Biology subjects

Chakraborty, A.

Publications and source records attributed to Chakraborty, A..

At least 55 records · Page 3Linked to original sources

bioRxiv2025

Organ-specific rewiring of mitochondrial integrity through COX7A dictates cellular ploidy control

To achieve proper cell and tissue size, cytoplasmic and nuclear growth must be coordinated. Disrupting this coordination causes birth defects and disease. In natures largest cells, nuclear growth occurs through polyploidization (whole-genome-duplication). How the massive nuclear growth of polyploid cells is coordinated with cytoplasmic growth processes such as mitochondrial biogenesis is relatively unclear. Here, focusing on one of natures most commonly polyploid organs-the heart-we uncover cross-talk between cytoplasmic mitochondrial biogenesis/integrity and nuclear growth/polyploidy. From a human-to-fly screen, we uncover novel regulators of cardiomyocyte ploidy, including mitochondrial integrity regulators. In comparing these cardiac hits with a parallel screen in another polyploid tissue, the salivary gland, we discovered two opposing roles for Cytochrome-c-oxidase-subunit-7A (COX7A). While salivary gland COX7A preserves mitochondrial integrity to promote polyploidy and optimal organ growth, cardiac COX7A instead suppresses mitochondrial biogenesis to repress polyploidy and prevent hypertrophic organ growth. Among all electron transport chain genes, only COX7A functions as a cardiac growth repressor. Fly hearts with compromised COX7A show abnormally high cardiac output. Human COX7A1, a mitochondrial-localized protein, similarly represses polyploidy of human iPSC-derived cardiomyocytes. In summary, our human-fly-human approach reveals conserved rewiring of mitochondrial integrity in heart tissue that switches COX7As role from ploidy promotion to repression. Our findings reveal fundamental cross-talk between mitochondrial biogenesis and genome duplication that are critical in growing metazoan tissues.

developmental biology↗

bioRxiv2025

Bridging biology and statistics with hybrid Bayesian experimental design for drug dose-response assays

Dose-response cytotoxicity assays are central to evaluating drug potency, yet selecting concentrations that capture both the full response range and key parameters such as the half-maximal inhibitory concentration (IC50) and Hill slope remains challenging. Practical constraints, including limited replicates, variability across culture conditions, and the trial-and-error nature of current practice, make assays time- and resource-intensive. To address this, we introduce a Bayesian inference framework that quantifies uncertainty, incorporates prior knowledge, and extend it with a Bayesian optimal experimental design (OED) strategy to systematically refine concentration selection. We further propose a hybrid OED approach that integrates information-theoretic design with space-filling principles, aligning statistical rigor with biological intuition. Applied to ER+ breast cancer cells in monoculture and co-culture with stromal cells, this framework revealed differential drug responses while improving information efficiency. More broadly, our results highlight how Bayesian OED can bridge computational and biological perspectives, offering a path toward more efficient, reproducible, and interpretable experimental design in cancer research and beyond. Author summaryWe present a statistical framework that systematically captures uncertainty in drug cytotoxicity assays, which are standard laboratory tests that measure how sensitive cells are to drug treatments. Applying this approach to breast cancer cells grown with and without supportive bone marrow cells, we revealed how the surrounding environment can protect cancer cells from therapy. By guiding the selection of drug concentrations, our framework reduces trial-and-error, thereby saving time and resources. Beyond cancer research, this strategy offers a general way to design more efficient biological experiments, ultimately supporting the development of new treatments.

cancer biology↗

bioRxiv2025

Strain, procedures, and tools for reproducible genetic transformation and genome editing of Spirodela polyrhiza (L.) Schleid.

Duckweeds (Lemnaceae) have excellent potential for fundamental and applied research due to ease of cultivation, small size, and continuous fast clonal growth. However, their usage as model organisms and platforms for biotechnological applications is often limited by the lack of universal genetic manipulation methods necessary for transgene expression, gene editing, and other methods to modify gene expression. To identify suitable strains for genetic manipulation of the giant duckweed, Spirodela polyrhiza, we screened several genotypes for callus induction and regeneration and established genetic transformation. We have identified SP162 to be amenable to Agrobacterium-mediated transformation via tissue culture. The procedure is robust and reproducible across laboratories, allowing stable expression of different reporter genes and selectable markers, enabling CRISPR/Cas9-mediated genome editing. In addition, due to a weak small RNA-based silencing response, S. polyrhiza sustains prolonged periods of transgene activity in transient expression assays. To promote duckweed research and encourage the adoption of S. polyrhiza, we have made SP162 (ID#: 5676) and its genome publicly available and provide here detailed procedures for its cultivation and transformation. Furthermore, we created a web server to explore its genome, retrieve gene sequences, and implemented orthologous gene search and a gRNA design function for diverse CRISPR/Cas-based applications (https://agxu.uni-mainz.de/SP162/).

