Biology subjects

Chaturvedi, P.

Publications and source records attributed to Chaturvedi, P..

At least 19 recordsLinked to original sources

bioRxiv2026

A Smart Bait Dispenser with Real-time Monitoring for Species-Specific Bait Delivery

1. Remote image capture using trail cameras has become an essential part of many wildlife monitoring and management programs. Oral bait vaccination has been a valuable tool in wildlife disease management, but baits are generally dropped on the ground or delivered by devices that are typically developed to utilise physical and behavioural characteristics of the target species. Both approaches can result in a high proportion of baits being consumed by non-target species. Merging years of trail camera research knowledge with new technologies allows for development of more specific bait delivery options. 2. We developed an open-source smart bait dispenser that uses field-based image classification to selectively deliver baits to the target species. The system's long-range wide area network-based remote telemetry provides updates to an online dashboard in near real time to show camera triggers, target species detections, and baits dispensed. We trialled four dispensers with placebo baits for 62 nights across three field sites in Tasmania, Australia to deliver baits to Tasmanian devil (Sarcophilus harrisii). 3. The dispensers recorded 2106 motion triggers, of which 876 were triggered by animals. The onboard devil classification model achieved 98.6% accuracy, 75.0% precision, 77.4% recall, and a 76.2% F1-score. The dispenser released 41 placebo baits in total, of which 31 were target-specific distributions in response to presence of Tasmanian devils at the dispenser, hence achieving an overall bait delivery success rate of 75.6%. Importantly, no animal was able to forcibly retrieve baits from the dispenser. Observed average battery life across trials was 19.2 nights. 4. These results clearly demonstrate the effectiveness of our innovative bait dispenser for targeted bait delivery, offering a versatile ecological tool that can be tailored for various species by simply updating the image classification model. This adaptable system can support critical initiatives like wildlife oral bait vaccination and supplementary feeding after wildlife translocations.

ecology↗

bioRxiv2026

Free-Energy and Molecular Dynamics Analysis of Ser195 and Thr195 Rotameric States in Thrombin

Ser195 is highly conserved as the catalytic nucleophile in trypsin like serine proteases, yet the molecular basis for the strong preference for serine over the chemically similar threonine residue remains unclear. Here, we used well tempered metadynamics and microsecond long molecular dynamics (MD) simulations to characterize the conformational energetics of S195 and T195 in thrombin. Well tempered metadynamics revealed distinct rotameric free energy landscapes in both the apo and the ligand bound systems. In the apo state, S195 preferentially occupied a lower angle rotameric state, while T195 exhibited two low energy states with similar free energies. Unbiased simulations further showed greater conformational heterogeneity and increased higher angle sampling for T195. Evaluation of the His57 to residue 195 hydrogen bond further revealed that S195 maintained close hydrogen bond distances across both rotameric regions, whereas T195 exhibited longer and more variable His57 to hydroxyl distances with reduced distance based occupancy. Analysis of the local environment showed that the additional T195 methyl group was associated with redistributed protein contacts. Additional rotamer conditioned analyses demonstrated differences in local packing within corresponding conformational states. Furthermore, solvent accessibility and water contact analyses revealed that the effects of the substitution on the catalytic side chain environment depend on rotameric conformation. Together, these findings demonstrate that two complementary simulation approaches converge on similar conformational preferences and suggest that the S195T substitution alters the rotameric free energy landscape in association with changes in catalytic geometry, local packing, and solvation. These results provide insight into the evolutionary preference for serine over threonine at the catalytic position of thrombin.

biophysics↗

bioRxiv2026

Characterizing the Conformational Dynamics of an Intrinsically Disordered Localization Sequence

