Biology subjects

Motiani, R. K.

Publications and source records attributed to Motiani, R. K..

12 recordsLinked to original sources

bioRxiv2026

The IFN-γ-inducible GTPase IIGP1 regulates mitochondrial remodeling and immunometabolic reprogramming during inflammatory macrophage polarization

Macrophages are key effector cells of the innate immune system that undergo dynamic polarization into functionally distinct phenotypic states, ranging from classically activated inflammatory (M1) to alternatively activated (M2) macrophages. Inflammatory macrophage polarization requires coordinated integration of cytokine signaling, metabolic reprogramming, and mitochondrial remodeling. However, the molecular effectors that control inflammatory signaling for mitochondrial metabolism and remodeling remain poorly defined. Using quantitative mass spectrometry-based proteomics, we identified Interferon-inducible GTPase 1 (IIGP1) as one of the most robustly induced proteins during inflammatory macrophage polarization. Its expression is governed by the IFN-{gamma}-JAK-STAT1 signaling axis and is crucial for this polarization process. IIGP1-deficient macrophages exhibited significantly impaired iNOS induction, nitric oxide production, pro-inflammatory gene expression, co-stimulatory surface marker expression, and morphological remodeling. Further, IIGP1 associates with mitochondria and its absence impairs key characteristics of inflammatory macrophages, including glycolytic metabolic reprogramming, mitochondrial redox adaptation, membrane potential regulation, and structural remodeling. Consistent with these defects, IIGP1-deficient macrophages displayed impaired intracellular killing of Escherichia coli and Salmonella Typhimurium. Collectively, these results point to IIGP1 as an IFN-{gamma}-inducible effector that facilitates efficient inflammatory macrophage polarization and antibacterial immunity by linking inflammatory signaling to mitochondrial homeostasis and immunometabolic reprogramming.

immunology↗

bioRxiv2026

Alcohol-Evoked Calcium Signaling Drives Distinct Responses in Zebrafish Hepatocytes and Pancreatic Acinar Cells

Alcohol exposure perturbs intracellular calcium (Ca2+) homeostasis in digestive organs, yet whether common or organ-specific mechanisms coordinate this response remains unclear. Using an acute ethanol paradigm in zebrafish, single-cell transcriptomics revealed broad up-regulation of Ca2+-signaling genes in hepatocytes and pancreatic acinar cells. In vivo Ca2+ buffering with SpiCee, a genetically encoded chelator, demonstrated a shared requirement for Ca2+ flux: in hepatocytes, lineage-restricted buffering was associated with pronounced cytoplasmic vacuolation composed of lipid-negative vesicles, consistent with stalled lysosomes or autophagosomes; in pancreatic acinar cells, it was associated with accumulation of aggregated/misfolded protein. Mechanistic experiments using pharmacological inhibitors implicated distinct molecular contributors in each tissue. In hepatocytes, inhibition of Pikfyve or its downstream effector, the lysosomal Ca2+ channel TRPML1, phenocopied Ca2+ buffering. While, in acinar cells, Pick1 inhibition produced analogous associations. These data position Pikfyve and Pick1 as organ-specific components linked to the Ca2+-coupled alcohol response. Notably, pharmacologic activation of TRPML1 in hepatocytes recapitulated alcohol-like Ca2+ dynamics but increased macrophage recruitment and cell death, indicating that Ca2+ signaling is required for the alcohol response yet can be detrimental when amplified. Together, our results support a model in which alcohol elicits a shared Ca2+ dynamics across liver and pancreas, modulated by tissue-specific molecular nodes.

cell biology↗

bioRxiv2026

TRPM2 couples cell-autonomous type-I interferon signaling to pigmentation homeostasis

