Biology subjects

Rodriguez, D.

Publications and source records attributed to Rodriguez, D..

At least 19 recordsLinked to original sources

bioRxiv2026

Closing the phenotyping gap: non-invasive electromagnetic induction enables high-throughput screening of crop root function in the field

Crop root systems play a critical role in water uptake and yield improvement under drought. However, a lack of methods to measure root system function at scale, particularly in the field, poses significant challenges to explore them in crop improvement programs. To address this bottleneck, we introduce a novel approach to directly screen water uptake and root activity in the field through integrating electromagnetic induction (EMI) sensing, inversion modelling and machine learning. This study was conducted as an on-farm field experiment comprising a factorial combination of 13 sorghum genotypes and three row spacings with three replications. Time-lapse EMI surveys were conducted from flowering to maturity using a customised DualEM-21S sensing system. Artificial neural network models were developed to predict 3D soil moisture ({theta}) using depth-specific true soil electrical conductivity estimated from EMI inversion algorithms, and the prediction accuracy was evaluated. Crop water use and functional root traits were derived from changes in {theta} profiles between successive EMI surveys using cubic splines fitted within a linear mixed model framework. Results indicate that dynamics of {theta} can be predicted with high accuracy (R2 = 0.83). There was a significant positive correlation between the EMI-derived crop water use and root length density at the depths without water stress. The functional root traits were significantly affected by growth stages, genotypes, row spacings, and their interactions. In general, narrower row spacing resulted in deeper rooting depth by flowering, which was associated with a larger canopy and higher grain number. In contrast, wider row spacing promoted greater post-flowering root growth, leading to increased post-flowering water use and, consequently, larger grain weight. Overall, our approach enables rapid characterisation of variation in crop water use, rooting depth and root growth in the field. It provides a valuable framework for understanding the dynamic interactions between root function and yield traits across variable field environments, with potential to accelerate genotype selection and optimise crop design in breeding and agronomy programs.

plant biology↗

bioRxiv2026

A reversible GNAS-PKA-RUNX2 checkpoint governs skeletal stem cell fate in fibrous dysplasia

Fibrous dysplasia (FD) arises from activating GNAS mutations that increase cAMP signaling in skeletal stem/progenitor cells (SSPCs), but the underlying mechanisms and whether the resulting fate imbalance can be reversed remain unclear. Using controlled GNAS induction with transcriptomic, pharmacologic, and cell-fate analyses, we identify a PKA-dependent checkpoint governing SSPC lineage commitment. GNASR201C redirects SSPCs toward osteoclastogenesis while suppressing osteoblast maturation; genetic or pharmacologic PKA inhibition reverses both effects, revealing this fate decision is dynamically reversible. Mechanistically, PKA phosphorylates RUNX2, the master regulator of osteogenesis, enforcing its cytoplasmic retention and blocking osteogenic transcription; a non-phosphorylatable RUNX2 mutant restores nuclear localization and osteoblast differentiation, defining RUNX2 phosphorylation as a disease-initiating molecular switch. Critically, PKA inhibition in vivo reduces fibrosis and osteoclast burden in established bone lesions, demonstrating reversibility even in advanced FD. These findings define a druggable GNAS-PKA-RUNX2 checkpoint and identify PKA inhibition as a strategy to reverse existing disease in FD.

cell biology↗

bioRxiv2026

Elevated cholesterol in APOE4 astrocytes drives mitochondrial cristae collapse and ATP synthase dysfunction

