a phase 3 trial for therapy-resistant OC patients receiving bevacizumab in combination with chemotherapy. In correlative RNA-seq data, inflammasome signaling through ZNFX1 correlates with abnormal vasculogenesis. ZNFX1 controls PMR signaling through the mitochondria and may serve as a biomarker to facilitate offering personalized therapy in OC patients, highlighting the strong translational significance of our findings. Significance statementDNA methyltransferase and poly(ADP-ribose) polymerase inhibitors upregulate expression of a novel nucleic-acid sensor, ZNFX1 that serves as a mitochondrial gateway to STING-dependent interferon/inflammasome signaling with tumor suppressor properties in ovarian cancer.

cancer biology↗

bioRxiv2024

Two liverworts from same habitats developed many similar but few distinct seasonal adaptive strategies: Insights from a transcriptomic approach

Liverworts are among the first land plants to experience seasonal changes after terrestrialization Therefore, we conducted a seasonal transcriptome analysis of two representative liverwort species from India, Dumortiera hirsuta and Plagiochasma appendiculatum, during their distinct growing seasons: the pre-monsoon, monsoon, post-monsoon, and fruiting season to understand the seasonal adaptations. Phylogenetic trees and evolutionary timescale analyses showed that D. hirsuta is primitive compared to P. appendiculatum. The RNA-seq analysis showed that D. hirsuta primarily modifies its transcriptome in the post-monsoon season but mainly induces specific genes in the fruiting season, most likely to develop reproductive organs and to adapt strategically by conserving energy in the fruiting season to deal with the harsh environmental conditions of both seasons. Conversely, P. appendiculatum exhibited significant transcriptome variability during both the fruiting and post-monsoon seasons, albeit to a lesser degree than D. hirsuta. This suggests that, to survive in the harsh conditions of both seasons, it strategically modulated its necessary gene expression levels over an extended period of time while taking energy conservation into consideration. This study offers the first comprehensive view of seasonal adaptive strategies employed by two liverworts that coexist in the same habitat but diverged at different stages in their evolutionary history.

genomics↗

bioRxiv2023

ACSL4 activity drives TNBC metastasis by positively regulating Histone H3 Acetylation mediated SNAIL expression

Triple-Negative Breast Cancer (TNBC) has profound unmet medical need globally for its devastating clinical outcome associated with rapid metastasis and lack of targeted therapies. Recently, lipid metabolic reprogramming has emerged as a major driver of breast cancer metastasis. Here, we unveil a strong association between the heightened expression of fatty acid metabolic enzyme, acyl-CoA synthetase 4 (ACSL4) and TNBC, which is primarily attributed by the selective absence of progesterone receptor (PR). Loss of ACSL4 function, either through genetic ablation or pharmacological inhibition significantly reduces metastatic potential of TNBC. Global transcriptome analysis reveals that ACSL4 activity markedly influences the gene expression pattern associated with TNBC migration. Mechanistically, ACSL4 alters fatty acid oxidation (FAO) and cellular acetyl-CoA levels, leading to the hyper-acetylation of particularly H3K27Ac and H3K9Ac marks resulting in overexpression of SNAIL during the course of TNBC metastatic spread to lymph node and lungs. Further, human TNBC metastasis exhibits positive correlation between ACSL4 and SNAIL expression. Altogether, our findings provide new molecular insights regarding the intricate interplay between metabolic alterations and epigenetic modifications, intertwined to orchestrate TNBC metastasis and posit a rational understanding for the development of ACSL4 inhibitors as a targeted therapy against TNBC.

cancer biology↗

bioRxiv2023

Molecular basis of RanGTP-activated nucleosome assembly with Histones H2A-H2B bound to Importin-9

Padavannil et al. 2019 show that Importin-9 (Imp9) transports Histones H2A-H2B from the cytoplasm to the nucleus using a non-canonical mechanism whereby binding of a GTP-bound Ran GTPase (RanGTP) fails to evict the H2A-H2B cargo. Instead, a stable complex forms, comprised of equimolar RanGTP, Imp9, and H2A-H2B. Unlike the binary Imp9*H2A-H2B complex, this RanGTP*Imp9*H2A-H2B ternary complex can release H2A-H2B to an assembling nucleosome. Here, we define the molecular basis for this RanGTP-activated nucleosome assembly by Imp9. We use hydrogen-deuterium exchange coupled with mass spectrometry and compare the dynamics and interfaces of the RanGTP*Imp9*H2A-H2B ternary complex to those in the Imp9*H2A-H2B or Imp9*RanGTP binary complexes. Our data are consistent with the Imp9*H2A-H2B structure by Padavannil et al. 2019 showing that Imp9 HEAT repeats 4-5 and 18-19 contact H2A-H2B, as well as many homologous importin*RanGTP structures showing that importin HEAT repeats 1 and 3, and the h8 loop, contact RanGTP. We show that Imp9 stabilizes H2A-H2B beyond the direct binding site, similar to other histone chaperones. Importantly, we reveal that binding of RanGTP releases H2A-H2B interaction at Imp9 HEAT repeats 4-5, but not 18-19. This exposes DNA- and histone-binding surfaces of H2A-H2B, thereby facilitating nucleosome assembly. We also reveal that RanGTP has a weaker affinity for Imp9 when H2A-H2B is bound. This may ensure that H2A-H2B is only released in high RanGTP concentrations near chromatin. We delineate the molecular link between the nuclear import of H2A-H2B and its deposition into chromatin by Imp9. SignificanceImp9 is the primary importin for shuttling H2A-H2B from the cytoplasm to the nucleus. It employs an unusual mechanism where the binding of RanGTP alone is insufficient to release H2A-H2B. The resulting stable RanGTP*Imp9*H2A-H2B complex gains nucleosome assembly activity as H2A-H2B can be deposited onto an assembling nucleosome. We show that H2A-H2B is allosterically stabilized via interactions with both N- and C-terminal portions of Imp9, reinforcing its chaperone-like behavior. RanGTP binding causes H2A-H2B release from the N-terminal portion of Imp9 only. The newly-exposed H2A-H2B surfaces can interact with DNA or H3-H4 in nucleosome assembly. Imp9 thus plays a multi-faceted role in histone import, storage, and deposition regulated by RanGTP, controlling histone supply in the nucleus and to chromatin.