Mitochondrial localization peptides (MLPs) play a critical role in directing proteins to mitochondria, yet how subtle sequence variations influence their conformational behavior remains poorly understood. Here, we investigate the conformational dynamics of a 15-residue MLP derived from the androgen receptor, together with a comprehensive panel of single-residue variants generated by systematic substitution at the second position. Across all variants, the peptide remains intrinsically disordered, exhibiting broad conformational heterogeneity and no stable folded state. Global measures of compactness show that single-residue substitutions induce only modest changes to overall peptide dimensions. In contrast, residue-level analysis reveals that the identity of the second residue subtly reshapes local structural preferences, particularly near the N-terminus. Small or hydrophobic substitutions enhance transient -helical sampling, whereas polar or charged substitutions favor increased disorder and {beta}- or polyprolinelike conformations. Comparison across variants further distinguishes mutations that preserve wild-type-like structural behavior from those that produce more pronounced deviations in the conformational ensemble. Enhanced sampling simulations highlight the complexity and ruggedness of the underlying free-energy landscape and demonstrate the challenges associated with achieving convergence for short intrinsically disordered peptides. Collectively, these results show that even minimal sequence changes can bias the dynamic structural ensemble of mitochondrial localization peptides, suggesting a potential mechanism by which targeting efficiency may be modulated. More broadly, this work underscores the importance of advanced sampling strategies for accurately characterizing intrinsically disordered localization signals and provides a framework for connecting sequence variation to functional targeting behavior.

biophysics↗

bioRxiv2026

HSF1-dependent, long-range directional HSPA1 gene motion to nuclear speckles is coupled to DDX39B condensate dynamics

Approximately 5-10% of the genome, enriched [~]10-fold in the most highly active genes, positions near deterministically within several hundred nm of nuclear speckles (NS). This includes the HSPA1 locus, among the most strongly NS-associated genes. Here we address how HSPA1 endogenous genes and transgenes stably position adjacent to the nuclear speckle periphery, facilitating their gene expression amplification. Live-cell imaging demonstrates a transcription-dependent speckle-anchoring of HSPA1 genes. Removal of this anchoring reveals sustained, long-range, curvilinear oscillations of the HSPA1 genes away from and back to nuclear speckles or between nuclear speckles. Curvilinear transgene movements are dependent on the transcription factor HSF1, largely independent of nuclear actin or cohesin, but unexpectedly spatially and temporally correlated with DDX39B condensate dynamics and dependent on DDX39B protein levels. We propose that stable nuclear speckle association of HSPA1 results from combining metastable speckle anchoring with constitutive, restorative long-range chromosome movements driven by DDX39B condensate dynamics.

cell biology↗

bioRxiv2025

Acidic transcription factors position the genome at nuclear speckles through transcription dependent and independent mechanisms

A small fraction of the genome positions reproducibly near nuclear speckles (NS), increasing expression and/or splicing efficiency of NS-associated genes. How specific genomic regions target to NS remains unclear. Here, we demonstrate that establishment of genome-wide NS-association occurs independent of active transcription. We show that DNA sequences derived from NS-associated regions integrated as transgenes autonomously target to NS. By systematically dissecting one such genomic locus, the COL1A1-SGCA locus, we identified redundant NS-targeting cis regulatory elements, including a [~]600 bp fragment with 17 binding motifs for 8 transcription factors (TFs). Four NS-targeting TFs within this fragment contain acidic activation domains (AADs) that provide both chromatin-context and transcription-dependent NS-targeting, a property that appears common among several other tested AADs. A subset of acidic activator TFs contain an additional, transcription-independent NS-targeting activity. Our findings establish diverse and partially redundant NS-targeting activities, which may facilitate dynamic gene positioning at NS periphery for context-specific transcriptional responses.

cell biology↗

bioRxiv2025

A study on edge devices for image classification of the Tasmanian devil (Sarcophilus harrisii) for vaccine delivery

