Biology subjects

Jindal, S.

Publications and source records attributed to Jindal, S..

At least 19 recordsLinked to original sources

bioRxiv2026

Machine-assisted systematic review and meta-analysis of citrus flavonoids in streptozotocin-induced painful diabetic neuropathy

BackgroundNeuropathic pain is a common and debilitating condition with persistently poor translation from preclinical research to effective clinical therapies. Preclinical systematic reviews and meta-analyses (SRMAs) can provide more robust estimates of intervention effects and help identify sources of bias and heterogeneity, but their use is limited by the time and effort required to synthesise rapidly expanding literatures. Automation may help overcome these barriers. MethodsWe developed Neuropathic Pain - Systematic Online Living Evidence Summaries (NP-SOLES), a machine-assisted approach used in this study to support systematic identification of animal studies relevant to neuropathic pain. Using NP-SOLES, we conducted a pilot systematic review and meta-analysis of controlled animal studies assessing the effects of citrus flavonoids on thermal and mechanical pain-related outcomes in streptozotocin-induced models of painful diabetic neuropathy. ResultsIn this pilot application, machine-assisted evidence synthesis supported reduced manual screening burden, illustrating the potential of such approaches to facilitate timely preclinical SRMAs. Fourteen studies met inclusion criteria for meta-analysis on thermal pain-related outcomes. Pooled estimates suggested a reduction in thermal pain-related behaviours following flavonoid treatment; however, substantial heterogeneity and widespread methodological limitations were observed. Nine studies were eligible for qualitative analysis on mechanical pain-related outcomes, which showed an overall reduction in pain-related behaviours following treatment but had similar methodological limitations. Reporting of key measures to reduce risk of bias, including randomisation and blinding, were consistently poor, and subgroup analyses were constrained by sparse data. ConclusionsAvailable evidence from preclinical animal studies suggests a potential analgesic effect of citrus flavonoids on thermal and mechanical pain-related outcomes in diabetic neuropathy models, but confidence in this estimate is limited by significant risks of bias. More rigorously designed and transparently reported preclinical studies are required before clinical translation can be justified.

neuroscience↗

bioRxiv2026

SOX9 and SEMA7A regulate cell plasticity in the postpartum mammary gland with implications for breast cancer

Postpartum mammary gland involution is a coordinated process of cell death and remodeling that returns the tissue to a near pre-pregnant state following lactation and weaning. In models of postpartum breast cancer, defined as breast cancer diagnosed in women under age 45 and within 10 years of recent childbirth, involution induces durable phenotypes in breast tumor cells that promote progression and are associated with increased risk for therapeutic resistance, metastasis, and death in patients. SRY-Box Transcription Factor 9 (SOX9), a known regulator of mammary stem and progenitor cells, also promotes resistance to therapy and metastasis in breast cancers. Yet the contribution of SOX9 to the involution process is not well understood. We utilized single-cell RNA sequencing of mouse mammary glands during involution to delineate Sox9-expressing cell populations during lactation and involution. We found that Sox9 mRNA is primarily expressed in luminal progenitor cells that are largely absent during lactation and present during early involution. We also reveal that Sox9 is involved in a shift in cell state from lactational to non-lactational and is expressed in the surviving cells during involution. Prior work revealed that Semaphorin-7a (SEMA7A) also promotes cancer stem cell and pro-survival phenotypes in luminal progenitor cells during involution, and we observe a population of luminal progenitor cells that co-expresses Sox9 and Sema7a during involution. Mechanistically, we demonstrate that knockdown of Sox9 in cultured mammary epithelial cells results in increased SEMA7A expression, mesenchymal phenotypes, and loss of lactogenic differentiation capacity, identifying a potential regulatory axis where SOX9 balances SEMA7A expression in normal mammary epithelium and that disruption of this balance results in a dedifferentiated state that resembles mesenchymal cells. We validated a spatial relationship between SOX9 and SEMA7A proteins in a unique set of breast tissue samples from healthy human donors to show co-expression during early involution. In breast cancer datasets, we observe elevated expression of SOX9 and SEMA7A in triple-negative breast cancers, as well as in the mesenchymal subtype of triple-negative breast cancers, suggesting disruption of this regulatory axis in breast cancer. Finally, we observe that co-expression increases metastatic risk in both estrogen receptor-negative and -positive breast cancers. Collectively, these findings define a novel SOX9-SEMA7A relationship in healthy mammary tissues and illustrate how studies of normal progenitor cell phenotypes can delineate cellular mechanisms that contribute to breast tumor progression.

