Biology subjects

Saha, T.

Publications and source records attributed to Saha, T..

11 recordsLinked to original sources

bioRxiv2026

Retrieval uncertainty drives rapid consolidation of confabulated memories in retrosplenial cortex

Learning to make associations in the absence of direct reinforcement, based on previous memories of reward or punishment, is an evolutionary trait quintessential for survival. Little is known about if such higher order associations (HOA) can lead to fabrication of new memories for unreal events. We probed if uncertainty during retrieval of old memory representations can trigger such confabulation. Using chemogenetics, in vivo imaging, and behavior we show that uncertainty in memory perception can not only transfer conditioning to hitherto unexperienced novel context but also can get rapidly consolidated. We find while acquisition of HOA require dorsal-CA1 parvalbumin interneuron activity in intact hippocampus, retrieval requires retrosplenial cortex (RSc). We develop an automated Large-scale Brainwide Correlated Activity Mapping (LaBCAM) method to identify functional connectivity changes across different brain regions in such scenarios. LaBCAM finds that only uncertain remote retrieval engages canonical fear circuitry while regular retrieval does not. Longitudinal imaging of dendritic spines in RSc, a region involved in detection and context-dependent conflict resolution, reveals differential reorganization of spines during acquisition and rapid systems consolidation of HOA. Interestingly, we find HOA is implicated early in AD mice model (APP/PS1) and also in aged animals.

neuroscience↗

bioRxiv2026

Actin-dependent mechanotransduction controls nucleocytoplasmic partitioning of DNMT3a through ERK1/2 signaling during cutaneous wound healing

Cutaneous wound healing is a multifaceted physiological process that requires a cell state transition from homeostasis to tissue repair. An important contributor to this reprogramming is the wound-induced mechanical cues that are perceived by epidermal keratinocytes. Previously we identified that the nuclear translocation of the de novo DNA methyltransferase 3A (DNMT3a) upon wounding as an important regulator of this cell-state transition. However, the molecular mechanisms linking mechanotransduction to epigenetic regulation remain incompletely understood. Here we show that under homeostasis, active ERK1/2 phosphorylates DNMT3a, intramolecularly masking its nuclear localization signal (NLS), resulting in its cytoplasmic sequestration. Upon wounding actin cytoskeleton remodeling leads to the downregulation of the Extracellular Signal-Related Kinase 1/2 (ERK 1/2) pathway. Wound-induced ERK1/2 inactivation unmasks the NLS, enabling DNMT3a nuclear translocation. Collectively, these findings define a mechanotransduction-driven signaling axis linking cytoskeletal dynamics to epigenetic regulation and confirms an active role for differentiated keratinocytes in initiating early wound repair programs.

cell biology↗

bioRxiv2026

A strategically designed homogeneous antibody-drug conjugate improves safety and therapeutic index in renal cell carcinoma

Renal cell carcinoma (RCC) is the most common form of kidney cancer. It is also one of deadliest cancers, with a 5-year survival rate of less than 20% in advanced cancer patients with distant metastasis. Current treatments rely on targeted therapies, such as sunitinib, which are limited in eHicacy. Recently, antibody drug conjugates (ADCs) have emerged as a promising treatment modality by delivering cytotoxic payloads specifically to cancer cells. Here, we describe the engineering of a novel ADC (LT-025), where the antibody targets the kidney injury molecule (KIM) -1 receptor over-expressed on RCC cells to deliver a toxic maytansinoid (DM1) payload. Unlike prior attempts to engineer a KIM-1 targeted ADC that were limited by a heterogenous product, here we used a microbial transglutaminase (MTGase)-based conjugation strategy, which achieved a site-specific conjugation of the drug-linker to the antibody, resulting in a homogenous ADC with a drug-to-antibody ratio (DAR) of 2. The ADC exhibited prolonged stability, excellent antigen-binding capability, and KIM-1 expression-dependent cellular internalization and cytotoxicity in RCC, including in sunitinib-resistant RCC. Excitingly, LT-025 was well tolerated in vivo, and combining LT-025 and sunitinib exhibited a synergistic antitumor eHicacy in a RCC mouse model. Combining an ADC with a targeted therapeutic could emerge as a paradigm shift in the management of advanced RCC.