Cholesterol imbalance is a hallmark of major human diseases, including atherosclerosis and Alzheimers disease, both of which are also associated with mitochondrial dysfunction, yet the mechanistic links between cholesterol and mitochondria remain poorly understood. Here we show that elevated intracellular cholesterol in murine astrocytes expressing the Alzheimers disease risk variant APOE4 disrupts the inner mitochondrial membrane, manifesting as sparse, truncated cristae alongside an excess of cristae junction complexes. These structural abnormalities are accompanied by loss of respiratory chain complexes I and IV, and reduced respiration; nevertheless, low proton leak and reverse ATP synthase activity combine to generate an elevated mitochondrial membrane potential. Strikingly, APOE4 astrocytes are hypersensitive to the ATP synthase inhibitor oligomycin, demonstrating a profound dysfunction of the enzyme, and cholesterol sequestration with methyl-{beta}-cyclodextrin abrogates this toxicity, establishing elevated cholesterol as its proximate cause. Another method of targeting ATP synthase, epicatechin, which prevents the enzyme operating in reverse, attenuated both cholesterol- and respiratory chain inhibitor-induced cell death. Reciprocally, reducing intracellular cholesterol via nutrient restriction restored cristae architecture and partially rescued respiratory chain complex abundance. These findings identify mitochondrial cholesterol as a critical determinant of cristae architecture and ATP synthase function and suggest that cholesterol-driven mitochondrial dysfunction may be a unifying feature of cholesterol-related disorders from neurodegeneration to atherosclerosis. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=145 SRC="FIGDIR/small/742391v1_ufig1.gif" ALT="Figure 1"> View larger version (27K): org.highwire.dtl.DTLVardef@2e80a1org.highwire.dtl.DTLVardef@fecd32org.highwire.dtl.DTLVardef@51ba3corg.highwire.dtl.DTLVardef@199f3c6_HPS_FORMAT_FIGEXP M_FIG Elevated intracellular cholesterol in APOE4 astrocytes disrupts the inner mitochondrial membrane and ATP synthase, resulting in sparse cristae, increased ATP hydrolysis and heightened oligomycin sensitivity. The vulnerability to oligomycin is abrogated by cholesterol sequestration with cyclodextrin, indicating causality. These findings position cholesterol as a key determinant of the cristae landscape and ATP synthase function, highlighting the importance of the emerging field of mitochondrial-cholesterol crosstalk. C_FIG

cell biology↗

bioRxiv2025

The potential role of volatile organic compounds on the colonisation of deadwood by saproxylic beetles

Volatile organic compounds (VOCs) emitted by deadwood are increasingly recognised as key olfactory cues used by saproxylic beetles to locate suitable substrates. However, how beetle communities respond to these cues during the colonisation of deadwood remains poorly understood. To address this gap, it is essential to quantify both VOC emissions and beetle assemblages, and to experimentally disentangle the main ecological drivers of the deadwood volatilome (i.e., tree species, sun exposure). To explore the potential role of VOCs on saproxylic beetle colonisation, we exposed 1200 freshly cut branches of oak, beech, spruce, and pine across Central Europe. To mimic natural variation in deadwood and disturbance processes, the bundles were experimentally manipulated either by sterilisation (to reduce endogenous fungi), fungal inoculation with a brown rot fungus (Fomitopsis pinicola), a white rot fungus (Fomes fomentarius), or burning. From each deadwood bundle, we sampled 448 substances, 89 of which were identified as VOCs and reared 134 saproxylic beetle species. We observed distinct VOC profiles emitted by broadleaf and conifer species, aligning with the beetles tree-type preferences. In conifers, bark beetles, longhorn beetles, and jewel beetles were associated with different chemical cues, whereas beetle taxonomic separation was not observed in broadleaf species. Although experimental treatments altered VOC composition, they could not explain beetle colonisation patterns. Our study highlights that VOCs emitted during the early decay stage of deadwood are associated with distinct saproxylic beetle assemblages. VOC composition varied across tree species and treatments, suggesting that chemical variation reflects multiple ecological factors. These findings indicate that deadwood diversity, particularly in tree species, contributes to chemical heterogeneity, which is linked to broader beetle assemblages. Forest conservation efforts may therefore need to consider the role of chemical variation in deadwood, as it could influence colonisation patterns of saproxylic beetles and affect the success of biodiversity management strategies.

ecology↗

bioRxiv2025

Environmental Detection of the Amphibian Chytrid Fungus in Water Bodies Predicts Host Infection Along a Deforestation Gradient