developmental biology↗

bioRxiv2026

Spatial immune hubs defined by conserved activated dendritic cells are remodeled by immunotherapy

Dendritic cells (DCs) orchestrate anti-tumor immune responses, yet the full extent of their phenotypic diversity, and spatial dynamics within the tumor microenvironment (TME) remains incompletely understood. Here, we constructed an integrated atlas of tumor-infiltrating DCs by harmonizing single-cell transcriptomic data from 12 murine tumor studies and 28 published human cancer datasets, together with newly generated single-cell-resolved multiplexed tissue imaging across immunotherapy conditions in a murine model. We noted conserved transcriptional states across species, including canonical conventional type 1 DCs (cDC1s), diverse type 2 DC (cDC2) subpopulations, and two activation states characterized by CCR7 expression (CCR7+ DCs) or interferon-stimulated gene expression (ISG DCs). Spatial transcriptomics analyses from human TMEs revealed that CCR7+ DCs and ISG DCs reside in distinct T cell-enriched regions that are embedded within distinct signaling environments. High-dimensional multiplexed proteomic imaging demonstrated that these DC-T cell niches undergo divergent remodeling across multiple immunotherapy conditions. Notably, this spatial reorganization occurred despite minimal detectable changes in DC transcriptional states. This study delineates conserved DC activation states and their spatial organization within tumors and captures the therapy-dependent remodeling, providing a framework for studying therapy-associated remodeling of DC immune programs in cancer.

immunology↗

bioRxiv2025

Macrophage-Dendritic Cell-T-Cell Tetrads Orchestrate Antitumor Immunity and Response to Checkpoint Blockade

Immune checkpoint inhibitors (ICIs) elicit durable responses in only a subset of patients with solid tumors, underscoring the need to define the cellular architectures that govern effective antitumor immunity. Here we identify a spatially organized multicellular immune unit comprising macrophages, cDC1s, CD4 T-cells, and CD8 T-cells that emerges in response to anti-CTLA-4 or dual checkpoint blockade. We term these structures tetrads. Using multiplexed imaging and spatial transcriptomics in mouse and human tumors, we show that tetrads assemble early during immune priming, depend on the ICOS-ICOSL pathway, and are enriched for ICOS Th1-like CD4 T cells and ICOSLhigh cDC1s. CD8 T-cells within tetrads exhibit an activated, non-terminally differentiated state, while tetrad-associated macrophages display an interferon-{gamma}-responsive program that sustains CD8 T-cell function and prevents dysfunction. Functionally, ICOSL cDC1s are required for tumor eradication in vivo. In patients with bladder cancer treated with neoadjuvant dual checkpoint blockade, tetrad, but not triad or dyad formation correlates with clinical response. These findings establish tetrads as a fundamental cellular unit coordinating antitumor immunity and responsiveness to ICIs.

immunology↗

bioRxiv2025

Coordinated action of CRK2 and QSK1 regulate osmotic stress response in Arabidopsis