cancer biology↗

bioRxiv2025

Mindin-α-M-Integrin endocytosis activates STAT3 to maintain keratinocyte stemness

Signalling networks rely not only on specific protein interactions but also on the subcellular locales at which these occur. Such spatial control is fundamental to cellular behaviour in both physiological and pathological scenarios. In studying keratinocyte stemness, we previously identified a Mindin-M{beta}2 (CD11b/CD18) interaction that triggers STAT3 activation. Here, we delineate the mechanism underlying this response and show that the F-Spondin domain of Mindin constitutes the minimal integrin-binding module required to initiate downstream signalling. F-Spondin binding to the integrin at the plasma membrane does not elicit the full activation state of the integrin. Instead, it promotes Src-kinase dependent endocytosis of the integrin receptor to the early endosomes. Analysis of integrin conformational dynamics reveals that this compartmental shift is essential as the acidic environment of early endosomes drives the transition into a signalling-competent state. This mechanism extends to pathological contexts, as we demonstrate a requirement for endocytosis in activating STAT3 signalling and preserving stem-like properties in a cancer stem cell model. Together, these findings highlight a previously unrecognized layer of spatial control in integrin signalling, confirming endosomal trafficking as a critical determinant of stem cell behaviour and offering new conceptual and therapeutic opportunities across regenerative biology and cancer.

cell biology↗

bioRxiv2025

Discovery and therapeutic exploitation of Master Regulatory miRNAs in Glioblastoma

Glioblastoma is a fatal primary malignant brain tumor, with an average survival of only 15 months despite surgical resection, chemotherapy, and radiation therapy. Due to the concurrent deregulation of numerous genes in glioblastoma, molecular monotherapies have not improved clinical outcomes. Evidence suggests that effectively targeting multiple deregulated molecules is essential for better therapies; however, this is limited by the lack of suitable drugs and the increased toxicity of combination therapies. To address this, we hypothesized that miRNAs, small gene-regulatory RNAs that suppress multiple target genes via sequence complementarity, could be developed to inhibit multiple deregulated genes simultaneously, leading to more effective treatments. We identified master regulatory miRNAs--those that target several deregulated genes in glioblastoma--using PAR-CLIP screenings in glioblastoma cells and analyzed TCGA tumor data to find which targets were deregulated. An algorithm ranked these targets based on their significance in glioblastoma malignancy. We selected two tumor suppressor master regulatory miRNAs, miR-340 and miR-382, and one oncogenic miRNA, miR-17. Validation showed that these miRNAs target critical glioblastoma pathways and significantly inhibit cell growth, survival, invasion, and tumor growth in vivo. We developed an innovative therapeutic delivery approach using Brain Penetrating Nanoparticles in combination with MRI-guided focused ultrasound and microbubbles, resulting in reduced tumor volume and extended survival in glioblastoma-bearing mice. This strategy offers a promising pathway for translating miRNA-based therapies into clinical trials for glioblastoma and other cancers. One Sentence SummaryWe developed and used new computational, experimental, and therapeutic approaches to identify and therapeutically deliver master regulatory miRNAs to inhibit the growth of glioblastoma, the most common and deadly primary brain tumor.