Understanding pathogen dynamics during environmental life stages is vital for comprehending wildlife diseases, especially for those with a free-living phase. In amphibians affected by the chytrid fungus Batrachochytrium dendrobatidis (Bd), most studies have focused on host-pathogen interactions, with less emphasis on Bds environmental stage. We tested whether the distribution of Bd in natural aquatic environments can predict host infection patterns. We sampled four tropical amphibian species across eight rainforest landscapes with varying habitat loss, testing whether environmental and host Bd detection varied along gradients of habitat change. Using a high-capacity water filtration method coupled with digital and real-time PCR detection assays, we quantified Bd in water and amphibian samples. Our results revealed a strong positive correlation between Bd DNA concentrations in water samples and infection loads on amphibian skin samples. Forest cover and habitat split were the primary predictors of Bd distribution in both free-living and host-associated forms. We identified Bd-GPL and Bd-Asia-2/Brazil lineages across our study landscapes. Our study introduces and validates a robust protocol for detecting Bd in environmental samples, with the potential to enhance monitoring and inform management strategies. Moreover, our work contributes novel, well-replicated spatial data on Bd associations between hosts and their environments.

ecology↗

bioRxiv2025

Key role of TLR3 in type I IFN expression and apoptosis induction in IBDV-infected chicken fibroblast cells

Infectious Bursal Disease Virus (IBDV) (Avibirnavirus genus, Birnaviridae family) is a non-enveloped virus with a double-stranded RNA (dsRNA) genome. IBDV causes a highly contagious and immunosuppressive disease in domestic chickens (Gallus gallus), representing a major threat to the global poultry industry. Apoptotic cell death and exacerbated innate immune responses have been implicated in IBDV pathogenesis. Previous studies from our laboratory demonstrated the crucial role of type I interferon (IFN) in triggering apoptosis in IBDV-infected cell cultures. Genomic IBDV dsRNA is recognized by the cytoplasmic pattern recognition receptor (PRR) melanoma differentiation-associated gene 5 (MDA5) in chicken cells, triggering type I IFN responses. However, the contribution of the endosomal PRR Toll-like receptor 3 (TLR3) dsRNA sensor on type I IFN production upon IBDV infection has not been studied, despite several studies have demonstrated that its expression is significantly upregulated upon IBDV infection. Here, we demonstrate that ablation of TLR3 gene expression in DF-1 chicken fibroblasts results in a complete blockade of IBDV-induced apoptosis, a marked reduction in IFN production, and a significant enhancement of virus progeny yields. Notably, this effect appears to be specific to IBDV, as similar outcomes were not observed with other RNA viruses. Our findings also suggest that TLR3 may also play a role in viral release into the extracellular space. Additionally, receptor interacting protein kinase 1 (RIPK1), a protein that interacts with TLR3 through the adaptor Toll/IL-1 receptor (TIR) domain-containing adaptor-inducing interferon-{beta} (TRIF), was shown to contribute to both IFN production and apoptosis in response to IBDV infection or dsRNA stimulation in DF-1 cells. Overall, this study provides new insights into the innate immune recognition of IBDV, highlighting the central role of TLR3 in mediating antiviral responses in chicken cells. Author summaryInfectious Bursal Disease Virus (IBDV) is the etiological agent of a devastating syndrome known as IBD or Gumboro disease, which affects domestic chickens (Gallus gallus), leading to significant economic losses in the poultry industry worldwide. The virus primarily targets immature B lymphocytes causing their killing by apoptosis, resulting in severe immunosuppression. Diverse studies have suggested that exacerbation of the innate immune response is related to IBDV pathogenesis. Here, we have investigated the role of the cellular sensor of double-stranded RNA (dsRNA) Toll-like receptor 3 (TLR3) on the fate of IBDV-infected cells. Deletion of the TLR3 gene completely blocks apoptosis induced in IBDV-infected DF-1 cells, drastically reduces interferon (IFN) production and improves viral replication efficiency. We also demonstrated the participation of receptor interacting protein kinase 1 (RIPK1), a known downstream mediator of TLR3, in both, IFN production and apoptosis induction in response to IBDV infection. These findings provide new molecular insights into the mechanisms underlying the robust type I IFN response observed during IBDV infection and contribute to a deeper understanding of IBDV pathogenesis.