O_LIA target-specific bait dispenser is required for oral bait vaccination of the endangered Tasmanian devil (Sarcophilus harrisii) against the deadly devil facial tumour disease. Development of a broadly extendible dispenser would be a beneficial ecological tool. A camera-based edge device with an onboard deep learning model can be used to identify the target species using image classification. It is important to choose a suitable edge device for the dispensers effective application in power constrained environments. C_LIO_LIWe evaluated four edge devices for the smart bait dispenser-ArduinoPro Nicla Vision, ArduinoPro Portenta H7 with Vision Shield (LoRa), Raspberry Pi 3 Model B+ and Raspberry Pi Zero 2 W. Two simple convolutional neural networks, and four fine-tuned pretrained models (MobileNetV2, MobileNetV3Small, ResNet50V2, and ResNet152V2) were trained on trail camera images of devil and non-devil species. These models were evaluated across four metrics and post-training quantised for deployment. The edge devices were assessed on inference latency of each model in seconds (s) and power consumption in watt (W). C_LIO_LIWe found that a simple CNN based image classification model yielded the best overall result of [~]96% across all metrics. Raspberry Pi 3 Model B+ and Zero 2 W could run all the six models whereas Portenta failed to run the two ResNet models and Nicla Vision failed to run all four pretrained models. All edge devices were found to be quick enough for dispenser application (average inference latency of 0.718 s). Portenta consumed the lowest power during inference (0.19 W), idle (0.185 W), and light sleep (0.045 W) states whereas Nicla Vision consumed the lowest power during deep sleep (0.002 W). C_LIO_LIOverall, we found that simple traditional CNNs was best suited for species classification in a camera-based smart bait dispenser for vaccination of Tasmanian devils. We conclude that ArduinoPro Portenta H7 with Vision Shield is the most competent device for this edge AI application. We suggest integration of this system with a microcontroller unit like ATtiny85 to minimise inactive power consumption of the dispenser. This method can be readily adapted for other species in vaccination and supplemental feeding projects. C_LI

ecology↗

bioRxiv2025

Highly active chromosome regions preferentially associate with two perispeckle networks that partition the interchromatin space

A subset of highly active chromosomal "hot zones" reproducibly positions adjacent to nuclear speckles (NS). Genes within these regions amplify their expression only with NS contact. However, gene expression differences inversely correlate with differences in NS distance, genome-wide. We hypothesized the existence of additional gene expression "niches" away from, but spatially correlated with, NS. Here we report the identification of two dynamic perispeckle patterns of protein concentrations extending outwards from NS and persisting even after NS are eliminated. Highly active chromosome regions which weakly associate with NS instead show close, NS-independent association with these perispeckle patterns. Additionally, transcripts from model intron-containing versus intronless genes associate differentially with these two patterns. While genes within NS-associated genomic regions are predominantly downregulated upon NS depletion, genes associated with perispeckle patterns are biased towards upregulation. We suggest the interchromatin space is partitioned into additional gene expression "niches"- surrounding and extending from NS - that may be involved in mRNA and gene dynamics.

cell biology↗

bioRxiv2025

Bridging the gap of late-gestation nephrogenesis using a non-human primate model

BackgroundHuman nephrogenesis is complete at 34-36 weeks gestation, with 60% of nephrons forming during the third trimester through lateral branch nephrogenesis (LBN). Currently, no mechanism exists for LBN as there are no late gestation human kidney transcriptional datasets. We hypothesized that a differentiated but dividing population of nephron progenitor cells (NPCs) would contribute to the amplification of nephrons in late gestation. We used the rhesus macaque, an established model of LBN, to help identify potential mechanisms. MethodsSingle-cell RNA-sequencing (scRNA-Seq) was performed on cortically-enriched fetal rhesus kidneys (n=9) from late second trimester and third trimester during LBN. This data was integrated with publicly available human scRNA-seq datasets from 8-18 weeks gestation kidneys (n=8) using state-of-the-art bioinformatics pipelines. Differentially expressed genes and ligand-receptor interactions were assessed and validated using RNAScopeTM on human and rhesus archival tissue. ResultsscRNA-Seq of 64,782 rhesus cells revealed 37 transcriptionally distinct cell populations, including 7,879 rhesus NPCs. Pseudotime analyses identified a late gestation-specific lineage branch of differentiated NPC in rhesus that was not observed in mid-gestation humans. Differential expression analyses identified increased SFRP1, FZD4, and TLE2 and decreased FZD7, SHISA2, SHISA3, and TLE4 within the late-gestation rhesus NPC compared to mid-gestation human NPC and increased SEMA3D within the rhesus ureteric bud (UB) tip, suggesting a compositional shift in WNT and SEMA signaling components within the naive NPC population during LBN. ConclusionThe rhesus macaque uniquely enables molecular studies of late-gestation primate nephrogenesis. Our study suggests the hypothesis that a transitional state of self-renewing NPCs supported by compositional shifts in key pathways may underlie the switch from branching phase nephrogenesis to lateral branch nephrogenesis and support ongoing nephron formation in late gestation. Translational statementNo transcriptional data exists for the late-gestation human kidney, during which 60% of the nephron endowment is formed through a primate-specific process called lateral branch nephrogenesis (LBN). In this study, we used single-cell RNA sequencing the late gestation rhesus macaque as a model for this developmental stage. Our findings suggest a potential mechanism for LBN, in which a transitional state of self-renewing nephron progenitor cells (NPCs) is supported by compositional shifts in key pathways, allowing for continued nephron formation in late gestation.

