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Joshi, S.

Publications and source records attributed to Joshi, S..

At least 55 records · Page 3Linked to original sources

Pioneering New Breeding Pathways with 3PaTec: Unlocking the Power of Polyspermy-Induced Triparentage for Sustainable Crop Development

Over the last few centuries, advancements in plant breeding have revolutionized agriculture, driving significant increases in global food production. Polyploidy, the increase in chromosome copies, can positively affect plant performance and is assumed to have played a critical role in the domestication of crop plants. Polyploidy is thought to be primarily caused by sperm that, due to meiotic aberrations, deliver unreduced chromosome sets. We have recently identified an alternative pathway to polyploidization by demonstrating that polyspermy, the fertilization of an egg cell by more than one sperm, occurs in planta and results in viable triploid plants. Capitalizing on a novel high-throughput polyspermy detection tool, we have shown that polyspermy involving two pollen donors can generate plants with three parents, one mother and two fathers. This 3PaTec technology not only speeds up breeding processes through an instant combination of beneficial traits from three parents; it also allows selective polyploidization of the egg cell, thereby bypassing the central cell-derived embryo-nourishing endosperm, a major hybridization barrier. Here, we further explore the genetic and developmental factors influencing polyspermy and show that the frequency of polyspermy and triparental plant formation varies among ecotypes and depends on pollen availability, suggesting that polyspermy is an adaptive trait. Additionally, we extend the application of 3PaTec to crops by successfully generating triparental sugar beet in-field using a wind pollination strategy. Our findings highlight the potential of 3PaTec for major crop plants. This innovative breeding technology does not rely on genetic engineering, requiring minimal technical expertise and infrastructure. As a result, it is highly accessible to a wide range of users, contributing to the democratization of plant breeding by empowering individuals from all backgrounds to collaborate and contribute to developing resilient and sustainable crops.

plant biology↗

Acoustofluidic patterning for improved microtissue histology

Microtissues (such as organoids or spheroids) are useful to model healthy or diseased human tissues and organs, with application in drug testing, tissue engineering, and personalized medicine. To characterize their morphology and phenotype, microtissues are often monitored using histology and high-resolution imaging. While this technique is well-established and standard for macroscopic tissue sections, it remains challenging to execute and has low throughput in the context of these sub-millimeter structures, mainly due to the fact that their size makes them difficult to locate within the embedding medium. To enhance the efficiency of microtissue histology, we developed an acoustofluidic device that arranges microtissues in a coplanar arrangement within HistoGel, a widely used embedding medium. Pre-patterning microtissues within HistoGel streamlines downstream processing by positioning the centers of mass of the microtissues at a pre-defined plan in the sample, simplifying sectioning, labeling, and imaging. We validated the method using differently sized HepG2 microtissues and tumor microtissues from osteosarcoma cells (Saos-2, MG-63, HOS), demonstrating that this method can be applied to different microtissue shapes, sizes, and types, without modifying established post-processing techniques. Our approach substantially enhances the microtissue histology workflow and broadens possibilities for microtissue analysis.

bioengineering↗

RAS pathway activation drives clonal selection and monocytic differentiation in FLT3 and BCL2 inhibitor resistance

Despite efficacy of FLT3 and BCL2 inhibition in acute myeloid leukemia (AML), relapse limits survival. Mutation status and AML monocytic differentiation are implicated in resistance. On-treatment tumor evolution may select for genetically distinct clones or shifts in differentiation not resolvable by bulk sequencing. We performed multiomic single cell (SC) DNA/protein and RNA/protein profiling of patients treated on a clinical trial of the BCL2 inhibitor venetoclax and the FLT3 inhibitor gilteritinib (Ven/Git) to characterize immunophenotypic, transcriptional, and genetic clonal evolution on therapy. We found that while Ven/Gilt effectively eliminated FLT3 mutant clones, it selected for RAS mutations, RAS pathway activation and RAS-associated monocytic differentiation. In an in vitro model of monocytic differentiation associated with heightened RAS pathway activation, we demonstrated that MEK inhibition re-sensitized to Ven/Gilt. These data indicate RAS signaling is central to FLT3 and BCL2 inhibitor resistance, is tightly coupled to monocytic differentiation and can be overcome by RAS pathway inhibition. COIC.C.S. has provided educational talks for Astellas Pharma, served on advisory boards for Genentech/Abbvie and received research funding from Abbvie. B.C., Y.S. and J.H. are employees of Abbvie. M.S. and H.H. are or were previously employees of Genentech. FundingThis work was supported in part by Abbvie. C.C.S. is a Leukemia & Lymphoma Society Scholar in Clinical Research and a Damon Runyon-Richard Lumsden Foundation Clinical Investigator supported (in part) by the Damon Runyon Cancer Research Foundation (CI-99-18). Statement of SignificanceMutational and non-mutational RAS signaling activation drives clonal selection, monocytic differentiation and treatment resistance to FLT3 and BCL2 inhibition in AML. MEK inhibition can resensitize resistant AML cells, suggesting therapeutic potential for combined FLT3, BCL2 and RAS pathway inhibition in AML.