plant biology↗

bioRxiv2025

The club cell circadian clock regulates temporal patterns in leukocyte trafficking in chronic allergic airways disease

RationaleAsthma displays temporally variable symptoms which worsen overnight, corresponding with a nocturnal increase in airway eosinophils. The molecular clock within the club cell of the bronchial epithelium is a key driver of lung rhythmic processes, however, its role in chronic allergic airways disease (AAD) is not known. Elucidating the role of the club cell clock in regulating rhythmic inflammation in AAD could lead to new therapeutic advances. ObjectivesTo investigate the club cell molecular clock regulation of leukocyte trafficking in chronic AAD. Methodsccsp-bmal1 KO mice (which lack a functional clock in club cells) and littermate control mice underwent a 5-week chronic house dust mite (HDM) model of AAD, following which leukocyte populations and cytokines from blood, lung and airway compartments were quantified in a 24-hour time-course. Airway epithelial cells were cultured and transepithelial electrical resistance measured to explore circadian variability in barrier permeability and impact of pharmacological modulation of the clock. Main ResultsLeukocyte populations accumulate in the blood, lung and airways of HDM exposed mice in a time-of-day dependent manner, with time of peak accumulation dependent on cell type. This temporal gating of leukocyte accumulation is controlled and coordinated by the club cell circadian clock, which also regulates airway barrier integrity. Targeting REVERBa (a component of the molecular circadian clock), was effective at modifying airway barrier permeability achieving reduced transepithelial leukocyte migration. ConclusionsThe club cell clock gates leukocyte trafficking signals and airway barrier integrity by time of day in chronic allergic airway inflammation.

immunology↗

bioRxiv2025

Huntingtin preserves mitochondrial genome integrity in neurons, which is impaired in Huntingtons disease

Huntingtin (HTT) function is enigmatic, as the native protein plays critical roles in neuronal health, while mutant HTT (mHTT), carrying an expanded polyglutamine stretch, triggers neurotoxicity and contributes to the pathogenesis of Huntingtons disease (HD). We recently found that HTT is part of a nuclear transcription-coupled DNA repair (TCR) complex with DNA repair enzymes including polynucleotide-kinase-3-phosphatase (PNKP). This complex resolves DNA lesions during transcription to maintain genome integrity, while in HD, mHTT impairs the activity of this complex, resulting in accumulation of DNA lesions. Using molecular, cellular biology and computational methods, we find that HTT has a role in assembling a functional DNA repair complex in mitochondria. Together with mitochondrial RNA polymerase and transcription factors, HTT resolves mitochondrial DNA lesions to preserve mitochondrial genome integrity and function. Pathogenic mHTT impairs this activity, resulting in persistent DNA lesions and reduced mitochondrial function in HD. Importantly, restoring activity of this complex in a Drosophila HD model through ectopic HTT or PNKP expression significantly improves mitochondrial genome integrity and ameliorates motor deficits. HIGHLIGHTSO_LIHTT organizes a functional, multifactorial mitochondrial DNA repair complex C_LIO_LIMutant HTT impairs the mitochondrial DNA repair complex causing DNA damage accumulation C_LIO_LIHTT-associated repair complex resolves mitochondrial DNA lesions and DNA integrity C_LIO_LIRestoring repair activity in HD flies rescues mitochondrial DNA integrity and motor defects C_LI

neuroscience↗

bioRxiv2025

Treatment Resistant Persister Cells Exploit Macrophage Lipid Metabolism to Sustain Glioblastoma Growth