microbiology↗

bioRxiv2025

Ultrasmall chemogenetic tags with group-transfer ligands

Chemogenetic tags are valuable tools for studying functions of a given protein-of-interest (POI) lacking small-molecule ligands, but most tags are too large for several POIs. Here, we report two ultrasmall chemogenetic tags (mgTag and cTag) of 36 and 50 amino acids (aa) that, to the best of our knowledge, are the smallest reported. These tags exhibit transferase-type reactivity with their ligands to append any moiety of interest to the tag. cTag utilizes an engineered C1 domain-bearing cysteine that undergoes group-transfer reaction with its ligand. Likewise, mgTag utilizes an engineered zinc-finger domain-bearing cysteine that undergoes group-transfer reaction with its molecular-glue ligand in the presence of cereblon (CRBN). We applied these tags in the context of cell signaling and proximity induction. While the fusion of HaloTag (297 aa) to the KRASG12D (188 aa) disrupted its ability to activate the growth-signaling pathway, fusion of mgTag or cTag did not. Group-transfer of BRD4 binder to tags appended to Abelson kinase (ABL) induced proximity between ABL and BRD4, resulting in the latters phosphorylation. Deletion of the transferase-type reactivity reduced phosphorylation levels, suggesting that proximity-inducing chimeras with group-transfer design may be more efficacious in certain scenarios. We envision these ultrasmall tags to have wide-ranging applications, including in basic science, biotechnology, and medicine. TOC O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=168 SRC="FIGDIR/small/653252v1_ufig1.gif" ALT="Figure 1"> View larger version (36K): org.highwire.dtl.DTLVardef@1f99888org.highwire.dtl.DTLVardef@18d3e0corg.highwire.dtl.DTLVardef@1e4a044org.highwire.dtl.DTLVardef@d610ee_HPS_FORMAT_FIGEXP M_FIG C_FIG

synthetic biology↗

bioRxiv2024

Optogenetic Central Amygdala Stimulation is Highly Reinforcing and Strongly Outcompetes Fentanyl Self-Administration in Male, but not Female, Rats

The central nucleus of the amygdala plays a key role in many aspects of substance use disorders, in particular biasing behaviors toward one drug or natural reward over another, yet the role of this region in opioid self-administration remains unclear. Here, we report that pharmacological inactivation of the central amygdala reduces fentanyl self-administration, while intra-central amygdala opioid receptor antagonism dose-dependently increases fentanyl self-administration. We tested whether optogenetic activation of the central amygdala would increase motivation for fentanyl in rats with a long fentanyl self-administration history. While pairing fentanyl delivery with optogenetic central amygdala activation increased fentanyl self-administration, optoactivation itself was highly reinforcing and, in choice settings, was pursued over fentanyl despite mounting effort requirements, delays, and sporadic reward availability. Of note, under free response conditions, these effects were limited to male rats; while female rats avidly responded for optogenetic activation of the central amygdala, this activation was less effective in enhancing fentanyl intake, and it was not preferred over fentanyl. In contrast, under discrete trial choice which precluded independent regulation of intake of the two options, both females and males preferred optoactivation to fentanyl. These results demonstrate that optogenetic stimulation of neural activity within the anterior central amygdala does not appear to potentiate the reinforcing effects of fentanyl, but is itself highly reinforcing regardless of sex, and, in male rats can robustly outcompete fentanyl. In contrast, in females, fentanyl intake is relatively insensitive to competing opportunities for optoactivation, except when opportunities to obtain drug or optoactivation are sparse.

neuroscience↗

bioRxiv2024

T cell epitope mapping reveals immunodominance of evolutionarily conserved regions within SARS-CoV-2 proteome.

As SARS-CoV-2 variants continue to emerge capable of evading neutralizing antibodies, it has become increasingly important to fully understand the breadth and functional profile of T cell responses to determine their impact on the immune surveillance of variant strains. Here, sampling healthy individuals, we profiled the kinetics and polyfunctionality of T cell immunity elicited by mRNA vaccination. Modeling of anti-spike T cell responses against ancestral and variant strains of SARS-CoV-2 suggested that epitope immunodominance and cross-reactivity are major predictive determinants of T cell immunity. To identify immunodominant epitopes across the viral proteome, we generated a comprehensive map of CD4+ and CD8+ T cell epitopes within non-spike proteins that induced polyfunctional T cell responses in convalescent patients. We found that immunodominant epitopes mainly resided within regions that were minimally disrupted by mutations in emerging variants. Conservation analysis across historical human coronaviruses combined with in silico alanine scanning mutagenesis of non-spike proteins underscored the functional importance of mutationally-constrained immunodominant regions. Collectively, these findings identify immunodominant T cell epitopes across the mutationally-constrained SARS-CoV-2 proteome, potentially providing immune surveillance against emerging variants, and inform the design of next-generation vaccines targeting antigens throughout SARS-CoV-2 proteome for broader and more durable protection. One Sentence SummaryPolyfunctional CD8+ and CD4+ T cells directed against SARS-CoV-2 target mutationally constrained regions of the viral proteome.