cancer biology↗

VAMP8 Deficiency Attenuates AngII-Induced Abdominal Aortic Aneurysm Formation via Platelet Reprogramming and Enhanced Extracellular Matrix Stability

BACKGROUNDAs vascular sentries, platelets, and their ability to release a host of bioactive molecules, are critical for vascular homeostasis as well as hemostasis. Despite data linking platelet activation to abdominal aortic aneurysms (AAA) and rupture, the underlying mechanisms remain poorly understood. This study addresses the hypothesis that VAMP8, the primary v-SNARE controlling platelet exocytosis, contributes to AAA formation. METHODS AND RESULTSIn an AngII-infused hypercholesterolemic mouse model, we observed significant platelet consumption, indicated by decreased platelet counts at both acute (5-day) and chronic (28-day) time points. Platelets accumulated at sites of elastin degradation and within false lumens of the abdominal aorta after 28 days of AngII infusion. Bulk RNA sequencing analysis of washed platelets and their releasates after 5 days of AngII infusion revealed significant transcriptomic changes, suggesting rapid reprogramming of platelet function. Parallel RNA-seq analysis of suprarenal aortic tissue highlighted changes in genes associated with extracellular matrix (ECM) organization, inflammation, and platelet signaling, linking platelets to vascular remodeling suggesting a "platelet-aorta axis". Laser speckle imaging in a FeCl injury model confirmed that VAMP8 deficiency impaired platelet function, resulting in delayed thrombosis. In vivo experiments demonstrated that VAMP8-/- mice were protected against AngII-induced AAA and aortic rupture. Aortic diameter analysis further revealed that VAMP8 deficiency significantly attenuated AngII-driven aortic pathology. RNA-seq analysis of platelets and aortic tissue suggests that loss of VAMP8 affects expression of genes controlling ECM degradation and aortic wall stability consistent with the protective effect of VAMP8 loss on AAA. CONCLUSIONShort-term AngII infusion appears to reprogram the platelet transcriptome, which may affect the aorta and contribute to AAA formation. Controlling cargo release from platelets via VAMP8 deficiency results in profound attenuation of aortic aneurysms. This introduces a novel paradigm for understanding the impact of reprogrammed platelet cargo secretion and function in aortopathies. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=139 SRC="FIGDIR/small/635525v1_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@22bcecorg.highwire.dtl.DTLVardef@a15207org.highwire.dtl.DTLVardef@1106fd8org.highwire.dtl.DTLVardef@1ee0ea_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIPlatelet transcriptome is altered at early aneurysmal stage. C_LIO_LIVAMP8 deficiency attenuates aortic aneurysms, potentially via enhanced ECM stability. C_LIO_LIVAMP8 deficiency significantly alters various genes contributing to aortic wall structure and stability in both platelets and suprarenal aortic tissue. C_LI

physiology↗

Enhanced yield and subtype identity of hPSC-derived midbrain dopamine neuron by modulation of WNT and FGF18 signaling