Glioblastoma (GBM) displays pronounced intratumoral heterogeneity, posing significant challenges to understanding its biology and developing effective treatments. Using spatial multi-omics, in vivo functional assays, and systems-level analysis, we delineate the diverse metabolic and immune architecture of GBM. We identify a lipid-dependent lineage of treatment-resistant persister cells (TRPCs) that engage tumor-associated macrophages (TAMs) in a spatially organized, metabolically specialized crosstalk. TRPCs co-opt CCR2, CSF1R, CD163+ TAMs for lipid scavenging and acquisition, promoting a pro-tumorigenic and immunosuppressive microenvironment. This cooperative axis is critically dependent on lipid chaperones like FABPs, whose targeting disrupts TAM recruitment, remodels immune composition, and suppresses tumor growth. Retrospective clinical analyses reveal that elevated TRPC-associated transcriptome may serve as stratification criteria to identify patients benefiting from lipid-lowering therapies like statins. Our findings uncover a targetable immunometabolic circuit between TRPCs and TAMs and support the development of precision therapies that disrupt lipid-fueled tumor-immune cooperation in GBM. In briefTreatment-resistant persister cells (TRPCs) in glioblastoma spatially engage TAMs to facilitate lipid transfer, thereby sustaining tumor growth and promoting immune evasion. Targeting this TRPC-TAM metabolic axis reprograms the immunosuppressive microenvironment and improves therapeutic outcomes, revealing a clinically actionable metabolic vulnerability with potential for precision immune-metabolic interventions in GBM. HighlightsO_LIGBM exhibits spatially resolved heterogeneity revealing correlative metabolic and immune micro-niches C_LIO_LITRPC lineage promotes a pro-tumorigenic and immunosuppressive microenvironment by recruiting lipid-specialized TAMs C_LIO_LITRPC lineage hijacks TAMs for metabolic support via stimulating lipid transfer and acquisition C_LIO_LIDisruption of the TRPC-TAM lipid axis, including through FABP3 targeting, reprograms the immune landscape, limits TAM recruitment, and impairs tumor progression C_LIO_LITRPC lineage and associated micro-niche transcriptomic profiles can serve as criteria for patient stratification and identification of responders to lipid-lowering therapies, such as statins C_LI

cancer biology↗

bioRxiv2025

A detailed study Biofilm forming Polyextremophilic Firmicutes from the Himalayas and exploration of their plant growth promoting potential.

Polyextremophily has been an intriguing point of interest among the scientists, a phenomenon in which organisms can thrive in combinations of multiple extreme conditions. In the foreseeable future, climatic conditions are predicted to change, and agriculture is bound to suffer with these changes in the environment. The situation necessitates some solutions that can ensure agricultural sustainability, in the changed, harsh environmental conditions without more toxic chemical intrusions in the agricultural fields. The bacterial strains Brevibacillus parabrevis BRAM_Y3 and Mesobacillus subterraneous BRAM_Y2 in this study, isolated from the waters of Yamunotri showed incredible properties of Polyextremophily like tolerance to not only temperatures as high as 70{degrees}C, pH range (4-10), saline, drought, UV and heavy metals such as mercury, iron, silver and arsenic but also their two-dimensional and three-dimensional combinations. The elevated levels of biofilm formation when subjected to stress revealed their property of using biofilm as their shield against harsh conditions. Furthermore, they were studied for their plant growth promotion like nutrient sequestration (N, P, K, plant hormone production etc.) biocontrol properties (cell wall degrading enzymes, siderophores, VOCs etc.) and the ability to confer abiotic stress resistance (ACC Deaminase) to plants. The in-vivo experiments conducted on Zea mays. L yielded conclusive and promising results. The use of these polyextremophiles for agriculture in harsh environments may serve as a solution for the global warming mediated climate and environmental changes.

microbiology↗

bioRxiv2025

Local adaptation in climate tolerance at a small geographic scale contrasts with broad latitudinal patterns

While climate adaptation is typically quantified across broad gradients, the potential for adaptation to the same environmental variables at small scales is rarely tested. If local-scale environmental heterogeneity can generate patterns of adaptation similar to broad gradients, then we currently underestimate adaptive capacity to global change. We quantified population variation in climate tolerance traits and their plasticity for five populations of Drosophila melanogaster from a 3000-km latitudinal gradient, which we contrasted with eight local populations from an environmentally heterogeneous 600x300km area. Population variation in stress tolerance at the local scale was comparable to that across latitude. Consistent with local adaptation, populations from warmer and drier environments showed greater heat and desiccation tolerance, and populations from more predictable environments showed greater plasticity. Climate adaptation at smaller geographic scales can therefore be comparable to adaptation across broad geographic scales. However, patterns of adaptation often changed across geography, which makes predicting responses to global change challenging.