Transient Receptor Potential Melastatin 2 (TRPM2), a Ca{superscript 2}-permeable cation channel, regulates innate and adaptive immunity and has recently been implicated in vitiligo, an autoimmune pigmentary disorder. However, whether TRPM2 exerts cell-autonomous immunoregulatory functions and how such signaling intersects with pigmentation remain unknown. Here, we reveal an unexpected role for TRPM2 as an intrinsic suppressor of pigmentation through type-I interferon (IFN) signaling in melanocytes. Pharmacological inhibition, genetic silencing, and gain-of-function approaches demonstrate that TRPM2 negatively regulates melanogenesis in vitro. Notably, TRPM2-deficient zebrafish and TRPM2-/- mice exhibit enhanced pigmentation in vivo, establishing physiological relevance. Transcriptomic profiling uncovers autonomous activation of the type-I-IFN pathway upon TRPM2 loss, leading to induction of interferon-stimulated gene 15 (ISG15). Mechanistically, ISG15 attenuates global ubiquitination and stabilizes microphthalmia-associated transcription factor (MITF), the master regulator of melanogenesis, thereby promoting pigmentation. Collectively, our findings define a previously unrecognized TRPM2-type-I-IFN-ISG15-MITF signaling axis that functionally integrates cell-autonomous immune surveillance pathways with pigmentary control. Further, it provides a conceptual framework linking type-I interferon signaling to pigmentation homeostasis and pigmentary disorders. Key highlights of the studyO_LITRPM2 negatively regulates pigmentation in vitro (mouse and primary human cells) and in vivo (zebrafish and mice). C_LIO_LIUnbiased RNA-sequencing identifies ISG15 as a positive regulator of melanogenesis downstream of TRPM2 silencing. C_LIO_LITRPM2 knockdown generates a cell autonomous type-I-IFN response in melanocytes that induces ISG15 expression. C_LIO_LIISG15 antagonizes global ubiquitination and regulates stability of MITF, the master regulator of pigmentation. C_LI

cell biology↗

bioRxiv2025

Orai3 orchestrates gemcitabine resistance in pancreatic cancer via NFATc1-SLIT3 axis

Pancreatic Cancer (PC) is one of the most aggressive cancers and is associated with poor prognosis. Gemcitabine is the first-line chemotherapy for PC. While gemcitabine-based regimes offer survival benefits, the acquired gemcitabine resistance leads to reoccurrence, metastasis and the long-term survival rate remains dismal. Although there is substantial clinical evidence for gemcitabine resistance, the cellular and molecular mechanisms driving gemcitabine resistance remain largely unappreciated. Here, we reveal that Orai3, a Ca2+ selective channel, is a crucial driver of gemcitabine resistance. We demonstrate that Orai3 is upregulated in gemcitabine-resistant PC cells. Orai3 silencing in these cells decreases proliferation, induces cell cycle arrest, enhances apoptosis, and moderates stemness characteristics. Notably, studies in zebrafish model corroborate the significance of Orai3 in gemcitabine resistance in vivo. Mechanistically, our unbiased RNA-seq analysis coupled with robust functional studies show that SLIT3 works downstream of Orai3 to drive gemcitabine resistance. Finally, we report that NFATc1 transcription factor bridges Orai3 to SLIT3 transcription. Taken together, this study identifies Orai3 as a key orchestrator of gemcitabine resistance and uncovers a unique Orai3-NFATc1-SLIT3 signaling module that drives chemoresistance. Hence, this work reveals Orai3 as a promising target for synergistic therapeutic approach to combat chemoresistance. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=197 HEIGHT=200 SRC="FIGDIR/small/671673v1_ufig1.gif" ALT="Figure 1"> View larger version (32K): org.highwire.dtl.DTLVardef@16fea2org.highwire.dtl.DTLVardef@1a77a3forg.highwire.dtl.DTLVardef@4aef28org.highwire.dtl.DTLVardef@19b62a3_HPS_FORMAT_FIGEXP M_FIG C_FIG Highlights{Rightarrow} Orai3 is upregulated in gemcitabine-resistant pancreatic cancer cells {Rightarrow}Orai3 is essential for chemo-sensitivity, migration, invasion and stemness in vitro {Rightarrow}Orai3 regulates metastasis of gemcitabine-resistant pancreatic cancer cells in vivo {Rightarrow}Unbiased RNA-seq reveals that SLIT3 functions downstream of Orai3 to drive gemcitabine resistance {Rightarrow}NFATc1 transcription factor bridges Orai3 to SLIT3 transcription

cancer biology↗

bioRxiv2025

SNHG10 promote tumorigenesis via miR-150/VEGFA/EGFR/AKT/ERK/mTOR axis and gemcitabine resistance in PDAC