cancer biology↗

bioRxiv2025

Bioengineered visible polymeric mesh to enhance urogynaecological health

Pelvic floor disorders, including pelvic organ prolapse and stress urinary incontinence, are prevalent health concerns, affecting approximately 50% of females over their lifetime, with about 75% of women over 65 years of age being impacted. Traditional surgical interventions, such as transvaginal mesh implants, have led to numerous complications, resulting in their prohibition in several countries. This study introduces an innovative composite mesh designed to mitigate these issues by combining polymethylmethacrylate and thermoplastic polyurethane, further enhanced with iodine-doped carbon nanoparticles to enable visibility via medical imaging. The mesh is coated with 2-methacryloyloxyethyl phosphorylcholine polymer to prevent protein adsorption and promote tissue regeneration. In vitro studies showed high cell viability and low protein adsorption, indicating excellent biocompatibility. Implantation of mesh (with or without iodine) in mice revealed no adverse effects on overall animal health. Mouse spleen weight (an indicator of inflammation) was similar between groups; however, levels of some cytokines (i.e., IL-10, IL-17A and GM-CSF) were elevated following implantation of iodinated mesh in mice suggesting that further refinement of the composite mesh is required. Analysis of the fecal microbiome, which is correlated with physiological states, showed that sham and iodinated mesh implant groups maintained consistent microbial profiles with stable diversity (richness and evenness) measures over time. In contrast, the non-iodinated mesh group exhibited decreased species richness post mesh implantation, likely due to a distinct starting microbiome composition prior to implantation. This research is envisaged to contribute to a safer and more effective solution for treating pelvic floor disorders, providing non-invasive post-implantation monitoring and enhanced mechanical compatibility of surgical mesh with native tissue. Our findings demonstrate that this composite mesh possesses mechanical properties that closely mimic human tissue, ensuring biocompatibility, strength, and flexibility without stimulating significant inflammatory or foreign body responses.

bioengineering↗

bioRxiv2024

Mechanical Strain Activates Planar Cell Polarity Signaling to Coordinate Vascular Cell Dynamics

Mechanical stimuli, particularly laminar blood flow, play a crucial role in shaping the vascular system. Changes in the rate of blood flow manifest in altered shear stress, which activates signaling cascades that drive vascular remodeling. Consistently, dysregulation of the endothelial response to fluid shear forces and aberrant flow patterns both lead to pathological conditions, including impaired blood vessel development and atherosclerosis. Despite its importance, the mechanisms driving the coordinated cell behavior underlying vascular remodeling are not fully understood. Combining classical cell biological approaches with advanced image analysis, mathematical modeling, biomimetic strategies, and in vivo studies, we identify the planar cell polarity (PCP) protein Vangl1 as an enforcer of flow-dependent cell dynamics in the vascular system. We demonstrate that shear stress triggers the relocation of Vangl1 from an internal reservoir to the plasma membrane at the initiation of cell remodeling. Membrane enrichment of Vangl1 is mediated by a Coronin1C-dependent shift in the equilibrium between endo- and exocytosis and results in the spatial reorganization of another essential PCP protein, Frizzled6 (Fzd6). The resulting mutual exclusion of the core PCP proteins Fzd6 and Vangl1 augments differential junctional and cytoskeletal dynamics along the flow axis. Loss of Vangl1 limits the ability of endothelial cells to respond to shear forces in a coordinated fashion, resulting in irregular cell alignment along the flow direction and erroneous vessel sprouting. Together, these studies introduce core PCP signaling as a determinant of collective cell dynamics and organization of the vascular system.

cell biology↗

bioRxiv2023

Designer grass pea for transgene-free minimal neurotoxin-containing seeds with CRISPR-Cas9

Grass pea seeds are consumed as food in several South Asian and Sub-Saharan African nations. However, the presence of the neurotoxic compound N-oxalyl-L-diamino propionic acid ({beta}-ODAP) has restricted its cultivation. Although various cultivars with low {beta}-ODAP levels have been developed, their cultivation is still limited due to the risk of neurolathyrism from long-term grass pea seed ingestion. In this study, we employed the CRISPR-Cas9-mediated gene editing technique to generate grass pea seeds with zero or minimal {beta}-ODAP levels. We targeted the BAHD-AT3 gene that encodes a key enzyme in the {beta}-ODAP biosynthesis pathway. We developed bahd-at3 knock-out lines using three gRNAs targeting different regions of this gene and characterized them. Cas9-free independent lines from each event carrying the desired on-target mutation were selected and backcrossed twice with the wild-type to eliminate any off-target mutation present therein. Various agronomical parameters were analyzed from the backcrossed mutant lines and they displayed no phenotypic abnormalities. Interestingly, the seed {beta}-ODAP content ranged between 0.001 % - 0.002 % of dry weight which is 99 % lower than the wild-type. Together, our study reports the development of transgene-free, genome-edited grass peas with insignificant levels of {beta}-ODAP in seeds for safer food in the future.