developmental biology↗

bioRxiv2025

Engineering Large-Scale and Innervated Functional Human Gut for Transplantation

A confined culture system (CCS) establishes methods to generate complex functional human gastrointestinal tissues. This approach yields large-scale innervated small intestinal, colonic and gastric organoids with an elongated tubular shape for both in vitro and in vivo studies. Transcriptomic and electrophysiological data demonstrate the co-development of a functional de novo enteric nervous system, which is absent from conventional organoids. When compared to traditional methods, CCS derived small intestinal, colonic and gastric organoids reached maturation supporting transplantation in half of the time, resulting in enhanced engraftment rates and sizes. Murine luminal content exposure within CCS organoids in vivo further augmented function, supporting the potential translational benefits required to model complex intestinal diseases. In summary, the CCS methodology simplifies current protocols while adding complexity and expediting the generation of clinically relevant functional gut tissues.

bioengineering↗

bioRxiv2025

Energy trade-offs under fluctuating light govern bioenergetics and growth in Chlamydomonas reinhardtii

Rapidly changing light intensity is a natural challenge that photosynthetic organisms can tolerate. Regulatory mechanisms of light harvesting and alternative electron pathways are critical in dissipating and distributing energy under fluctuating light intensities (FL), but less is known about downstream metabolic regulations. Here, we compared the cellular responses of Chlamydomonas reinhardtii grown under FL to cells acclimated to constant high (HL) or low light (LL), either under high (2 %) or low (0.04 %) CO2. Under low CO2, the physiology of FL cells resembled HL cells and proteomics revealed an induction of the ATP consuming carbon-concentrating mechanism, and photorespiration particularly under FL. High CO2 promoted growth under FL, albeit by a lesser extent than under HL and led to higher ATP contents than under low CO2. To fuel ATP production under low CO2, cells upregulated mitochondrial respiration under FL, while enhanced cyclic electron flow and redox shuttling between intracellular compartments was most evident under FL and LL. Chloroplastic carbon metabolism rapidly responded to light changes, independent of CO2 availability, whereas metabolites associated with mitochondrial bioenergetics responded slower, and remained abundant under high CO2. The accumulation of enzymes involved in starch synthesis and breakdown under FL, together with the transient accumulation of hexoses and hexose phosphates, indicated that cells relied on sugars as a transient carbon pool to meet changing metabolic demands under FL. We conclude that the interplay between light intensity and CO2 availability drives critical energy trade-offs, balancing photoprotection, repair, and carbon allocation, that regulate growth under FL.

plant biology↗

bioRxiv2025

Molecular Insights into Cholesterol Concentration Effects on Planar and Curved Lipid Bilayers for Liposomal Drug Delivery