evolutionary biology↗

bioRxiv2025

Variation in temperature but not diet determines the stability of latitudinal clines in tolerance traits and their plasticity

Latitudinal clines are routinely used as evidence of adaptation across broad climatic gradients. However, if environmental variation influences the strength of latitudinal clines, then clinal patterns will be unstable, and using patterns of adaptation to predict population responses to global change will be difficult. To test whether environmental variation influences latitudinal clines, we sampled five populations of Drosophila melanogaster spanning 3000-km of east coast Australia, and measured stress tolerance (heat, cold and desiccation) and body size on flies that developed in six combinations of temperature (13{degrees}C, 25{degrees}C and 29{degrees}C) and diet (standard and low-calorie) treatments. We found latitudinal clines where populations further from the equator had larger wings, higher cold tolerance and lower heat tolerance. For all traits, temperature determined the strength of latitudinal clines, whereas diet had little influence. Steeper clines often emerged in warmer treatments, created by latitudinal clines in plasticity. In the warmest temperature, higher latitude populations showed larger increases in heat tolerance, larger reductions in desiccation tolerance but smaller decreases in cold tolerance. Heat tolerance was the only trait that supported the climate variability hypothesis and a trade-off between plasticity and tolerance. Environment-dependent latitudinal clines are therefore likely to determine variation in population responses to global change.

evolutionary biology↗

bioRxiv2025

KIF5B and Dynein regulate adhesion-dependent Golgi organization and microtubule acetylation

Cell-matrix adhesion regulates Golgi organization through Arf1-mediated dynein recruitment, maintaining its juxtanuclear localization. On loss of adhesion, Arf1 activation drops, causing loss of dynein, promoting differential disorganization of cis- vs trans-Golgi along microtubules. Golgi regulates microtubule nucleation and stability. In fibroblasts, acetylated tubulin levels drop on loss of adhesion, recovering on re-adhesion with time. Active Arf1 overexpression in preventing Golgi disorganization sustains microtubule acetylation, also seen in T24 bladder cancer cells. Active Arf1 binds KIF5B, recruiting it to the Golgi. KIF5B and dynein knockdown disorganize the Golgi as ministacks, with cis- and trans-Golgi. Dynein knockdown disrupts MTOC positioning, causing ministacks to disperse, preventing Golgi reorganization upon re-adhesion. Dispersed ministacks interestingly maintain microtubule acetylation in adherent and non-adherent cells. The joint KIF5B-dynein knockdown causes the Golgi to lose its ribbon morphology, becoming compact while keeping cis- and trans-Golgi together. This also causes a change in spreading, aspect ratio and migration of knockdown cells, which could be regulated by their Golgi phenotype. In evaluating adhesion-dependent Golgi organization, we reveal the Arf1-KIF5B-dynein crosstalk to regulate Golgi-dependent tubulin acetylation and cell function. SummaryKIF5B and dynein are vital microtubule-associated motors that drive organelle positioning and organization. Adhesion-dependent Arf1 activation mediates KIF5B and dyneins recruitment to the Golgi, regulating its organization and position. This, in turn, regulates microtubule acetylation levels, localization, and cellular functions.

cell biology↗

bioRxiv2025

LOSS OF ROR2 TYROSINE KINASE RECEPTOR IS ASSOCIATED WITH ENDOTHELIAL DYSFUNCTION IN PAH VIA INAPPROPRIATE INTEGRIN BETA 1 ACTIVATION