SNHG10 emerged as a key regulator in progression and metastasis of cancers. However, potential of SNHG10 in PDAC tumorigenesis, gemcitabine resistance, and underlying mechanisms remains poorly understood. We observed significant upregulation of SNHG10 in 179 PDAC cases, revealing a positive correlation with clinical stages. Our results showed a significant SNHG10 overexpression in several PDAC cells. Downregulation of SNHG10 significantly decreased the proliferation, clonogenicity, EMT, tumor growth in the xenograft model, and the induction of cell cycle arrest and apoptosis of PDAC cells. Mechanistically, SNHG10 knockdown significantly inhibited the expression of vimentin, N-cadherin, survivin, CDK4, CDK6, cyclin B1, cyclin D1, aurora kinase A, and B, with an increased expression of E-cadherin and p21. RNA Immunoprecipitation data displayed physical interaction among SNHG10, miR-150-5p, and VEGFA in PDAC cells. SNHG10 silencing led to the significant induction of miR-150-5p, which repressed VEGFA expression in PDAC cells. SNHG10 downregulation enhanced gemcitabine sensitivity in PDAC cells. SNHG10 silencing suppressed the phosphorylation of EGFR, AKT, ERK1/2, mTOR, and c-MET pathways. Silencing SNHG10 and its regulated signalling offers a novel prospective therapeutic strategy.

cancer biology↗

bioRxiv2025

IP3R2 mediated inter-organelle Ca2+ signaling orchestrates melanophagy

Organelle dynamics and crosstalk play a critical role in cellular functions thereby regulating physiological processes and pathological conditions. A variety of cellular processes are outcome of a balance between organelle biogenesis and degradation. Pigmentation is one such homeostatic state that is a result of melanosome biogenesis and melanosome degradation. Although melanosome biogenesis is partially understood, the melanosome degradation i.e. melanophagy remains largely unappreciated. Here, we reveal that Inositol 1,4,5-trisphosphate receptor 2 (IP3R2) is a negative regulator of melanophagy. In this study, we developed two de novo ratio metric imaging probes to study melanophagy in live-cells. Using these probes, biochemical assays, ultrastructural studies, confocal microscopy, molecular analyses and calcium imaging; we demonstrate that IP3R2, but not IP3R1 or IP3R3, keeps melanophagy in check. In vivo studies in zebrafish model system further substantiate IP3R2s functional relevance in pigmentation. Mechanistically, IP3R2 silencing decreases mitochondrial Ca2+ uptake, augments ADP/ATP ratio and thereby activates melanophagy. Simultaneously, IP3R2 knockdown increases ER-lysosome proximity, enhances lysosomal Ca2+ levels and decreases lysosomal pH. This in turn activates lysosomal TRPML1 channel and stimulates nuclear translocation of TFEB transcription factor, which facilitates transcription of key autophagy and two known melanophagy drivers. Taken together, we uncover that IP3R2-mediated Ca2+ signaling across organelles is a critical determinant of melanophagy and thereby skin pigmentation. Hence, this signaling cascade offers potential therapeutic prospects for the management of pigmentary disorders and skin malignancies. Highlights[tpltrtarr] IP3R2, but not IP3R1 and IP3R3, is a critical positive regulator of melanogenesis in vitro and in vivo. [tpltrtarr]Generation and validation of two de novo ratio-metric live cells imaging probes reveal crucial role of IP3R2 in melanophagy. [tpltrtarr]IP3R2 knockdown decreases mitochondrial Ca2+ uptake, augments ADP/ATP ratio and thereby activates melanophagy via AMPK-ULK1 pathway. [tpltrtarr]IP3R2 silencing enhances ER-lysosomal proximity, elevates lysosomal Ca2+ levels and reduces lysosomal pH. [tpltrtarr]IP3R2 knockdown stimulates lysosomal TRPML1 channel activity thereby facilitating nuclear translocation of TFEB transcription factor. [tpltrtarr]TFEB transcriptionally upregulates genes involved in the melanophagy process, leading to enhanced degradation of melanosomes and decreased pigmentation.