Liposomal carriers provide a flexible and effective strategy for delivering therapeutics across a broad spectrum of diseases. Cholesterol is frequently included in these systems to improve membrane rigidity and limit permeability. Despite its widespread use, the optimal cholesterol-to-lipid proportion for achieving stable and efficient liposome performance remains to be fully determined. In this work, we apply all-atom molecular dynamics simulations to explore how different cholesterol concentrations influence the structural and dynamic characteristics of 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC) bilayers, considering both planar and curved membrane geometries. Bilayers with cholesterol molar ratios of 0%, 10%, 20%, 30%, 40%, and 50% were simulated, and key biophysical parameters including area per lipid (APL), membrane thickness, leaflet interdigitation, and deuterium order parameters (SCD) were analyzed. In planar bilayers, increasing cholesterol concentration led to a progressive decrease in APL from approximately 60 {degrees}A2 to 40 {degrees}A2, accompanied by increased membrane thickness and lipid ordering, consistent with cholesterols classical condensing effect. In contrast, curved bilayers exhibited a cholesterol-induced expansion effect, particularly in the inner leaflet, where APL increased from approximately 60 {degrees}A2 to 90 {degrees}A2 with rising cholesterol levels. SCD profiles showed that cholesterol enhanced tail ordering up to 40% concentration, beyond which the effect plateaued or slightly declined, suggesting structural saturation or packing frustration. Membrane thickness displayed a monotonic increase in planar bilayers but followed a nonlinear trend in curved systems due to curvature-induced stress. These findings highlight that cholesterols influence on membrane properties is highly dependent on bilayer geometry and asymmetry. While planar bilayers exhibit predictable responses, curved systems reveal nonclassical behaviors that challenge traditional models of cholesterol-lipid interactions. This work provides molecular-level insights and establishes a computational framework for the rational design of liposomal systems, emphasizing the need to account for curvature and asymmetry in membrane engineering.

biophysics↗

bioRxiv2025

Regulation of early-stage tomato fruit growth by the splicing factor RS2Z36

Fruit growth is mediated by cell division and expansion. In tomato, the model for fleshy fruit development, both processes are tightly linked to changes in gene expression, including transcriptional regulation and RNA processing. While several transcription factors are implicated in fruit developmental programs, the role of splicing regulators remains largely unexplored. Expression profiling of splicing-related genes revealed expression patterns. The serine/arginine-rich splicing factor RS2Z36 is expressed in ovaries and during early fruit development. Loss-of-function mutations in RS2Z36 result in ovaries with altered patterning and in smaller, ellipsoid fruits. rs2z36 mutants display elongated pericarp cells along the longitudinal axis of pre-anthesis ovaries, indicating that RS2Z36-dependent expansion patterns are established before anthesis. RNA-seq uncovered widespread alternative splicing of genes across diverse biological processes, while proteome analysis revealed altered protein abundance and peptides derived from novel splice variants. In addition, rs2z36-1 pericarps show increased deposition of LM6-recognized arabinan and AGP epitopes. Together, these findings identify RS2Z36 as a regulator of ovary and fruit development and highlight a previously underappreciated role for splicing control in shaping early fruit morphology.

developmental biology↗

bioRxiv2024

A fast-track, high-throughput screening platform for biological nitrification inhibitors discovery based on soil relevant ammonia oxidizing strains

Nitrogen cycling is critical for ecosystem functioning, with nitrification playing a central role. Excessive nitrification, triggered by heavy nitrogen fertilisation, contributes to reduced nitrogen use efficiency, nitrate leaching, and nitrous oxide emissions. While synthetic nitrification inhibitors (SNIs) help reduce these impacts, their erratic performance and environmental risks have shifted focus to biological nitrification inhibitors (BNIs) as sustainable alternatives. In vitro bioassays with ammonia-oxidising microorganisms (AOM) are valuable for BNI discovery and research, but often have low throughput and rely on a limited number of mostly non-soil-relevant or genetically modified ammonia-oxidizing bacteria (AOB) strains, lacking validation with established BNIs. We present a refined fast-track, high-throughput assay for BNI screening, utilizing soil-relevant, ecophysiologically and phylogenetically diverse AOB (Nitrosospira multiformis, Nitrosomonas ureae, Nitrosomonas communis) and ammonia-oxidizing archaea (AOA) strains (Nitrososphaera viennensis, "Ca. Nitrosocosmicus franklandianus"), achieving consistent cellular activity and density. The assay was validated with established SNIs and BNIs, showing differences in inhibition efficacy and strain sensitivity, consistent with literature. As a proof of concept, root exudates from diverse wheat genotypes were screened, demonstrating distinct inhibition profiles. The proposed system advances previously available screening systems and, when integrated with realistic soil tests, will facilitate the discovery of novel BNIs and BNI-producing plant genotypes.