immunology↗

bioRxiv2024

Earlier endosomal escape improves the catalytic activity of delivered enzyme cargo

There is enormous interest in strategies to efficiently traffic biologics-proteins, nucleic acids, and complexes thereof-into the mammalian cell cytosol and internal organelles. Not only must these materials reach the appropriate cellular locale fully intact and in therapeutically relevant concentrations, they must also retain activity upon arrival. The question of residual activity is especially critical when delivery involves exposure to the late endocytic pathway, whose acidic lumenal environment can denature and/or degrade internalized material. ZF5.3 is a compact, stable, rationally designed mini-protein that efficiently escapes intact from late endocytic vesicles, with or without covalently linked protein cargo. Here, using insights from mechanistic studies on the pathway of endosomal escape and classic principles of zinc(II) bioinorganic chemistry, we re-designed the sequence of ZF5.3 to successfully alter the timing (but not the efficiency) of endosomal escape. The new mini-protein we describe, AV5.3, escapes earlier than ZF5.3 along the endocytic pathway with no loss in efficiency, with or without enzyme cargo. More importantly, earlier endosomal escape translates into higher enzymatic activity of a pH-sensitive enzyme upon arrival in the cytosol. Delivery of the pH-sensitive enzyme dihydrofolate reductase (DHFR) with AV5.3 results in substantial catalytic activity in the cytosol, whereas delivery with ZF5.3 does not. The activity of AV5.3-DHFR upon delivery is sufficient to rescue a genetic DHFR deletion in CHO cells. This work provides evidence that programmed trafficking through the endosomal pathway is a viable strategy for the efficient cytosolic delivery of therapeutic proteins. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=77 SRC="FIGDIR/small/615476v2_ufig1.gif" ALT="Figure 1"> View larger version (21K): org.highwire.dtl.DTLVardef@1de8a77org.highwire.dtl.DTLVardef@154c401org.highwire.dtl.DTLVardef@105ce8org.highwire.dtl.DTLVardef@43c1f3_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2024

Endoglin regulates the integrity of the bone marrow vasculature

Endoglin (Eng) is an accessory receptor for transforming growth factor-{beta} (TGF-{beta}) that is critical for maintaining vascular integrity. Mutations in Eng cause hereditary hemorrhagic telangiectasia (HHT), resulting in arteriovenous malformations (AVMs) and blood abnormalities. Despite the known association between Eng deficiency and AVMs, the underlying mechanisms are unclear. In addition, the role of the bone marrow (BM), a major source of immune and blood cells, in endothelial Eng (EC-Eng) deficiency is unexplored. We show that BM blood vessels conditionally deficient in Eng (cKO) undergo a structured remodeling process over four weeks, with distinct proliferative and resolution phases. These phases involve angiogenic set points, the involvement of integrins, and the modulation of vascular integrity. In addition, we observe changes in hematopoietic stem and progenitor cells (HSPC) and circulating granulocytes, along with reduced red blood cells and platelets due to splenic sequestration. Using a conditional heterozygous EC-Eng deficient mouse model, reflecting the genetics of HHT patients, we identify vascular changes similar to those in the cKO model. Taken together, using multiple in vivo approaches, we suggest that reduced Eng expression in the endothelium drives significant BM vascular remodeling, sharing mechanisms with early vascular processes associated with AVM formation. Explanation of NoveltyOur findings reveal that BM blood vessels deficient in endoglin undergo an orchestrated remodeling process with distinct proliferative and resolution phases over several weeks. We identify specific angiogenic set points and profound alterations in vascular integrity, along with hematopoietic changes starting at the level of hematopoietic stem and progenitor cells. These findings advance our understanding of the role of Eng in vascular remodeling and may provide novel therapeutic targets for HHT. Key PointsO_LIConditional EC-Eng deficiency leads to vascular remodeling in the BM of mice in a temporally orchestrated manner. C_LIO_LIEC-Eng facilitates vascular integrity, hematopoietic homeostasis, and immune cell mobilization. C_LI

molecular biology↗

bioRxiv2024

Elevated prevalence of the global panzootic chytrid strain in 1 Ecuadorian anurans of the Amazonian lowlands