Precise control of intercellular communication is essential for normal growth and stress responses in all multicellular organisms. In Arabidopsis, two membrane-localized receptor like kinases (RLKs), the Cysteine-rich RLK CRK2 and the Leucine-rich repeat (LRR) RLK QSK1 relocalize from the general plasma membrane (PM) to plasmodesmata (PD) in response to osmotic stress. Both these RLKs regulate callose deposition thereby modulating PD permeability. However, unchecked callose deposition can block the PD and disrupt proper intercellular communication. Here, we show that under normal growth conditions, CRK2 phosphorylates and sequesters QSK1 at the general PM, preventing unnecessary callose deposition at PD. We show that osmotic stress-induced enrichment of QSK1 at PD requires functional CRK2 and establish that phosphorylation of QSK1 in its C-terminal region is inhibitory in this process. We propose that osmotic stress triggers dephosphorylation and release of QSK1 from the CRK2-QSK1 complex, enabling its relocalization from general PM to PD, where it promotes stress-induced callose deposition. Subsequently, CRK2 relocalizes to PD where it negatively influences callose deposition. Our work reveals a tightly coordinated distribution of QSK1 and CRK2 at PM, establishing a dynamic gating mechanism that balances growth and stress responsiveness.

plant biology↗

bioRxiv2025

CYSTEINE-RICH RLK2 regulates development via callose synthase-dependent symplastic transport in Arabidopsis

CYSTEINE-RICH RECEPTOR-LIKE PROTEIN KINASEs (CRKs) play an important role in plant development and stress responses. One of the best described members of the Arabidopsis CRK family is CRK2, which was proposed as a crucial regulator of intercellular transport facilitated by plasmodesmata (PD). As intercellular channels allowing symplastic communication, PD-mediated transport is predominantly regulated by callose synthase (CALS)-mediated callose deposition. This process can impact not just the distribution of molecules between adjacent cells, but also the symplastic loading of vascular tissue, thereby influencing plant stress responses and developmental processes. Here we described the overlapping expression pattern of genes encoding phylogenetically closely related CALS1 and CALS3. Both CALSs were phosphorylated in vitro by CRK2, and the genetic interaction between genes encoding CRK2 and CALS1 or CALS3 revealed their impact on callose deposition, rosette growth, primary root length, and development, represented as a decreased number of true leaves. Importantly, we observed significant accumulation of starch in crk2 mutant plants, especially in developmentally older leaves, which was reverted by the independent introduction of cals1.5 and cals3.1 into the crk2 mutant background. The observed starch accumulation was accompanied by photosynthesis inhibition. We propose that the growth and developmental alterations of crk2 are caused by decreased phloem loading, which resulted in starch accumulation in source organs, and subsequent sink tissue starvation. Our results propose CRK2 as negative regulator of CALS1 and CALS3 regulating source to sink transport, which impacts plant growth and development.

plant biology↗

bioRxiv2025

Spatial multi-omics and deep learning reveal fingerprints of immunotherapy response and resistance in hepatocellular carcinoma

Despite advances in immunotherapy treatment, nonresponse rates remain high, and mechanisms of resistance to checkpoint inhibition remain unclear. To address this gap, we performed spatial transcriptomic and proteomic profiling on human hepatocellular carcinoma tissues collected before and after immunotherapy. We developed an interpretable, multimodal deep learning framework to extract key cellular and molecular signatures from these data. Our graph neural network approach based on spatial proteomic inputs achieved outstanding performance (ROC-AUC > 0.9) in predicting patient treatment response. Key predictive features and associated spatial transcriptomic profiles revealed the multi-omic landscape of immunotherapy response and resistance. One such feature was an interface niche expressing restrictive extracellular matrix factors that physically separates tumor tissue and lymphoid aggregates in nonresponders. We integrate this and other spatially-resolved signatures into SPARC, a multi-omic "fingerprint" comprising scores for immunotherapy response and resistance mechanisms. This study lays groundwork for future patient stratification and treatment strategies in cancer immunotherapy.

cancer biology↗

bioRxiv2024

Light quantity impacts early response to cold and cold acclimation in young leaves of Arabidopsis