cell biology↗

bioRxiv2024

NFAT2 drives both Orai3 transcription and protein degradation by harnessing the differences in epigenetic landscape

The functional significance of any protein in physiological processes and pathological conditions is largely dependent on its expression profile. Therefore, nature has evolved several autonomous mechanisms to regulate protein expression such as transcription, translation, post-translational modifications and epigenetic changes. These processes are typically controlled by distinct molecular players with no overlapping roles. Here, we reveal that same transcription factor, NFAT2 regulates both transcription and lysosomal degradation of Orai3 oncochannel in a context dependent manner. We demonstrate that NFAT2 drives Orai3 transcription and thereby increases Orai3 levels in non-metastatic cancerous cells. While in invasive and metastatic cancerous cells, NFAT2 induces Orai3 lysosomal degradation by transcriptionally enhancing the levels of MARCH8 E3 ubiquitin ligase. Our biochemical and super-resolution microscopy data show that MARCH8 physically interacts with Orai3 eventually resulting in its degradation. Mechanistically, the dichotomy in regulation of Orai3 expression emerges from the differences in the epigenetic landscape of MARCH8. We uncover that the MARCH8 promoter is highly methylated in non-metastatic cancerous cells and hence NFAT2 does not induce MARCH8 mediated Orai3 degradation in these cells. Importantly, we demonstrate that MARCH8 restricts pancreatic cancer metastasis by targeting Orai3 degradation thereby highlighting pathophysiological importance of this signaling module. Taken together, we report a unique and clinically relevant scenario wherein nature has commissioned the same transcription factor to both enhance and curtail the expression of a target protein. Highlights[tpltrtarr] NFAT2 transcriptionally upregulates Orai3 Ca2+ channel in non-metastatic cells [tpltrtarr]NFAT2 induces Orai3 lysosomal degradation via MARCH8 E3 ubiquitin ligase in metastatic cells [tpltrtarr]The dichotomy in NFAT2s function is an outcome of differences in the methylation status of MARCH8 promoter [tpltrtarr]MARCH8 inhibits pancreatic cancer metastasis by driving Orai3 degradation Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=199 SRC="FIGDIR/small/628081v2_ufig1.gif" ALT="Figure 1"> View larger version (48K): org.highwire.dtl.DTLVardef@1292480org.highwire.dtl.DTLVardef@1b5b61org.highwire.dtl.DTLVardef@c4483org.highwire.dtl.DTLVardef@16607f8_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2024

LncRNA TAAL is a Modulator of Tie1-Mediated Vascular Function in Diabetic Retinopathy

Diabetic retinopathy (DR), a leading cause of vision impairment and blindness, is characterized by abnormal retinal vascular changes due to chronic hyperglycemia. The Tie-1 signaling pathway, essential for vascular growth and remodeling, has emerged as a key therapeutic target, though its molecular mechanisms and interactome remain largely unclear. Through a protein-centric approach, we identified a novel lncRNA and named it Tie1-associated angiogenic lncRNA (TAAL). TAAL lncRNA regulates endothelial cell migration, proliferation, tube formation, and permeability by modulating ER-calcium homeostasis and cytoskeleton dynamics. In zebrafish, taal modulation led to angiogenic defects, which were rescued by human TAAL orthologue. Our molecular studies further revealed that TAAL negatively regulates Tie1 protein via ubiquitin-mediated degradation. Notably, TAAL expression is upregulated in the blood of DR patients and downregulated in endothelial DR cell models. Overexpression of TAAL restored endothelial permeability and VE-cadherin surface expression. These findings establish TAAL as a novel regulator of Tie1 protein turnover, with potential therapeutic implications for diabetic retinopathy.