While clinical trials are ongoing using human pluripotent stem cell-derived midbrain dopamine (mDA) neuron precursor grafts in Parkinsons disease (PD), current protocols to derive mDA neurons remain suboptimal. In particular, the yield of TH+ mDA neurons after in vivo grafting and the expression of some mDA neuron and subtype-specific markers can be further improved. For example, characterization of mDA grafts by single cell transcriptomics has yielded only a small proportion of mDA neurons and a considerable fraction of contaminating cell populations. Here we present an optimized mDA neuron differentiation strategy that builds on our clinical grade ("Boost") protocol but includes the addition of FGF18 and IWP2 treatment ("Boost+") at the mDA neurogenesis stage. We demonstrate that Boost+ mDA neurons show higher expression of EN1, PITX3 and ALDH1A1. Improvements in both mDA neurons yield and transcriptional similarity to primary mDA neurons is observed both in vitro and in grafts. Furthermore, grafts are enriched in authentic A9 mDA neurons by single nucSeq. Functional studies in vitro demonstrate increased dopamine production and release and improved electrophysiological properties. In vivo analyses show increased percentages of TH+ mDA neurons resulting in efficient rescue of amphetamine induced rotation behavior in the 6-OHDA rat model and rescue of some motor deficits in non-drug induced assays, including the ladder rung assay that is not improved by Boost mDA neurons. The Boost+ conditions present an optimized protocol with advantages for disease modeling and mDA neuron grafting paradigms.

cell biology↗

Regular Plasmodium falciparum importation onto Bioko Island, Equatorial Guinea, hampers malaria elimination from the island

The Bioko Island Malaria Elimination Project (BIMEP) has made significant progress in reducing the prevalence of Plasmodium falciparum on Bioko Island, Equatorial Guinea. However, like other malaria endemic islands like Sao Tome and Principe and Zanzibar, Tanzania, elimination efforts are hampered by imported infections. In an effort to understand the local transmission dynamics and the influence of importation on Bioko Islands P. falciparum population, whole-genome sequences were generated from field samples collected during the BIMEPs 2019 Malaria Indicator Survey (MIS). Within the sub-Saharan African context, we observed Bioko Island parasites did not significantly differentiate from nearby continental neighbors. Among Bioko infections, within-host diversity and the quantity of polyclonal infections appear similar to an area of moderate malaria transmission. However, we observed higher than expected genetic diversity among Bioko parasites, similar to high transmission areas, suggesting imported strains are contributing to transmission on the island. Among Biokos closest geographical neighbors, the flow of parasites with Bioko appeared more pronounced with the Gabonese parasite population, implying more importation may be coming from this region than others. Overall, despite significant investment in malaria control, results illustrate the challenges of eliminating malaria without both interrupting local transmission and accounting for importation from higher transmission areas, likely due to human migration. For there to be sustained progress towards elimination, the BIMEP needs, if feasible, to conduct targeted interventions of outgoing/incoming travelers, and ideally expand malaria control interventions to the continental region of Equatorial Guinea. ImportancePlasmodium falciparum accounts for the majority of malaria deaths globally, with over 500,000 estimated deaths in 2023, predominantly in sub-Saharan Africa, despite strong investment in control and elimination interventions. Incorporating sequencing technologies into malaria surveillance is viewed as a powerful tool to improve control strategies, including identifying parasite transmission pathways. Here, we provide the first genomic characterization of P. falciparum on Bioko Island and Equatorial Guinea since malaria control began in 2004, and use these data to better understand the contribution of neighboring regions to Biokos P. falciparum population. Results highlight the need to account for offisland contributors to transmission and to understand how unmitigated transmission in neighboring regions can hamper progress. This study furthers our understanding of how the flow of parasites between regions impacts infectious disease control and provides foundational genomic data in a previously undescribed region that can be used to inform malaria elimination efforts.

genomics↗

Identification of genetic variation in genes linked to buparvaquone resistance in Theileria sp infecting dairy cattle in India.