Considerable attention has been directed to studying the infection dynamics of the fungal pathogen, Batrachochytrium dendrobatidis (Bd), affecting amphibians in the high elevations of the Neotropics. Lowland forests of the same realm, on the other hand, remain relatively understudied in this context. Herein, an attempt to bridge this gap was made by investigating the occurrence of Bd in several anuran taxa inhabiting the Amazonian lowlands in the northeast of Ecuador. To this end, 207 anurans belonging to 10 different families, 25 different genera, and 55 distinct host species were sampled for Bd DNA in 2008. Data on the taxonomy, morphology (i.e., weight and snout-vent length), and life-long aquatic dependency of hosts (i.e., aquatic index) were also collated to serve as potential predictors of infection prevalence. Genotyping via quantitative PCR revealed the presence of the global panzootic lineage of Bd (Bd-GPL) in the Ecuadorian Amazon. The overall infection prevalence of Bd was determined to be 58%, which is a relatively high prevalence rate of Bd reported for any amphibian population from the lowlands of the Neotropics to date. A total of 88% of sampled anuran families tested positive for the infection at varying proportions. A logistic regression analysis showed a significant negative relationship between host weight and the proportion of Bd infections (p < 0.05). However, no significant associations were observed between host taxonomy, aquatic dependency, or snout-vent length and Bd prevalence. Our findings contribute to the understanding of Bd dynamics in the Neotropical lowlands and emphasize the need for future research on the ecological factors influencing Bd in the Amazon and their implications for amphibian conservation.

evolutionary biology↗

bioRxiv2024

Genotype-specific expression of uncle fester suggests a role in allorecognition education in a basal chordate

Histocompatibility is the ability to discriminate between self and non-self tissues, and has been described in species throughout the metazoa. Despite its universal presence, histocompatibility genes utilized by different phyla are unique-those found in sponges, cnidarians, ascidians and vertebrates are not orthologous. Thus, the origins of these sophisticated recognition systems, and any potential functional commonalities between them are not understood. A well-studied histocompatibility system exists in the botryllid ascidians, members of the chordate subphylum, Tunicata, and provides an opportunity to do so. Histocompatibility in the botryllids occurs at the tips of an extracorporeal vasculature that come into contact when two individuals grow into proximity. If compatible, the vessels will fuse, forming a parabiosis between the two individuals. If incompatible, the two vessels will reject-an inflammatory reaction that results in melanin scar formation at the point of contact, blocking anastomosis. Compatibility is determined by a single, highly polymorphic locus called the fuhc with the following rules: individuals that share one or both fuhc alleles will fuse, while those who share neither will reject. The fuhc locus encodes multiple proteins with roles in allorecognition, including one called uncle fester, which is necessary and sufficient to initiate the rejection response. Here we report the existence of genotype-specific expression levels of uncle fester, differing by up to 8-fold at the mRNA-level, and that these expression levels are constant and maintained for the lifetime of an individual. We also found that these differences had functional consequences: the expression level of uncle fester correlated with the speed and severity of the rejection response. These findings support previous conclusions that uncle fester levels modulate the rejection response, and may be responsible for controlling the variation observed in the timing and intensity of the reaction. The maintenance of genotype specific expression of uncle fester is also evidence of an education process reminiscent of that which occurs in mammalian Natural Killer (NK) cells. In turn, this suggests that while histocompatibility receptors and ligands evolve via convergent evolution, they may utilize conserved intracellular machinery to interpret binding events at the cell surface.

immunology↗

bioRxiv2024

Identification of type 2 diabetes- and obesity-associated human β-cells using deep transfer learning