microbiology↗

bioRxiv2024

Chromatin-associated lncRNA-splicing factor condensates regulate hypoxia responsive RNA processing of genes pre-positioned near nuclear speckles

Hypoxia-induced alternative splicing (AS) regulates tumor progression and metastasis. Little is known about how such AS is controlled and whether higher-order genome and nuclear domain (ND) organizations dictate these processes. We observe that hypoxia-responsive alternatively spliced genes position near nuclear speckle (NS), the ND that enhances splicing efficiency. NS-resident MALAT1 long noncoding RNA, induced in response to hypoxia, regulates hypoxia-responsive AS. MALAT1 achieves this by organizing the SR-family of splicing factor, SRSF1, near NS and regulating the binding of SRSF1 to pre-mRNAs. Mechanistically, MALAT1 enhances the recruitment of SRSF1 to elongating RNA polymerase II (pol II) by promoting the formation of phase-separated condensates of SRSF1, which are preferentially recognized by pol II. During hypoxia, MALAT1 regulates spatially organized AS by establishing a threshold SRSF1 concentration near NSs, potentially by forming condensates, critical for pol II-mediated recruitment of SRSF1 to pre-mRNAs.

molecular biology↗

bioRxiv2024

Fetal Liver-like Organoids Recapitulate Blood-Liver Niche Development and Multipotent Hematopoiesis from Human Pluripotent Stem Cells

The fetal liver is a hematopoietic organ, hosting a diverse and evolving progenitor population. While human liver organoids derived from pluripotent stem cells (PSCs) mimic aspects of embryonic and fetal development, they typically lack the complex hematopoietic niche and the interaction between hepatic and hematopoietic development. We describe the generation of human Fetal Liver-like Organoids (FLOs), that model human hepato-hematopoietic interactions previously characterized in mouse models. Developing FLOs first integrate a yolk sac-like hemogenic endothelium into hepatic endoderm and mesoderm specification. As the hepatic and hematopoietic lineages differentiate, the FLO culture model establishes an autonomous niche capable of driving subsequent progenitor differentiation without exogenous factors. Consistent with yolk sac-derived waves, hematopoietic progenitor cells (HPCs) within FLOs exhibit multipotency with a preference for myeloid lineage commitment, while retaining fetal B and T cell differentiation potential. We reconstruct in FLOs the embryonic monocyte-to-macrophage and granulocyte immune trajectories within the FLO microenvironment and assess their functional responses in the liver niche. In vivo, FLOs demonstrate a liver engraftment bias of hematopoietic cells, recapitulating a key phenomenon of human hematopoietic ontogeny. Our findings highlight the intrinsic capacity of liver organoids to support hematopoietic development, establishing FLOs as a platform for modeling and manipulating human blood-liver niche interactions during critical stages of development and disease.

developmental biology↗

bioRxiv2024

Analyzing Biodiversity in Protected vs. Unprotected Tide Pools

This study assessed the efficacy of marine protected areas (MPAs) by comparing tide pool biodiversity in Half Moon Bay, California between a protected site (Fitzgerald Marine Reserve) and an unprotected site (Mavericks Beach). Samples were collected through random quadrat surveys. ImageJ and RStudio were used to analyze data through three methods utilizing the Simpsons Diversity Index along with three metrics. The unprotected Mavericks Beach displayed greater biodiversity through two metrics while the third did not find significant difference in biodiversity between sites. 30 unique species were present at each site with 43 species between both. These mixed findings prevent us from drawing definitive conclusions on the effectiveness of Half Moon Bays tide pool MPAs but suggest the MPA is less biodiverse. Potential explanations include the Intermediate Disturbance Hypothesis, lack of sample size, or that the MPA is not effective in maintaining biodiversity.