Buparvaquone (BQO) is used for treatment of bovine theileriosis, a tickborne disease caused by parasites of the Theileria genus. Studies on T. annulata have linked the mechanism of BQO resistance predominantly to genetic variations in the parasite cytochrome b (cytb) gene. In addition, cryptic mechanisms of resistance involving the parasite peptidyl-prolyl isomerase (pin1) and dihydroorotate dehydrogenase (dhodh) genes require assessment. In India, where bovine theileriosis is endemic, and BQO is widely used for treatment, monitoring and establishing the link between genetic variations in cytb, dhodh and pin1 genes and BQO resistance is essential. In this study, multiplexed PCR amplification and nanopore sequencing approaches were used for genotyping the complete gene loci of the target genes. Analysis of 420 T. annulata field samples collected from seven different states of India revealed the presence of previously reported variations S129G, A146T and P253S in cytb and A53P in pin1, which are linked to BQO resistance. The role of A146T, a highly prevalent variation which mostly co-occurred with I203V, in BQO resistance needs to be evaluated. From 60 samples having T. orientalis infection, the genetic variations identified from the three genes were found to be mostly natural variations based on the reference genotypes and were distinct from T. annulata variations. This study has revealed the presence of BQO resistance-linked cytb gene mutations in T. annulata infecting dairy cattle in India and establishes a nanopore sequencing method for molecular surveillance of genetic variation in field samples. Author SummaryBuparvaquone (BQO) is the most effective drug that is available for the treatment of cattle theileriosis caused by parasites of the Theileria genus. However, treatment failure due to drug resistance is reported, and gene mutations linked to BQO resistance have been reported in African and Middle Eastern countries. These mutations occur in the cytochrome b (cytb) and peptidyl-prolyl isomerase (pin1) genes in the parasite. In this study, genetic variations occurring in these two genes and the parasite dihydroorotate dehydrogenase (dhodh) gene have been mapped using PCR amplification and Nanopore sequencing from field samples collected from seven different states of India. At least three previously reported mutations linked to BQO resistance were found in the T. annulata cytb gene sequence obtained from field samples. While one of these mutations (A146T) was highly prevalent, the other two were found to occur in only a few samples. Similarly, the BQO resistance linked to A53P mutation in the T. annulata pin1 gene was present in only a few samples. Despite their low frequency, the presence of these mutations reveals the existence of BQO resistance due to genetic variations in the parasite population present in India. This is the first comprehensive report of BQO resistance-conferring mutations occurring in Theileria parasites affecting dairy cattle in India and establishes a scalable method for large-scale molecular surveillance studies.

genomics↗

A novel Pbunavirus phage from the Ganges River in biocontrol of multidrug-resistant Pseudomonas aeruginosa in wastewater by downregulation of las/rhl systems and reduction of KPC carbapenemases

Wastewater systems are reservoirs of multidrug-resistant (MDR) pathogens, including Pseudomonas aeruginosa, mirroring those found in the hospital effluents. Effective treatment of raw sewage for MDR P. aeruginosa abatement is critical for safe effluent disposal. This study reports the isolation of a novel phage ({phi}PRS-1) within the Pbunavirus genus from a pristine stretch of the Ganges River, targeting the MDR strain of P. aeruginosa. Antibacterial and antibiofilm assays of {phi}PRS-1 against P. aeruginosa PAO1 demonstrated robust efficacy, achieving log10 reductions of 8.14 in planktonic cells, 9.83 in biofilm inhibition, and 7.01 in biofilm disruption after 24 hours. Additionally, {phi}PRS-1 effectively disrupted the preformed biofilm of various other MDR P. aeruginosa strains with >5 log10 reductions. Mechanistically, {phi}PRS-1 downregulated quorum-sensing genes and reduced the carbapenemase resistance KPC cluster, validated through qRT-PCR analysis. Evaluation of {phi}PRS-1 in raw sewage revealed a 4.2 log10 CFU/mL reduction in the P. aeruginosa counts, highlighting its efficacy in complex environmental matrices. In vitro, cytotoxicity assays confirmed the phages safety. The use of {phi}PRS-1 for tackling MDR P. aeruginosa in the wastewater offers a sustainable alternative, potentially reducing reliance on other treatment methods. The findings have attempted to deliver SDGs 3, 6, 11, 14, and 15. HighlightsO_LIA novel phage ({phi}PRS-1) from an untapped location of the Ganges River C_LIO_LI{phi}PRS-1 reduces planktonic cells of MDR P. aeruginosa by 8.14 log10 in 24 hours C_LIO_LIAntibiofilm potentials: 9.83 log10 reduction (inhibition) >5 log10 reduction (disruption) C_LIO_LIDownregulates quorum-sensing genes and reduces KPC resistance in MDR P. aeruginosa C_LIO_LI{phi}PRS-1 shows 4.2 log10 CFU/mL reduction of MDR P. aeruginosa in raw sewage C_LI Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=111 SRC="FIGDIR/small/621487v1_ufig1.gif" ALT="Figure 1"> View larger version (49K): org.highwire.dtl.DTLVardef@14f4ac0org.highwire.dtl.DTLVardef@b4a8d9org.highwire.dtl.DTLVardef@b97a6borg.highwire.dtl.DTLVardef@114072c_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗

Antibody Profiles Elicited by Potent and Subpotent Whole Cell Pertussis Vaccines in Mice

Inactivated, whole cell pertussis (wP) vaccines remain at the frontline in the global fight against the resurgence of whooping cough, especially in low- and middle-income countries. However, the reliance on the intracerebral mouse potency test (ic-MPT or Kendrick assay) as the standard batch release assay is challenging for the production of commercial wP vaccines. The ic-MPT is technically challenging, labor intensive and, arguably, incongruous with modern animal welfare guidelines. Replacing the ic-MPT with a whole cell Bordetella pertussis ELISA, the so-called pertussis serology potency test (PSPT), has shown promise, but has been difficult to implement in practice. In this report, we tested the hypothesis that potent and subpotent wP vaccines have distinct serologic profiles in mice that could be developed as a substitute for the ic-MPT. We first established an accelerated decay (thermal stress) protocol in which wP, in the context of DTwP, was rendered >10-fold less effective than unstressed vaccine when evaluated in a mouse model of B. pertussis lung clearance following intranasal challenge. We then screened immune sera on a limited B. pertussis Tahoma I proteome array and identified >30 antigens whose antibody reactivity profiles increased, decreased or were unchanged as a function of wP potency. Interestingly, virtually all the "indicator" antigens identified are known virulence factors or reactive with human convalescent sera, thereby establishing a potential link between wP potency and pertussis infection and immunity. These results bode well for the development of a limited B. pertussis antigen array as a stability-indicating surrogate potency assay for the ic-MPT.

immunology↗

Grey Pineapple Mealybug, Dysmicoccus neobrevipes Beardsley: A New Devastating Pest of Tuberose in India

Tuberose (Agave amica) is a highly valued flower crop in India, known for its consistently high returns and ability to thrive in various climatic conditions. Severe outbreaks of the grey pineapple mealybug, Dysmicoccus neobrevipes Beardsley (Hemiptera: Pseudococcidae), have been observed in several villages of the Pune district, Maharashtra (India), since 2021, leading to significant economic losses. This report provides an illustrative morphological diagnosis of D. neobrevipes and basic information on its field establishment, impact on tuberose crops, nature of the damage and field symptoms etc. Surveys conducted in major tuberose-growing areas of Pune district (Maharashtra) revealed that the mealybug primarily infests the underground and basal portion of tuberose plants, causing stunted growth, drooping, and ultimately death of the plants. D. neobrevipes produces honeydew, fostering sooty mold growth that impairs photosynthesis, reduces flower yield. It also infests tuberose bulbs, spoiling them in storage. Based on primary scientific literature, this study represents the first scientifically confirmed record of D. neobrevipes as a new pest of tuberose in India. The ant species Solenopsis geminata was found attending to mealybug colonies, aiding their spread and creating a nuisance for farmers during routine field activities. A total of 87.09% of the surveyed fields (n=62) showed mealybug infestation, ranging from 68% to 97%. Raising awareness among the farmers and implementing regular monitoring in tuberose-growing areas are crucial steps for developing effective management practices and preventing the further spread of this pest to other regions.

zoology↗

Widespread Neuroanatomical Integration and Distinct Electrophysiological Properties of Glioma-Innervating Neurons