genetics↗

bioRxiv2024

Remodeling Ca2+ dynamics by targeting a promising E-box containing G-quadruplex at ORAI1 promoter in triple-negative breast cancer.

ORAI1 is an intrinsic component of store-operated calcium entry (SOCE) that strictly regulates Ca2+ influx in most non-excitable cells. ORAI1 has been extensively studied to have been overexpressed in various cancer phenotypes, and its signal transduction has been associated with oncotherapy resistance. There is extensive proteomic interaction of ORAI1 with other channels and effectors, resulting in various altered phenotypes. However, the transcription regulation of this gene is not well understood. We have found a putative G-quadruplex (G4) motif, ORAI1-Pu, in the upstream promoter region of the gene, having regulatory functions. High-resolution 3-D NMR structure elucidation suggests that ORAI1-Pu is a stable parallel-stranded G4, having an unusual 8-nt loop imparting dynamics without affecting the structural stability. The protruded loop further houses an E-box motif that provides a docking site for transcription factors like Zeb1. The G4 structure was also endogenously observed using Chromatin Immunoprecipitation (ChIP) with anti-G4 antibody (BG4) in the MDA-MB-231 cell line overexpressing ORAI1. Ligand-mediated stabilization suggested that the stabilized G4 represses transcription in cancer cell line MDA-MB-231. Downregulation of transcription further cascaded down to a decrease in Ca2+ entry by the SOCE pathway, as observed by Fura-2 confocal Ca2+ imaging.

biochemistry↗

bioRxiv2023

Telomeres control regulation of the human Telomerase (hTERT) gene through non-telomeric TRF2 and independent of Telomere looping

The function of the human telomerase reverse transcriptase (hTERT) in the synthesis and maintenance of chromosome ends, or telomeres, is widely understood. Whether and how telomeres, on the other hand, influence hTERT regulation is relatively less studied. We found hTERT was transcriptionally altered depending on telomere length (TL). This resulted from TL-dependent binding of TRF2 between telomeres and the hTERT promoter. hTERT promoter-bound TRF2 was non-telomeric and did not involve the looping of telomeres to the hTERT promoter. Cell lines from different tissue types (fibrosarcoma (HT1080), colon cancer (HCT116), and breast cancer (MDA-MB-231), engineered for either telomere elongation/shortening gave increase/decrease in hTERT, respectively. Mechanistically, we show hTERT promoter-bound non-telomeric TRF2 recruits the canonical PRC2-complex inducing repressor histone H3K27-trimethylation in a TL-dependent fashion. This was further supported by TL-dependent promoter activity from an exogenously inserted hTERT reporter. Increase in TL over days followed by gradual decline, resulted in activation followed by repression of hTERT in a concerted manner, further implicating TL as a key factor for hTERT regulation. Notably on reprogramming primary fibroblasts to induced pluripotent stem cells (iPSCs), TRF2 loss from the hTERT promoter was evident along with telomere elongation and hTERT upregulation. Conversely, on telomere shortening in iPSCs, hTERT promoter-bound TRF2 was restored with marked reduction in hTERT further supporting the causal role of TL in hTERT transcription. Mechanisms of tight control of hTERT by TL shown here are likely to have major implications in telomere-related physiologies, particularly, cancer, ageing and pluripotency. TeaserTelomere length controls hTERT expression by modulating TRF2 distribution and PRC2-mediated repression, highlighting a self-regulatory mechanism in cancer.