ecology↗

bioRxiv2024

Integration of multi-omics and deep phenotyping provides novel insights into multiple abiotic stress responses in potato

Potato is highly water and space efficient but susceptible to abiotic stresses such as heat, drought, or flooding, which are severely exacerbated by climate change. Understanding of crop acclimation to abiotic stress, however, remains limited. Here, we present a comprehensive molecular and physiological high-throughput profiling of potato (Solanum tuberosum, cv. Desiree) under heat, drought and waterlogging applied as single stresses or in combinations designed to mimic realistic future scenarios. Stress-responses were monitored via daily phenotyping and multi-omics analyses of leaf samples comprising transcriptomics, proteomics, metabolomics and hormonomics at several timepoints during and after stress treatments. Additionally, critical metabolites of tuber samples were analysed at the end of the stress period. Integrative analysis of multi-omics data was performed using a bioinformatic pipeline, which was established here, based on machine learning and knowledge networks. Overall, waterlogging had the most immediate and dramatic effects on potato plants, interestingly activating ABA-responses similar to drought stress. In addition, we observed distinct stress signatures at multiple molecular levels in response to heat or drought and to a combination of both. In response to all treatments, we found a downregulation of photosynthesis at different molecular levels, an accumulation of minor amino acids and diverse stress induced hormones. Our integrative multi-omics analysis provides global insights into plant stress responses, facilitating improved breeding strategies towards climate-adapted potato varieties. One Sentence SummaryIntegrated multi-omics analysis of high-throughput phenotyping in potato reveals distinct molecular signatures of acclimation to single and combined abiotic stresses related to climate change.

plant biology↗

bioRxiv2023

Reduced nephron endowment in the common Six2-TGCtg mouse line is due to Six3 misexpression by aberrant enhancer-promoter interactions in the transgene

Lifelong kidney function relies on the complement of nephrons generated during mammalian development from a mesenchymal nephron progenitor cell (NPC) population. Low nephron endowment confers increased susceptibility to chronic kidney disease. We asked whether reduced nephron numbers in the popular Six2TGC transgenic mouse line1 was due to disruption of a regulatory gene at the integration site or to ectopic expression of a gene(s) contained within the transgene. Targeted locus amplification identified integration of the Six2TGC transgene within an intron of Cntnap5a on chr1. We generated Hi-C datasets from NPCs isolated from the Six2TGCtg/+ mice, the Cited1CreERT2/+control mice, and the Six2TGCtg/+;Tsc1+/Flox,2 mice that exhibited restored nephron number compared with Six2TGCtg/+mice, and mapped the precise integration of Six2TGC and Cited1CreERT2transgenes to chr1 and chr14, respectively. No changes in topology, accessibility, or expression were observed within the 50-megabase region centered on Cntnap5a in Six2TGCtg/+ mice compared with control mice. By contrast, we identified an aberrant regulatory interaction between a Six2 distal enhancer and the Six3 promoter contained within the transgene. Increasing the Six2TGCtg to Six2 locus ratio or removing one Six2 allele in Six2TGCtg/+ mice, caused severe renal hypoplasia. Furthermore, CRISPR disruption of Six3 within the transgene (Six2TGC{Delta}Six3CT) restored nephron endowment to wildtype levels and abolished the stoichiometric effect. Data from genetic and biochemical studies together suggest that in Six2TGC, SIX3 interferes with SIX2 function in NPC renewal through its C-terminal domain. SignificanceUsing high-resolution chromatin conformation and accessibility datasets we mapped the integration site of two popular transgenes used in studies of nephron progenitor cells and kidney development. Aberrant enhancer-promoter interactions drive ectopic expression of Six3 in the Six2TGCtgline which was correlated with disruption of nephrogenesis. Disruption of Six3 within the transgene restored nephron numbers to control levels; further genetic and biochemical studies suggest that Six3 interferes with Six2-mediated regulation of NPC renewal.

developmental biology↗