Gliomas are the most common malignant primary brain tumors and are often associated with severe neurological deficits and mortality. Unlike many cancers, gliomas rarely metastasize outside the brain, indicating a possible dependency on unique features of brain microenvironment. Synapses between neurons and glioma cells exist, suggesting that glioma cells rely on neuronal inputs and synaptic signaling for proliferation. Yet, the locations and properties of neurons that innervate gliomas have remained elusive. In this study, we utilized transsynaptic tracing with a pseudotyped, glycoprotein-deleted rabies virus to specifically infect TVA and glycoprotein-expressing human glioblastoma cells in an orthotopic xenograft mouse model, allowing us to identify the neurons that form synapses onto the gliomas. Comprehensive whole-brain mapping revealed that these glioma-innervating neurons (GINs) consistently arise at brain regions, including diverse neuromodulatory centers and specific cortical layers, known to project to the glioma locations. Molecular profiling revealed that these long-range cortical GINs are predominantly glutamatergic, and subsets express both glutamatergic and GABAergic markers, whereas local striatal GINs are largely GABAergic. Electrophysiological studies demonstrated that while GINs share passive intrinsic properties with cortex-innervating neurons, their action potential waveforms are altered. Our study introduces a novel method for identifying and mapping GINs and reveals their consistent integration into existing location-dependent neuronal network involving diverse neurotransmitters and neuromodulators. The observed intrinsic electrophysiological differences in GINs lay the groundwork for future investigations into how these alterations may correspond with the postsynaptic characteristics of glioma cells. SignificanceWe have developed a novel system utilizing rabies virus-based monosynaptic tracing to directly visualize neurons that synapse onto human glioma cells implanted in mouse brain. This approach enables the mapping and quantitative analysis of these glioma-innervating neurons (GINs) in the entire mouse brain and overcomes previous barriers of molecular and electrophysiological analysis of these neurons due to the inability to identify them. Our findings indicate that GINs integrate into existing neural networks in a location-specific manner. Long-range GINs are mostly glutamatergic, with a subset expressing both glutamatergic and GABAergic markers and local striatal GINs are GABAergic, highlighting a complex neuromodulatory profile. Additionally, GINs exhibit unique action potential characteristics, distinct from similarly selected neurons in non-tumor-bearing brains. This study provides new insights into neuronal adaptations in response to forming putative synapses onto glioma, elucidating the intricate synaptic relationship between GINs and gliomas.

cancer biology↗

Navigating the Landscape of Fear

O_LIAnimals searching for food must navigate complex landscapes with varying terrain, food availability, predator activity, and shelter. Where and when should they gather food? To what extent should they engage in anti-predator behaviors such as vigilance or seeking refuge if a predator is detected? Optimal foraging theory (OFT) posits that animals balance potentially conflicting goals (such as feeding versus escaping predation) by making decisions that maximize some expected utility or reward. However, OFT models have generally considered highly simplified landscapes, either ignoring spatial variability or assuming that the habitat consists of discrete, internally uniform habitat patches. As a result, OFT has largely avoided the question of how animals should move from one potential feeding area to another, or between feeding areas and refuges. C_LIO_LIWe develop methods based on stochastic dynamic programming to find optimal foraging strategies, including optimal movement paths, in a continuous landscape with spatially varying predation risk. Our approach accounts for switching from foraging to escape behavior when pursued by a predator. Because contingent escape paths are considered for all visited locations, they influence the optimal foraging path even before threats are encountered. The optimal strategy thus depends on the animals level of hunger, the distribution of food, and the perceived threat distribution. The realized path is further influenced by actual predator encounters. C_LIO_LIWe illustrate our approach with two numerical examples: the first hypothetical with two food-abundant regions accessible only via high-risk areas, the second based on empirical studies on foraging Samango monkeys, Cercopithecus albogularis schwarzi. We find that the shape of the foragers utility function (risk-averse, risk-neutral, or with state-dependent risk sensitivity) affects not only its choices of where to feed, but also the optimal paths to and from each feeding ground. C_LIO_LIOur methods make it possible to compare properties of observed foraging trajectories with those predicted for different goal functions. Foraging trajectories can then provide additional information, along with other behavioral choices, about what quantity, if any, animals aim to optimize while foraging. C_LI

ecology↗

Genome-Wide CRISPR-Cas9 Screening Identifies a Synergy between Hypomethylating Agents and SUMOylation Blockade in MDS/AML