cancer biology↗

bioRxiv2023

Mitochondrial calcium signaling mediated transcriptional regulation of keratin filaments is a critical determinant of melanogenesis

Mitochondria are versatile organelles that regulate several physiological functions. Many mitochondria-controlled processes are driven by mitochondrial Ca2+ signaling. However, role of mitochondrial Ca2+ signaling in melanosome biology remains unknown. Here, we show that pigmentation requires mitochondrial Ca2+ uptake. In vitro gain and loss of function studies demonstrated that Mitochondrial Ca2+ Uniporter (MCU) is crucial for melanogenesis while the MCU rheostats, MCUb and MICU1 negatively control melanogenesis. Zebrafish and mouse models showed that MCU plays a vital role in pigmentation in vivo. Mechanistically, MCU controls activation of transcription factor NFAT2 to induce expression of three keratins (keratin 5, 7 and 8), which we report as positive regulators of melanogenesis. Interestingly, keratin 5 in turn modulates mitochondrial Ca2+ uptake thereby this signaling module acts as a negative feedback loop that fine-tunes both mitochondrial Ca2+ signaling and melanogenesis. Mitoxantrone, an FDA approved drug that inhibits MCU, decreases physiological melanogenesis. Collectively, our data demonstrates a critical role for mitochondrial Ca2+ signaling in vertebrate pigmentation and reveal the therapeutic potential of targeting MCU for clinical management of pigmentary disorders. Given the centrality of mitochondrial Ca2+ signaling and keratin filaments in cellular physiology, this feedback loop may be functional in a variety of other pathophysiological conditions. HighlightsO_LIMCU complex mediated mitochondrial Ca2+ uptake is a novel regulator of vertebrate pigmentation C_LIO_LIKeratin filaments bridge mitochondrial Ca2+ signaling to melanosome biogenesis and maturation C_LIO_LITranscription factor NFAT2 connects mitochondrial Ca2+ dynamics to keratins expression C_LIO_LIMCU-NFAT2-Keratin 5 signaling module generates a negative feedback loop to maintain mitochondrial Ca2+ homeostasis and to ensure optimal melanogenesis C_LIO_LIInhibiting MCU with mitoxantrone, an FDA approved drug, leads to reduction in physiological pigmentation C_LI

cell biology↗

bioRxiv2023

Methotrimeprazine exerts antiviral and neuroprotective effects in Japanese encephalitis virus infection through activation of adaptive ER stress and autophagy

Japanese encephalitis virus (JEV) is the leading global cause of virus-induced encephalitis. Its pathogenesis is driven by a combination of neuronal cell death and neuroinflammation. We hypothesized that pharmacological upregulation of autophagy could exert a neuroprotective antiviral effect, and tested a panel of forty-two FDA-approved drugs that were shown to induce autophagy. Four drugs were tested in the JE mouse model based on in vitro protective effects on neuronal cell death, inhibition of viral replication, and anti-inflammatory effects in microglial cells. The antipsychotic phenothiazines Methotrimeprazine (MTP) and Trifluoperazine (TFP) showed a significant survival benefit with reduced virus titers in the brain, prevention of blood-brain barrier (BBB) breach, and inhibition of neuroinflammation. Both drugs were potent mTOR-independent autophagy flux inducers. Mechanistically MTP inhibited SERCA channel functioning, thereby resulting in rise in cytosolic calcium levels, and induction of a unique adaptive ER stress response. In virus infected drug treated cells, there was a strong transcriptional downregulation of type I interferon and interferon-stimulated genes and upregulation of cholesterol metabolic pathway genes. The drugs exerted an autophagy-dependent antiviral effect at the level of JEV protein translation/replication complex formation in diverse cell types. Inhibition of inflammatory cytokine/chemokine release from mouse microglial cells was partly autophagy-dependent. Our study suggests that MTP exerts a combined antiviral and anti-inflammatory effect in JEV infection, and has therapeutic potential to be repurposed for JE treatment.

microbiology↗