plant biology↗

bioRxiv2023

Toxic metal uptake by oyster mushrooms grown in sugarcane bagasse

Fungiculture or Mushroom Cultivation is rapidly expanding throughout the world. It is further catalyzed by its high nutritional value and increased market demand. Additionally, the method of cultivation does not require much space and the per unit production is very high compared to other crops. Bagasse is the byproduct left after extracting the juice from sugarcane. It has been speculated that bagasse can be an excellent substrate due to its high carbohydrate and mineral content (Hoa et al., 2015). In the experiment spawns of Oyster Mushroom were inoculated in bagasse collected from industries. Remarkable growth of mushrooms in short period of time was observed. However, the metal toxicity being our primary concern in this experiment, AAS (Atomic Absorption Spectrometry) Test was used to detect the heavy metal ion concentrations in both the bagasse and the fruiting bodies of mushrooms. The concentrations of metals calculated are as follows, in bagasse (mg/kg): Cadmium - 0.015, Arsenic - 8.56, Copper-12.47, Chromium-9.17, Mercury - less than 0.01, and in mushroom (mg/kg): Cd - 0.022, As - 10.12, Cu - 14.29, Cr - 7.59, Hg - less than 0.01. These results indicated biomagnification of metal ion concentration in the mushrooms up taken from bagasse. These metals, in such high concentrations are lethal and causes many diseases in humans and other animals. Hence, from this experiment it can be deduced, that though bagasse is an excellent substrate for mushroom cultivation, the toxicity of metal ions present in it in such high concentrations overshadow the earlier benefits of bagasse as a good substrate. However, it has been proposed that if such metals are removed or their concentrations are lowered then the earlier mentioned benefits of bagasse are restored.

ecology↗

bioRxiv2021

Protection of nascent DNA at stalled replication forks is mediated by phosphorylation of RIF1 intrinsically disordered region

RIF1 is a multifunctional protein that plays key roles in the regulation of DNA processing. During repair of DNA double-strand breaks (DSBs), RIF1 functions in the 53BP1-Shieldin pathway that inhibits resection of DNA ends to modulate the cellular decision on which repair pathway to engage. Under conditions of replication stress, RIF1 protects nascent DNA at stalled replication forks from degradation by the DNA2 nuclease. How these RIF1 activities are regulated at the post-translational level has not yet been elucidated. Here, we identified a cluster of conserved ATM/ATR consensus SQ motifs within the intrinsically disordered region (IDR) of mouse RIF1 that are phosphorylated in proliferating B lymphocytes. We found that phosphorylation of the conserved IDR SQ cluster is dispensable for the inhibition of DSB resection by RIF1, but is essential to counteract DNA2-dependent degradation of nascent DNA at stalled replication forks. Therefore, our study identifies a key molecular switch that enables the genome-protective function of RIF1 during DNA replication stress.

molecular biology↗

bioRxiv2020

Retromer retrieves the Wilson Disease protein, ATP7B from lysosomes in a copper-dependent mode

ATP7B utilizes lysosomal exocytosis to export copper from hepatocytes. We investigated the fate of ATP7B, post-copper export. At high copper ATP7B traffics to lysosomes and upon subsequent copper chelation, returns to Trans Golgi Network. At high copper, ATP7B co-localizes with lysosomal marker, Lamp1 and the core member of retromer complex, Vps35. Knocking down VPS35 did not alter copper-responsive vesicularization of ATP7B; rather upon subsequent copper chelation, ATP7B failed to relocalize to TGN that could be rescued by overexpressing wtVPS35. Using super-resolution microscopy and proximity ligation assays we demonstrate that VPS35 and ATP7B are juxtaposed on the same lysosomal compartment and their interaction is indirect. Utilizing in-cell photoamino acid-based UV-crosslinking and subsequent immunoprecipitation, we detected ATP7B and retromer subunits, VPS35 and VPS26 in a large complex in high copper conditions, hence confirming their interaction. We demonstrate that retromer regulates lysosome to TGN trafficking of the copper transporter ATP7B and it is dependent upon cellular copper level.

cell biology↗