Hypomethylating agents (HMAs) are frontline therapies effective at altering the natural course of Myelodysplastic Neoplasms (MDS) and Acute Myeloid Leukemia (AML). However, acquired resistance and treatment failure are hallmarks of HMA therapy. To address this clinical need, we performed a genome-wide CRISPR-Cas9 screen in a human MDS-derived cell line, MDS-L, and identified TOPORS as a highly ranked loss-of-function target that synergizes with HMAs, reducing leukemic burden and improving survival in xenograft models. We demonstrate that the depletion of TOPORS mediates sensitivity to HMAs by predisposing leukemic blasts to an impaired DNA damage response (DDR) accompanied by an accumulation of SUMOylated DNMT1 in HMA-treated TOPORS-depleted cells. Importantly, the combination of HMAs with targeting of TOPORS did not functionally impair healthy hematopoiesis. While inhibitors of TOPORS are currently unavailable, we show that inhibition of protein SUMOylation (upstream of TOPORS functions) with TAK-981 partially phenocopies HMA-sensitivity and DDR impairment. Overall, our data suggest that the combination of HMAs with the inhibition of SUMOylation or TOPORS demonstrates a favourable therapeutic index and is a rational treatment framework for High-Risk MDS (HR-MDS) or AML.

molecular biology↗

Determining Optimal Placement of Copy Number Aberration Impacted Single Nucleotide Variants in a Tumor Progression History

Intratumoral heterogeneity arises as a result of genetically distinct subclones emerging during tumor progression. These subclones are characterized by various types of somatic genomic aberrations, with single nucleotide variants (SNVs) and copy number aberrations (CNAs) being the most prominent. While single-cell sequencing provides powerful data for studying tumor progression, most existing and newly generated sequencing datasets are obtained through conventional bulk sequencing. Most of the available methods for studying tumor progression from multi-sample bulk sequencing data are either based on the use of SNVs from genomic loci not impacted by CNAs or designed to handle a small number of SNVs via enumerating their possible copy number trees. In this paper, we introduce DETOPT, a combinatorial optimization method for accurate tumor progression tree inference that places SNVs impacted by CNAs on trees of tumor progression with minimal distortion on their variant allele frequencies observed across available samples of a tumor. We show that on simulated data DETOPT provides more accurate tree placement of SNVs impacted by CNAs than the available alternatives. When applied to a set of multi-sample bulk exome-sequenced tumor metastases from a treatment-refractory, triple-positive metastatic breast cancer, DETOPT reports biologically plausible trees of tumor progression, identifying the tree placement of copy number state gains and losses impacting SNVs, including those in clinically significant genes.

cancer biology↗

Generative interpolation and restoration of images using deep learning for improved 3D tissue mapping

The development of novel imaging platforms has improved our ability to collect and analyze large three-dimensional (3D) biological imaging datasets. Advances in computing have led to an ability to extract complex spatial information from these data, such as the composition, morphology, and interactions of multi-cellular structures, rare events, and integration of multi-modal features combining anatomical, molecular, and transcriptomic (among other) information. Yet, the accuracy of these quantitative results is intrinsically limited by the quality of the input images, which can contain missing or damaged regions, or can be of poor resolution due to mechanical, temporal, or financial constraints. In applications ranging from intact imaging (e.g. light-sheet microscopy and magnetic resonance imaging) to sectioning based platforms (e.g. serial histology and serial section transmission electron microscopy), the quality and resolution of imaging data has become paramount. Here, we address these challenges by leveraging frame interpolation for large image motion (FILM), a generative AI model originally developed for temporal interpolation, for spatial interpolation of a range of 3D image types. Comparative analysis demonstrates the superiority of FILM over traditional linear interpolation to produce functional synthetic images, due to its ability to better preserve biological information including microanatomical features and cell counts, as well as image quality, such as contrast, variance, and luminance. FILM repairs tissue damages in images and reduces stitching artifacts. We show that FILM can decrease imaging time by synthesizing skipped images. We demonstrate the versatility of our method with a wide range of imaging modalities (histology, tissue-clearing/light-sheet microscopy, magnetic resonance imaging, serial section transmission electron microscopy), species (human, mouse), healthy and diseased tissues (pancreas, lung, brain), staining techniques (IHC, H&E), and pixel resolutions (8 nm, 2 {micro}m, 1mm). Overall, we demonstrate the potential of generative AI in improving the resolution, throughput, and quality of biological image datasets, enabling improved 3D imaging.

bioinformatics↗

DCs targeted therapy expands CD8 T cell responses to bona-fide neoantigens in lung tumors