biophysics↗

bioRxiv2022

Mechanism of RanGTP priming the release of H2A-H2B from Kap114 and Importin-9

Previously we showed that the nuclear import receptor Importin-9 wraps around the H2A-H2B core to chaperone and transport it from the cytoplasm to the nucleus. However, unlike most nuclear import systems where RanGTP dissociates cargoes from their importins, RanGTP binds stably to the Importin-9*H2A-H2B complex and formation of the ternary RanGTP*Importin-9*H2A-H2B complex facilitates H2A-H2B release to the assembling nucleosome. It was unclear how RanGTP and the cargo H2A-H2B can bind simultaneously to an importin, and how interactions of the three components position H2A-H2B for nucleosome assembly. Here we show cryo-EM structures of Importin-9*RanGTP and of its yeast homolog Kap114, including Kap114*RanGTP, Kap114*H2A-H2B, and RanGTP*Kap114*H2A-H2B to explain how the conserved Kap114 binds H2A-H2B and RanGTP simultaneously and how the GTPase primes histone transfer to the nucleosome. In the ternary complex, RanGTP binds to the N-terminal repeats of Kap114 in the same manner as in the Kap114/Importin-9*RanGTP complex, and H2A-H2B binds via its acidic patch to the Kap114 C-terminal repeats much like in the Kap114/Importin-9*H2A-H2B complex. Ran binds to a different conformation of Kap114 in the ternary RanGTP*Kap114*H2A-H2B complex. Here, Kap114 no longer contacts the H2A-H2B surface proximal to the H2A docking domain that drives nucleosome assembly, positioning it for transfer to the assembling nucleosome. Significance StatementHistones and their chaperone networks are typically conserved in eukaryotes. The yeast importin Kap114 and its human homolog Importin-9 share low sequence identity, but both are primary nuclear import receptors for the core histone heterodimer H2A-H2B. Cryo-EM structures of Kap114*H2A-H2B, Kap114*RanGTP and Importin-9*RanGTP complexes show homologous structure and function for Kap114 and Importin-9. In the nucleus, RanGTP binding to Kap114/Imp9*H2A-H2B does not release H2A-H2B, but RanGTP binds to form an atypical ternary complex. Structure of the ternary RanGTP*Kap114*H2A-H2B complex explains how the GTPase and cargo bind simultaneously to Kap114 and how the presence of Ran in the complex primes H2A-H2B transfer to assembling nucleosomes.

biochemistry↗

bioRxiv2021

The spatial transcriptomic landscape of the healing intestine following damage

The intestinal barrier is composed of a complex cell network defining highly compartmentalized and specialized structures. Here, we use spatial transcriptomics (ST) to define how the transcriptomic landscape is spatially organized in the steady state and healing murine colon. At steady state conditions, we demonstrate a previously unappreciated molecular regionalization of the colon, which dramatically changes during mucosal healing. Here, we identified spatially-organized transcriptional programs defining compartmentalized mucosal healing, and regions with dominant wired pathways. Furthermore, we showed that decreased p53 activation defined areas with increased presence of proliferating epithelial stem cells. Finally, we used our resource to map transcriptomics modules associated with human diseases demonstrating that ST can be used to inform clinical practice. Overall, we provide a publicly available resource defining principles of transcriptomic regionalization of the colon during mucosal healing and a framework to develop and progress further hypotheses.

systems biology↗

bioRxiv2018

Chromatin compaction states, nuclear shape fluctuations and auxeticity: A biophysical interpretation of the epigenetic landscape of stem cells

When embryonic stem cells differentiate, the mechanical properties of their nuclei evolve en route to their terminal state. Measurements of the deformability of cell nuclei in the transitional state that intervenes between the embryonic stem cell state and the differentiation primed state of mouse stem cells, indicate that such nuclei are auxetic i.e. have a negative Poissons ratio. We show, using a theoretical model, how this unusual mechanical behaviour results from the coupling between chromatin compaction states and nuclear shape. Our biophysical approach, which treats chromatin as an active polymer system whose mechanics is modulated by nucleosome binding and unbinding, reproduces experimental results while providing new predictions. We discuss ways of testing these predictions. Our model suggests a biophysical interpretation of the epigenetic landscape of stem cells.

biophysics↗