Plant reactions to stress vary with development stage and fitness. This study assessed the relationship between light and chilling stress in Arabidopsis acclimation. By analyzing the transcriptome and proteome responses of expanding leaves subjected to varying light intensity and cold, 2251 and 2064 early response genes and proteins were identified, respectively. Many of these represent as yet unknown part of early response to cold, illustrating development-dependent response to stress and a duality in plant adaptations. While standard light promoted photosynthetic upregulation, plastid maintenance, and increased resilience, low light triggered a unique metabolic shift, prioritizing ribosome biogenesis and lipid metabolism and attenuating expression of genes associated with plant immunity. The comparison of early response in young leaves with that in expanded ones showed striking differences, suggesting a sacrifice of expanded leaves to support young ones. Validations of selected DEGs in mutant background confirmed a role of HSP90-1, transcription factor FLZ13, and Phospholipase A1 (PLIP) in response to cold, and the PLIP family emerged as crucial in promoting acclimation and freezing stress tolerance. The findings highlight the dynamic mechanisms that enable plants to adapt to challenging environments and pave the way for the development of genetically modified crops with enhanced freezing tolerance.

plant biology↗

bioRxiv2024

A TORC1-PHR1 signaling axis regulates phosphorus starvation and immunity signaling network in Arabidopsis

The Target of Rapamycin Complex 1 (TORC1) is a crucial eukaryotic kinase that modulates growth in response to nutrient availability. Phosphorus (P) is an essential macronutrient, and its deficiency induces extensive reprogramming of growth and defense strategies in plants. This process involves Phosphate Starvation Response 1 (PHR1), a master regulator of the Phosphate Starvation Response (PSR). In this study, we identify a novel, non-canonical role for TORC1 in regulating P starvation responses in Arabidopsis. We demonstrate that P limitation activates TORC1, leading to the stabilization of PHR1. Inhibition of TORC1 increased sensitivity to P starvation, accompanied by disruption of starvation-induced transcriptional reprogramming. Additionally, our results reveal that the TORC1-PHR1 signaling axis plays a crucial role in reprogramming the expression of genes involved in the plant immune signaling network. This regulation is critical for the symbiotic association with the endophytic fungus Piriformospora indica under P starvation. These findings underscore the significant role of the TORC1-PHR1 module in orchestrating the PSR and highlight the evolutionary adaptation of TORC1 signaling pathways in plants.

plant biology↗

bioRxiv2024

Spatial Transcriptomics Reveals Inflammation and Trans-differentiation States of Acute Myeloid Leukemia in Extramedullary and Medullary Tissues

Acute myeloid leukemia (AML) is a heterogeneous disease of the bone marrow (medullary) but can also involve extramedullary tissues. While single cell dynamics of AML in suspension are previously explored, a comprehensive spatial transcriptomic assessment in AML remain underexplored. Here, we used Visium spatial transcriptomics to resolve medullary and extramedullary AML environments. We reveal spatial co-localization of monocytes and granulocyte-monocyte progenitors with leukemic populations in the bone marrow, sharing molecular signatures with extramedullary sites. Cell-cell communication via the CXCL12- CXCR4 axis correlated with PI3K/AKT/mTOR signaling in high inflammatory niches. Trans- differentiation states were concentrated in AML-infiltrated regions, with committed-like AML populations present in inflammatory niches and away from the trabeculae, while primitive-like AML cells localized near the endosteal niche. We validated these findings in GeoMx-based Digital Spatial profiling (DSP). Our study applied multimodal spatial transcriptomic approaches to characterize the spatial hierarchy and microenvironmental dynamics of AML differentiation states. We also demonstrated the feasibility of applying Visium-based spatial transcriptomics in decalcified bone tissues.