RationaleEndothelial dysfunction is a key feature of pulmonary arterial hypertension (PAH). We previously identified Wnt7a, a ligand of the Wnt planar cell polarity (PCP) pathway, as essential for pulmonary angiogenesis, with its loss linked to PAH. Given the importance of Wnt/PCP to lung endothelial function and angiogenesis, our goal is to elucidate how Wnt/PCP regulates angiogenic responses in pulmonary microvascular endothelial cells (PMVECs). ROR2, a tyrosine kinase receptor specific to Wnt/PCP, is crucial for cardiovascular development, but its role in PAH is unclear. We hypothesized that ROR2 supports endothelial homeostasis, and its loss would impair angiogenesis, contributing to PAH. MethodsEndothelial-specific ROR2 knockout (ROR2 ECKO) and wild-type (WT) mice were studied under normoxia and chronic hypoxia using echocardiography, hemodynamics, and lung morphometry. PMVECs from healthy and PAH lungs were transfected with ROR2 siRNA/constructs for functional and molecular studies. Focal adhesion (FA) activation and force generation were assessed via FRET-based methods. Bulk and single-cell transcriptomic analyses were performed on siROR2 PMVECs and ROR2 ECKO lungs. ResultsROR2 ECKO mice exhibited worsened pulmonary hypertension, right ventricular remodeling, microvascular loss, and muscularization in hypoxia. Single-cell RNA sequencing of lung endothelial cells showed dysregulation of pathways involved in barrier formation and angiogenesis. Evans blue dye extravasation confirmed reduced endothelial barrier integrity in ROR2 ECKO mice. ROR2-deficient PAH PMVECs displayed increased adhesion, permeability, and FA numbers, with reduced VE-cadherin at cell junctions. Confocal imaging revealed ROR2 localization in FAs, interacting with integrin {beta}1 (ITGB1). FRET analysis showed that ITGB1 remained in an active, adhesion-promoting state in ROR2-deficient cells. Restoring ROR2 in PAH PMVECs normalized adhesion, barrier function, and FA abundance. Transcriptomic analysis identified Rab12 as a key mediator of ROR2-ITGB1 crosstalk, with Rab12 knockdown mimicking ROR2 deficiency in PMVECs. ConclusionsROR2 regulates pulmonary angiogenesis by maintaining endothelial barrier integrity and facilitating integrin recycling. Restoring ROR2 signaling could be a potential therapeutic approach for PAH.

cell biology↗

bioRxiv2025

CES1 DEFICIENCY IS ASSOCIATED WITH METABOLIC REPROGRAMING AND ENDOTHELIAL DYSFUNCTION IN PULMONARY ARTERIAL HYPERTENSION

BackgroundPulmonary arterial hypertension (PAH) is a progressive disease characterized by pulmonary microvascular loss and obliterative remodeling, driven by metabolic reprogramming, oxidative stress, and endothelial dysfunction. While BMPR2 mutations contribute to metabolic shifts in pulmonary microvascular endothelial cells (PMVECs), their low penetrance suggests additional genetic modifiers play a role. A genetic screen of PAH PMVECs identified carboxylesterase 1 (CES1)--an endoplasmic reticulum (ER) enzyme involved in lipid metabolism and detoxification--as a candidate regulator of endothelial metabolism and angiogenesis. We hypothesize that CES1 loss promotes endothelial dysfunction via metabolic reprogramming, lipotoxicity, and oxidative stress. MethodsPAH and healthy PMVECs and lung tissues were obtained from transplant donors and commercial sources. CES1 expression was modulated in PMVECs using siRNA knockdown and plasmid overexpression. Mitochondrial and ER function were assessed via confocal microscopy and proteomics. CES1 heterozygous knockout (HET KO) and endothelial-specific knockout (ECKO) mice were exposed to normoxia or hypoxia, with lung tissues analyzed by single-cell RNA sequencing (scRNA-seq) and histopathology. ResultsCES1 expression was significantly reduced in PAH PMVECs and vascular lesions. CES1-deficient PMVECs exhibited increased apoptosis, reactive oxygen species (ROS) production, mitochondrial fragmentation, ER stress, and impaired angiogenesis. Confocal imaging and metabolic studies revealed lipid droplet accumulation, reduced fatty acid oxidation, and a glycolytic shift-- phenotypes reversed by CES1 restoration. Mechanistically, CES1 transcription was induced by BMPR2 via NRF2 activation, a key regulator of redox and metabolic homeostasis. In vivo, CES1-deficient mice developed severe pulmonary hypertension (PH) under hypoxia, with extensive vascular remodeling, right ventricular dysfunction, and dysregulated angiogenesis and lipid metabolism pathways, as confirmed by lung scRNA-seq. ConclusionsCES1 is essential for pulmonary endothelial homeostasis and serves as a critical modifier of BMPR2 signaling. Given the limited efficacy of current PAH therapies in reversing endothelial dysfunction and small-vessel loss, restoring CES1 expression represents a promising therapeutic strategy.

biochemistry↗

bioRxiv2025

Loofah, a newly characterized adhesion protein, suppresses cell death in long-lived Drosophila hindgut enterocytes