Cross-presentation by type 1 cDCs (cDC1) is critical to induce and sustain antitumoral CD8 T cell responses to model antigens, in various tumor settings. However, the impact of cross-presenting cDC1 and the potential of DC-based therapies in tumors carrying varied levels of bona-fide neoantigens (neoAgs) remains unclear. Here we generated a non-small cell lung cancer model with distinct ranges of TMB and MHC-I neoepitopes to test immunogenicity and response to Flt3L+CD40 (DC-therapy). We found that cDC1 are required to broaden the pattern of CD8 responses to basal and acquired neoAgs and DC-therapy strongly inhibits the growth of TMBhigh tumors. In contrast, TMBlow tumors induce weaker responses that are not sufficient to block progression. scRNA transcriptional analysis, immune profiling and functional assays show that DC-therapy triggers the accumulation of lung cDC1 with increased immunostimulatory properties and CD8 T cells with enhanced cytotoxic functions and reduced exhaustion, most prominently in neoAgshigh tumors. We conclude that boosting cDC1 activity is critical to broaden the diversity of anti-tumoral CD8 T cell responses and to leverage neoAgs content for therapeutic advantage.

immunology↗

Plasticity in inhibitory networks improves pattern separation in early olfactory processing

Distinguishing between nectar and non-nectar odors is challenging for animals due to shared compounds and varying ratios in complex mixtures. Changes in nectar production throughout the day - and potentially many times within a foragers lifetime - add to the complexity. The honeybee olfactory system, containing fewer than 1,000 principal neurons in the early olfactory relay, the antennal lobe (AL), must learn to associate diverse volatile blends with rewards. Previous studies identified plasticity in the AL circuits, but its role in odor learning remains poorly understood. Using a biophysical computational network model, tuned by in vivo electrophysiological data, and live imaging of the honeybees AL, we explored the neural mechanisms and functions of plasticity in the early olfactory system. Our findings revealed that when trained with a set of rewarded and unrewarded odors, the AL inhibitory network suppresses shared chemical compounds while enhancing responses to distinct compounds. This results in improved pattern separation and a more concise neural code. Our calcium imaging data support these predictions. Analysis of a graph convolutional neural network performing an odor categorization task revealed a similar mechanism for contrast enhancement. Our study provides insights into how inhibitory plasticity in the early olfactory network reshapes the coding for efficient learning of complex odors. Significance StatementBy combining biophysical modeling, machine learning, electrophysiology and analysis of calcium imaging data, we demonstrate that associative and nonassociative plasticity in the honeybee antennal lobe (AL) the first relay of the insect olfactory system - work together to enhance the contrast between rewarded and unrewarded odors. Training the ALs inhibitory network within specific odor environments enables the suppression of neural responses to common odor components while amplifying responses to distinctive ones. This study sheds light on the olfactory systems ability to adapt and efficiently learn new odor-reward associations across varying environments, and it proposes innovative, energy-efficient principles applicable to artificial intelligence.

neuroscience↗

Three-dimensional assessments are necessary to determine the true spatial tissue composition of diseased tissues

Methods for spatially resolved cellular profiling using thinly cut sections have enabled in-depth quantitative tissue mapping to study inter-sample and intra-sample differences in normal human anatomy and disease onset and progression. These methods often profile extremely limited regions, which may impact the evaluation of heterogeneity due to tissue sub-sampling. Here, we applied CODA, a deep learning-based tissue mapping platform, to reconstruct the three-dimensional (3D) microanatomy of grossly normal and cancer-containing human pancreas biospecimens obtained from individuals who underwent pancreatic resection. To compare inter-and intra-sample heterogeneity, we assessed bulk and spatially resolved tissue composition in a cohort of two-dimensional (2D) whole slide images (WSIs) and a cohort of thick slabs of pancreas tissue that were digitally reconstructed in 3D from serial sections. To demonstrate the marked under sampling of 2D assessments, we simulated the number of WSIs and tissue microarrays (TMAs) necessary to represent the compositional heterogeneity of 3D data within 10% error to reveal that tens of WSIs and hundreds of TMA cores are sometimes needed. We show that spatial correlation of different pancreatic structures decay significantly within a span of microns, demonstrating that 2D histological sections may not be representative of their neighboring tissues. In sum, we demonstrate that 3D assessments are necessary to accurately assess tissue composition in normal and abnormal specimens and in order to accurately determine neoplastic content. These results emphasize the importance of intra-sample heterogeneity in tissue mapping efforts.

cancer biology↗