cancer biology↗

bioRxiv2022

Myeloid-specific KDM6B inhibition sensitizes Glioblastoma to PD1 blockade

Glioblastoma (GBM) tumors are enriched in immune-suppressive myeloid cells and are refractory to immune checkpoint therapy (ICT). Targeting epigenetic pathways to reprogram the functional phenotype of immune-suppressive myeloid cells to overcome resistance to ICT remains unexplored. Single-cell and spatial transcriptomic analyses of human GBM tumors demonstrated high expression of an epigenetic enzyme - histone 3 lysine 27 demethylase (KDM6B) in intra- tumoral immune-suppressive myeloid cell subsets. Importantly, myeloid-cell specific Kdm6b deletion enhanced pro-inflammatory pathways and improved survival in GBM tumor-bearing mice. Mechanistic studies elucidated that the absence of Kdm6b enhances antigen-presentation, interferon response and phagocytosis in myeloid cells by inhibiting mediators of immune suppression including Mafb, Socs3 and Sirpa. Further, pharmacological inhibition of KDM6B mirrored the functional phenotype of Kdm6b deleted myeloid cells and enhanced anti-PD1 efficacy. Thus, this study identified KDM6B as an epigenetic regulator of the functional phenotype of myeloid cell subsets and a potential therapeutic target to improve response to ICT.

immunology↗

bioRxiv2022

Stepwise origin and evolution of a transcriptional activator and repressor system integrating nutrient signaling in plants

Plants possess a unique transcriptional regulatory system in which two related MYB-related transcription factors (TFs) coordinate gene expression according to phosphate (Pi) and nitrogen (N) availability. The Phosphorus Starvation Response (PSR) type TFs are transcriptional activators integrating the cellular Pi sensing machinery and gene regulation majorly under Pi starvation. The Hypersensitivity To Low Pi-Elicited Primary Root Shortening (HRS) type TFs are transcriptional repressors integrating the Pi and N availability signals through different feedback loops. They are highly connected through multiple signaling loops to finetune the transcriptional responses according to nutrient availability. Molecular functions of these TFs are fairly uncovered in model systems; however, how plants evolved this activator-repressor system is currently unknown. In this study, using sensitive evolutionary analysis, we identified a stepwise origin of the PSR-HRS regulatory system in plants. The PSR TFs were originated before the split of Prasinodermophyta and Chlorophyta. The HRS TFs were originated later in the Streptrophycean algae. We also identified the asymmetric expansion of this TF repertoire in land plants majorly shaped by genome duplication and triplication events. The phylogenetic reconstruction coupled with motif analysis revealed that the origin of the specific accessory motifs is a major contributing factor in the functional divergence which led to the evolution of different sub-families preceding the angiosperm radiation. The spatiotemporal gene expression analysis in different developmental stages and nutrient availability conditions in angiosperms identified a critical role of expression divergence in shaping the functions of these TF families which is essential for adaptive plasticity of plants.

plant biology↗

bioRxiv2021

Preventing ovariectomy-induced weight gain decreases tumor burden in rodent models of obesity and postmenopausal breast cancer.

Obesity and adult weight gain are linked to increased breast cancer risk and poorer clinical outcomes in postmenopausal women, particularly for hormone-dependent tumors. Menopause is a time when significant weight gain occurs in many women, and clinical and preclinical studies have identified menopause (or ovariectomy) as a period of vulnerability for breast cancer development and promotion. We hypothesized that preventing weight gain after ovariectomy (OVX) may be sufficient to prevent the formation of new tumors and decrease growth of existing mammary tumors. Here, we tested this hypothesis in a rat model of obesity and carcinogen-induced postmenopausal mammary cancer and validated our findings in a murine xenograft model with implanted human tumors. In both models, preventing weight gain after OVX significantly decreased obesity-associated tumor development and growth. Importantly, we did not induce weight loss in these animals, but simply prevented weight gain. In both lean and obese rats, preventing weight gain reduced visceral fat accumulation and associated insulin resistance. Similarly, the intervention decreased circulating tumor-promoting growth factors and inflammatory cytokines (ie, BNDF, TNF, FGF2), with greater effects in obese compared to lean rats. In obese rats, preventing weight gain decreased adipocyte size, adipose tissue macrophage infiltration, reduced expression of the tumor-promoting growth factor FGF-1, and reduced phosphorylated FGFR in tumors. Together, these findings suggest that the underlying mechanisms associated with the anti-tumor effects of weight maintenance are multifactorial, and that weight maintenance during the peri-/post-menopausal period may be a viable strategy for reducing obesity-associated breast cancer risk and progression in women.