Tissue maintenance in the presence of cell death-promoting insults requires a host of molecular mechanisms. Many studies focus on cell renewal through regeneration, while fewer studies explore mechanisms that promote cell longevity despite cell death stimuli. Here, we reveal that the adult Drosophila hindgut ileum is an excellent model to study tissue maintenance by long-lived cells. Hindgut ileal enterocytes resist the damaging detergent SDS and upstream caspase signaling by head-involution-defective (hid). This hid-induced death insensitivity arises early in adulthood and associates with numerous transcriptional changes. We interrogated 82 of these transcriptional changes in a candidate screen for enhancers of hid-induced death in the ileum. Top among our screen hits is an immunoglobulin family cell adhesion gene, CG15312. CG15312 maintains the adhesion protein FasIII on cell membranes. In hid-expressing ileal cells, CG15312 loss causes cell death and pyknotic nuclear clustering. We name this conserved gene low on-membrane fas and enhancer of hid (loofah). Our findings reveal a new mechanism linking cell adhesion and cell death resistance in a long-lived cell type. Our work establishes a new model to study tissue preservation.

cell biology↗

bioRxiv2025

Revealing the impact of Pseudomonas aeruginosa quorum sensing molecule 2'-aminoacetophenone on human bronchial-airway epithelium and pulmonary endothelium using a human airway-on-a-chip

Pseudomonas aeruginosa (PA) causes severe respiratory infections utilizing multiple virulence functions. Our previous findings on PA quorum sensing (QS)-regulated small molecule, 2-aminoacetophenone (2-AA), secreted by the bacteria in infected tissues, revealed its effect on immune and metabolic functions favouring a long-term presence of PA in the host. However, studies on 2-AAs specific effects on bronchial-airway epithelium and pulmonary endothelium remain elusive. To evaluate 2AAs spatiotemporal changes in the human airway, considering endothelial cells as the first point of contact when the route of lung infection is hematogenic, we utilized the microfluidic airway-on-chip lined by polarized human bronchial-airway epithelium and pulmonary endothelium. Using this platform, we performed RNA-sequencing to analyse responses of 2-AA-treated primary human pulmonary microvascular endothelium (HPMEC) and adjacent primary normal human bronchial epithelial (NHBE) cells from healthy female donors and potential cross-talk between these cells. Analyses unveiled specific signaling and biosynthesis pathways to be differentially regulated by 2-AA in epithelial cells, including HIF-1 and pyrimidine signaling, glycosaminoglycan, and glycosphingolipid biosynthesis, while in endothelial cells were fatty acid metabolism, phosphatidylinositol and estrogen receptor signaling, and proinflammatory signaling pathways. Significant overlap in both cell types in response to 2-AA was found in genes implicated in immune response and cellular functions. In contrast, we found that genes related to barrier permeability, cholesterol metabolism, and oxidative phosphorylation were differentially regulated upon exposure to 2-AA in the cell types studied. Murine in-vivo and additional in vitro cell culture studies confirmed cholesterol accumulation in epithelial cells. Results also revealed specific biomarkers associated with cystic fibrosis and idiopathic pulmonary fibrosis to be modulated by 2-AA in both cell types, with the cystic fibrosis transmembrane regulator expression to be affected only in endothelial cells. The 2-AA-mediated effects on healthy epithelial and endothelial primary cells within a microphysiological dynamic environment mimicking the human lung airway enhance our understanding of this QS signaling molecule. This study provides novel insights into their functions and potential interactions, paving the way for innovative, cell-specific therapeutic strategies to combat PA lung infections.

molecular biology↗

bioRxiv2025

Developing epidermis acquires nutrients from the external milieu by mTOR-dependent macropinocytosis

The vertebrate epidermis transitions from a simple bilayer to a complex, multilayered epithelium during development. Periderm, the outermost layer of the bilayered epidermis, is in direct contact with the external aquatic surrounding. While the barrier function of the epidermis is well established, the functional repertoire of the developing epidermis remains unclear. Using zebrafish, we uncover a function of the periderm in nutrient acquisition. Our findings reveal a developmentally regulated wave of macropinocytosis, which is further amplified by extracellular nutrients, leading to the formation of large endo-lysosomal compartments termed Stimulated Nutrient Activated Compartments (SNACs). We identify mTOR as a key integrator of developmental and nutrient-derived signals that modulate SNAC biogenesis. A global lipidomics analysis demonstrates that macropinocytosis-mediated uptake drives the production of metabolically relevant lipids and enhances animal survival when gut function is compromised. Our study identifies a hitherto unappreciated route of nutrient acquisition by the metabolically responsive developing epidermis. We propose that this mechanism fulfils a vital nutritional role, particularly during the transitional phase when maternally provided resources are depleting and the gut is not yet fully operational.