Diabetes affects >10% of adults worldwide and is caused by impaired production or response to insulin, resulting in chronic hyperglycemia. Pancreatic islet {beta}-cells are the sole source of endogenous insulin and our understanding of {beta}-cell dysfunction and death in type 2 diabetes (T2D) is incomplete. Single-cell RNA-seq data supports heterogeneity as an important factor in {beta}-cell function and survival. However, it is difficult to identify which {beta}-cell phenotypes are critical for T2D etiology and progression. Our goal was to prioritize specific disease-related {beta}-cell subpopulations to better understand T2D pathogenesis and identify relevant genes for targeted therapeutics. To address this, we applied a deep transfer learning tool, DEGAS, which maps disease associations onto single-cell RNA-seq data from bulk expression data. Independent runs of DEGAS using T2D or obesity status identified distinct {beta}-cell subpopulations. A singular cluster of T2D-associated {beta}-cells was identified; however, {beta}-cells with high obese-DEGAS scores contained two subpopulations derived largely from either non-diabetic or T2D donors. The obesity-associated non-diabetic cells were enriched for translation and unfolded protein response genes compared to T2D cells. We selected CDKN1C and DLK1 for validation by immunostaining in human pancreas sections from healthy and T2D donors. Both CDKN1C and DLK1 were heterogeneously expressed among {beta}-cells. CDKN1C was increased in {beta}-cells from T2D donors, in agreement with the DEGAS predictions, while DLK1 appeared depleted from T2D islets of some donors. In conclusion, DEGAS has the potential to advance our holistic understanding of the {beta}-cell transcriptomic phenotypes, including features that distinguish {beta}-cells in obese non-diabetic or lean T2D states. Future work will expand this approach to additional human islet omics datasets to reveal the complex multicellular interactions driving T2D.

bioinformatics↗

bioRxiv2023

Single-cell mechanical analysis reveals viscoelastic similarities between normal and neoplastic brain cells

Understanding cancer cell mechanics allows for the identification of novel disease mechanisms, diagnostic biomarkers, and targeted therapies. In this study, we utilized our previously established fluid shear stress assay to investigate and compare the viscoelastic properties of normal immortalized human astrocytes (IHAs) and invasive human glioblastoma (GBM) cells when subjected to physiological levels of shear stress that are present in the brain microenvironment. We used a parallel-flow microfluidic shear system and a camera-coupled optical microscope to expose single cells to fluid shear stress and monitor the resulting deformation in real-time, respectively. From the video-rate imaging, we fed cell deformation information from digital image correlation into a three-parameter generalized Maxwell model to quantify the nuclear and cytoplasmic viscoelastic properties of single cells. We further quantified actin cytoskeleton density and alignment in IHAs and GBM cells via immunofluorescence microscopy and image analysis techniques. Results from our study show that contrary to the behavior of many extracranial cells, normal and cancerous brain cells do not exhibit significant differences in their viscoelastic behavior. Moreover, we also found that the viscoelastic properties of the nucleus and cytoplasm as well as the actin cytoskeletal densities of both brain cell types are similar. Our work suggests that malignant GBM cells exhibit unique mechanical behaviors not seen in other cancer cell types. These results warrant future study to elucidate the distinct biophysical characteristics of the brain and reveal novel mechanical attributes of GBM and other primary brain tumors.

bioengineering↗

bioRxiv2023

Adapting to heat stress by sowing summer grain crops early in late winter: Sorghum root growth, water use, and yield