cancer biology↗

bioRxiv2021

Mutant strains of Escherichia coli lacking global regulators, arcA and fis, demonstrate better growth fitness by pathway reprogramming under acetate metabolism

Global regulatory transcription factors play a significant role in controlling microbial metabolism under genetic and environmental perturbations. A systems-level effect of carbon sources such as acetate on microbial metabolism under disrupted global regulators has not been well established. Acetate is one of the substrates available in a range of nutrient niches such as the mammalian gut and high-fat diet. Therefore, investigating the study on acetate metabolism is highly significant. It is well known that the global regulators arcA and fis regulate acetate uptake genes in E. coli under glucose condition. In this study, we deciphered the growth and flux distribution of E.coli transcription regulatory knockout mutants {Delta}arcA, {Delta}fis and double deletion mutant, {Delta}arcAfis under acetate using 13C-Metabolic Flux Analysis which has not been investigated before. We observed that the mutants exhibited an expeditious growth rate (~1.2-1.6 fold) with a proportionate increase in acetate uptake rates compared to the wild-type. 13C-MFA displayed the distinct metabolic reprogramming of intracellular fluxes, which conferred an advantage of faster growth with better carbon usage in all the mutants. Under acetate metabolism, the mutants exhibited higher fluxes in the TCA cycle (~18-90%) and lower gluconeogenesis flux (~15-35%) with the proportional increase in growth rate. This study reveals a novel insight by stating the sub-optimality of the wild-type strain grown under acetate substrate aerobically. These mutant strains efficiently oxidize acetate to acetyl-CoA and therefore are potential candidates that can serve as a precursor for the biosynthesis of isoprenoids, biofuels, vitamins and various pharmaceutical products. ImportanceUnravelling the role of global regulatory genes on microbial metabolism of substrates available in various growth niche is important. Studies have shown that the global transcriptional regulators arcA and fis, under glucose availability, suppress the acetate uptake genes indicating a link between nutrient source and gene regulatory control. This work is focused on deciphering the influence of these regulators on acetate metabolism in E.coli. Growth studies using knockout strains ({Delta}arcA, {Delta}fis and {Delta}arcAfis) and 13C Metabolic flux analysis defined precise metabolic phenotypes under acetate metabolism. Interestingly, the mutants showed metabolic readjustment to facilitate optimal biomass requirements and a better balance between energy and precursor synthesis, resulting in better growth, which lacked in the wild-type strain. The outcomes of this study will be leveraged in understanding the regulatory control under various nutrient shifts.

microbiology↗

bioRxiv2021

Oleate metabolism using kinetic 13C dilution strategy deciphered the potential role of global transcription regulator arcA in Escherichia coli

Microbial metabolism of long-chain fatty acids (LCFA; > C12) is of relevance owing to their presence in various nutrient niches. Microbes have evolved to metabolize LCFA by expressing relevant genes coordinated by various transcriptional regulators. Among the global transcriptional regulators, the metabolic control conferred by arcA (aerobic respiration control) under a LCFA medium is lacking. This work is targeted to unravel the metabolic features of E.coli MG1655 and its knockout strain {Delta}arcA under oleate (C18:1) as a sole carbon source, providing novel insights into the flexibility of the global regulators in maintaining the cellular physiology. Owing to the availability and cost of stable isotope LCFA tracers, we adopted a novel kinetic 13C dilution strategy. This allowed us to quantify the 13C dilution rates in the amino acids that retro-biosynthetically shed light on the central metabolic pathways in actively growing cells. Our data comprehensively mapped oleate oxidization in E.coli via the pathways of {beta}-oxidation, TCA cycle, anaplerotic and gluconeogenesis. Interestingly, arcA knockout showed expeditious growth (~60%) along with an increased oleate utilization rate (~55%) relative to the wild-type. {Delta}arcA also exhibited higher 13C dilution rates (> 20%) in proteinogenic amino acids than the wild-type. Overall, the study established the de-repression effect conferred by {Delta}arcA in E.coli, which resulted in a phenotype with reprogrammed metabolism favouring higher oleate assimilation. The outcomes suggest rational metabolic engineering of regulators as a strategy to develop smart cells for enhanced biotransformation of LCFA. This study also opens an avenue for adopting a kinetic 13C dilution strategy to decipher the cellular metabolism of a plethora of substrates, including other LCFA in microbes.