developmental biology↗

bioRxiv2025

Spatial ploidy inference using quantitative imaging

Polyploidy (whole-genome multiplication) is a common yet under-surveyed property of tissues across multicellular organisms. Polyploidy plays a critical role during tissue development, following acute stress, and during disease progression. Common methods to reveal polyploidy involve either destroying tissue architecture by cell isolation or by tedious identification of individual nuclei in intact tissue. Therefore, there is a critical need for rapid and high-throughput ploidy quantification using images of nuclei in intact tissues. Here, we present iSPy (Inferring Spatial Ploidy), a new unsupervised learning pipeline that is designed to create a spatial map of nuclear ploidy across a tissue of interest. We demonstrate the use of iSPy in Arabidopsis, Drosophila, and human tissue. iSPy can be adapted for a variety of tissue preparations, including whole mount and sectioned. This high-throughput pipeline will facilitate rapid and sensitive identification of nuclear ploidy in diverse biological contexts and organisms.

developmental biology↗

bioRxiv2025

Bioinformatics analysis identifies sequence determinants of enzymatic activity for the PHARC associated lipase ABHD12

In humans, PHARC (polyneuropathy, hearing loss, ataxia, retinitis pigmentosa, and cataract) is an early onset autosomal recessive neurological disorder caused by deleterious mutations to ABHD12 (/{beta}-hydrolase domain protein # 12). Biochemically, ABHD12 functions as a lipase, and catalyzes the hydrolysis of lysophosphatidylserine (lyso-PS) (lyso-PS lipase). By doing so, it controls the concentrations and signaling pathways regulated by this potent signaling lysophospholipid in the mammalian brain. While genetic mapping efforts have identified over 30 mutations in ABHD12 from human PHARC subjects, the biochemical activity of these pathogenic mutants remains unknown. To understand this, here, we performed an exhaustive bioinformatics survey, and collated ABHD12 protein sequences from various organisms across evolution. Next, based on sequence alignments and structural modeling, we identified functionally relevant conserved residues in the ABHD12 protein sequence that are potentially important for its enzymatic activity. To validate these in silico findings, we generated numerous mutants of murine ABHD12, including those associated with human PHARC subjects, and assayed them for their enzymatic activity. Taken together, these complementary in silico and biochemical studies provide the first thorough sequence-function relationship for mammalian ABHD12, especially relevant in the context of PHARC. Finally, our evolutionary analysis identified CG15111 as an ABHD12 ortholog in the fruit fly (Drosophila melanogaster), and enzymatic assays indeed confirmed that recombinant CG15111 has robust lyso-PS lipase activity. Flies serve as an excellent animal system to model various human neurological diseases, and the identification of CG15111 as a Drosophila melanogaster ABHD12 ortholog opens new avenues to study PHARC in fly models.

biochemistry↗

bioRxiv2025

Synaptic vesicle glycoprotein 2 enables viable aneuploidy following centrosome amplification

Amplified centrosome number causes genomic instability, most severely through division into more than two aneuploid daughter cells (multipolar mitosis). Several mechanisms that suppress multipolar division have been uncovered, yet mechanisms that favor viable multipolar division are poorly understood. To uncover factors that promote viability in cells with frequent centrosome amplification and multipolar division, we conducted an unbiased Drosophila genetic screen. In 642 mutagenized lines, we exploited the ability of intestinal papillar cells to form and function despite multipolar divisions. Our top hit is an unnamed gene, CG3168. We name this gene synaptic vesicle glycoprotein 2, reflecting homology to human Synaptic Vesicle Glycoprotein 2 (SV2) proteins. GFP-tagged SV2 localizes to the plasma membrane. In cells with amplified centrosomes, SV2 positions membrane-adjacent centrosomes, which prevents severe errors in chromosome alignment and segregation. Our results uncover membrane-based multipolar division regulation and reveal a novel vulnerability in cells with common cancer properties.

cell biology↗