CONTEXTDrought and extreme heat at flowering are common stresses limiting the yield of summer crops, which are likely to intensify and become more frequent as projected under climate change. OBJECTIVEThis study explores the idea that adaptation to these stresses could be increased by sowing summer crops early in late winter or spring, to avoid the overlap with critical crop stages around flowering. Here we report on the impacts of early sowing i.e., in late winter and spring on sorghum crop and root growth and function (i.e., water use), and final grain yield. METHODSTwo seasons of on-farm genotype (G) by environment (E) by management (M) sorghum experimentation were conducted in the Darling Downs region of Queensland, Australia. Each trial consisted of a factorial combination of three times of sowing (TOS, referred to as late winter, spring, and summer), two levels of irrigation, four plant populations, and six commercial genotypes. Treatments were replicated three times. Crop roots and shoot were sampled at the flag leaf stage for each TOS. Crop water use across the growing season was monitored using time-lapse electromagnetic induction (EMI) surveys. EMI was also used to calculate a root activity factor. Final grain yield and yield components were determined at maturity. RESULTSResults showed that TOS, irrigation levels, and their interactions significantly influenced crop root and shoot traits, water use, and yield, though results were not always consistent across seasons. In the first season which was dry and had large temperature contrasts between TOS, crop growth in the early sown crops was primarily limited by temperature. In contrast, the second season was much warmer and crop growth was instead primarily limited by water availability. Cold air and soil temperatures in the early sowing dates i.e., late winter and spring during the first season, lead to smaller crops with smaller rooting systems and root-to-shoot ratios, and roots having a larger average root diameter. In general terms, root length and root length density responded positively to increasing pre-flowering mean air temperatures ranging between 16 and 20{degrees}C, while root average diameters were larger below 19 {degrees}C or above 21{degrees}C. Early sowing advanced flowering and therefore decreased the risk of extreme heat during the critical stages around flowering and affected water use before and after flowering. The root activity factor was directly related to the crop root length density. The early sown crops increased yield by transferring water use from vegetative to reproductive stages. The larger yield of the early sown crop was associated with larger grain numbers, particularly for the tillers, and a larger water use efficiency. As expected, irrigated and summer-sown crops exhibited lowest water use efficiency. The early-sown crops left more water in the soil profile at maturity, particularly under irrigated conditions and with small plant populations. CONCLUSIONSWe conclude that early sown sorghum is a potential option to increase crop adaptation to hotter and drier environments. Here we propose that in the race to increase crop adaptation to heat stresses, plant breeding efforts should consider cold tolerance traits during crop germination, emergence, and early vegetative stages so that sorghum sowing windows could be significantly advanced.

plant biology↗

bioRxiv2022

Microscopic Imaging of Epigenetic Age

Biomarkers of biological age that predict the risk of disease and expected lifespan better than chronological age are key to efficient and cost-effective healthcare1-3. To advance a personalized approach to healthcare, such biomarkers must perform on the individual rather than population level, demonstrate single cell resolution, and provide scalable and cost-effective measurements. We developed a novel approach - image-based chromatin and epigenetic age (ImAge), that utilizes image texture features based on the patterns of chromatin and epigenetic marks in single nuclei. We observed the emergence of intrinsic trajectories of ImAge using dimensionality reduction without regression on chronological age. ImAge was correlated with chronological age in all tissues and organs examined and was consistent with the expected acceleration and/or deceleration of biological age in chronologically identical mice treated with chemotherapy or following a caloric restriction regimen, respectively. ImAge from chronologically identical mice inversely correlated with their locomotor activity (greater activity for younger ImAge), consistent with the essential role of locomotion as an aging biomarker. Finally, we demonstrated that ImAge is reduced upon partial reprogramming in vivo following transient expression of OSKM cassette in the liver and skeletal muscles of old mice and validated the power of ImAge to assess the heterogeneity of reprogramming. We propose that ImAge represents the first-in-class individual-level biomarker of aging and rejuvenation with single-cell resolution.

systems biology↗

bioRxiv2022

Coinfection with chytrid genotypes drives divergent infection dynamics reflecting broad epidemiological patterns

By altering the abundance, diversity, and distribution of species -- and their pathogens -- globalization may inadvertently select for more virulent pathogens. In Brazils Atlantic Forest, a hotspot of amphibian biodiversity, the pet trade has facilitated the co-occurrence of previously isolated enzootic and panzootic lineages of the pathogenic amphibian-chytrid ( Bd) and generated new virulent recombinant genotypes ( hybrid). Epidemiological data indicate that amphibian declines are most severe in hybrid zones, suggesting that coinfections are causing more severe infections or selecting for higher virulence. We investigated how coinfections involving these genotypes shaped virulence and transmission. Overall, coinfection favored the more virulent and competitively superior panzootic genotype, despite dampening its virulence and transmission. However, for the least virulent and least competitive genotype, coinfection increased both pathogen virulence and transmission. Thus, by integrating experimental and epidemiological data, our results provide mechanistic insight into how globalization can select for, and propel, the emergence of introduced hypervirulent lineages, such as the globally distributed panzootic lineage of Bd.

ecology↗