systems biology↗

bioRxiv2020

A negative feedback loop of the TOR signaling moderates growth and enables rapid sensing of stress signals in plants

TOR kinase is a central coordinator of nutrient-dependent growth in eukaryotes. Maintaining optimal TOR signaling is critical for the normal development of organisms. However, the mechanisms involved in the maintenance of optimal TOR signaling are currently unknown in plants. In this study, we describe a negative feedback loop of TOR signaling helping in the adaptability of plants in changing environmental conditions. Using an interdisciplinary approach, we identified a plant-specific zinc finger protein FLZ8, as a regulator of TOR signaling in Arabidopsis. In sugar sufficiency, FLZ8 is upregulated by TOR-dependent and -independent histone modifications. FLZ8 negatively regulates TOR signaling by promoting antagonistic SnRK11 signaling and bridging the interaction of SnRK11 with RAPTOR, a crucial accessory protein of TOR. This negative feedback loop moderates the TOR-growth signaling axis in the favorable condition and helps in the rapid activation of stress signaling in unfavorable conditions establishing its importance in the adaptability of plants.

plant biology↗

bioRxiv2020

A palette of fluorophores that are differentially accumulated by wild-type and mutant strains of Escherichia coli: surrogate ligands for bacterial membrane transporters

Our previous work had demonstrated that two commonly used fluorescent dyes that were accumulated by wild-type E. coli MG1655 were accumulated differentially in single-gene knockout strains, and also that they might be used as surrogates in flow cytometric transporter assays. We summarise the desirable properties of such stains, and here survey 143 candidate dyes. We triage them eventually (on the basis of signal, accumulation levels, and cost) to a palette of 39 commercially available and affordable fluorophores that are accumulated significantly by wild-type cells of the Keio strain BW25113, as measured flow cytometrically. Cheminformatic analyses indicate both their similarities and their (much more considerable) structural differences. We describe the effects of pH and of the efflux pump inhibitor chlorpromazine on the accumulation. Even the wild-type MG1655 and BW25113 strains can differ significantly in their ability to take up such dyes. We illustrate the highly differential uptake of our dyes into strains with particular lesions in, or overexpressed levels of, three particular transporters or transporter components (yhjV, yihN, and tolC). The relatively small collection of dyes described offers a rapid, inexpensive, convenient and valuable approach to the assessment of microbial physiology and transporter function.

microbiology↗

bioRxiv2020

S-nitrosylated and non-nitrosylated COX2 has differential expression and distinct subcellular localization in normal and breast cancer tissue

Immunohistochemical staining in breast cancer shows both gain and loss of COX2 expression with disease risk and progression. We investigated four common COX2 antibody clones and found high specificity for purified human COX2 for three clones; however, recognition of COX2 in cell lysates was clone dependent. Biochemical characterization revealed two distinct forms of COX2, with SP21 recognizing an S-nitrosylated form and CX229 and CX294 appearing to recognize the same non-nitrosylated COX2 antigen. We found S-nitrosylated and non-nitrosylated COX2 occupy different subcellular locations in normal and breast cancer tissue, implicating distinct synthetic/trafficking pathways and function. Dual stains of ~2000 breast cancer cases show early onset breast cancer has increased expression of both forms of COX2 compared to postmenopausal cases. Our results highlight the strengths of using multiple, highly characterized antibody clones for COX2 immunohistochemical studies and raise the prospect that S-nitrosylation of COX2 may play a role in breast cancer biology.

cancer biology↗